Резултати </ti= tle> <o:SmartTagType namespaceuri=3D"urn:schemas-microsoft-com:office:smarttags" name=3D"metricconverter"/>  <style>  </style>  </head> <body lang=3DBG link=3Dblue vlink=3Dpurple style=3D'tab-interval:36.0pt'> <div class=3DSection1> JOURNAL OF CULTURE COLLECTIONS<o:p></o:p> Volume 4, 2004-2005, pp. </span= >68-77<o:p></o:p></= i> <o:p> </o:p> <o:p> </o:p> <o:p> </o:p> Vaccine strain of tomato mosaic virus obtained after serial passages of attenuated strain of tobacco mosaic virus in tolerant to tobamoviruses tomato<o:p></o:p> <= o:p> </o:p> E= lisaveta Stoimenova1* and Angela Yordanova2<o:p></o:p><= /p> <o:p> </o:p> 1Institute of Genetics, Bulgarian Aca= demy of Sciences, 1113 Sofia, Bulgaria</spa= n>;<o:p></o:p></= span> 2<s= pan lang=3DEN-US style=3D'mso-bidi-font-size:12.0pt;font-family:"Times New Roma= n"; letter-spacing:-.1pt;mso-ansi-language:EN-US;font-weight:normal'>National B= ank for Industrial Microorganisms and Cell Cultures, 1113 Sofia, P.O.Box 239, Bulgaria<o:p></o:p> <o:p> </o:p> <o:p> </o:p> Summary<o:p></o:p></= span> <o:p> </o:p></s= pan> An attenuated strain of tobacco mosaic virus= </spa= n>(TMV<= span style=3D'mso-bidi-font-size:12.0pt;font-family:"Times New Roman";color:blac= k; font-weight:normal'>) <s= pan lang=3DEN-US style=3D'mso-bidi-font-size:12.0pt;font-family:"Times New Roma= n"; color:black;mso-ansi-language:EN-US;font-weight:normal'>isolated from tobac= co and symptopmless for tomato and pepper was put in fivefold passage in toler= ant to tobamoviruses tomato. The changes in the strain population after the pas= sages were studied by determination of the virion electrophoretic mobility, serological and biological properties of purified preparations of the obtai= ned isolates. All performed experiments pro= ved that as early as in the first passage TMV was replaced by a tomato mosaic virus (<s= pan lang=3DEN-US style=3D'mso-bidi-font-size:12.0pt;font-family:"Times New Roma= n"; color:black;mso-ansi-language:EN-US;font-weight:normal'>ToMV).= After five serial passages the presence of virions cau= sing mosaic in susceptible and/or tolerant tomato were not found. The study for vaccine properties of the new obtained isolate ToMV D52-V showed that it belonged to Pelham’s group “<st1:metricconverter ProductID=3D"1= ”" w:st=3D"on">1”</st1:metricconverter> since it was effective in tomato carrying Tm-1 gene. Its protective action did not give way to that of analo= gous vaccines applied in the practice and hence strain D52-V could be successful= ly used for protection of susceptible and tolerant tomato against pathogenic tobamovirus strains. <sp= an style=3D'mso-bidi-font-size:12.0pt;font-family:"Times New Roman";color:blac= k; font-weight:normal'><o:p></o:p> <o:p> </o:p> Introduction<o:p></o= :p> <o:p> </o:p> In the 70s and 80s of the twent= ieth century vaccine strains of tomato mosaic virus (ToMV) have been widely used= for tomato protection against pathogenic tobamovirus strains. The application o= f this method in the practice proved its effectiveness however, some problems aros= e. The main objections were related to the presence of a little of pathogenic particles (up to 1 %) in the vaccine preparations and to the risk of appearance of new highly virulent virus strains. Apprehensions were stated about the vaccine strains translation upon other crops, for which these str= ains were pathogenic [7, 24]. The method application in tomato was restrict= ed as consequence of the new cultivars creation - tolerant (Tm-1) or resistant= to tobamoviruses ones (Tm-2 and Tm-22). However, the broad use of t= hese cultivars led to raising of ToMV strains from group “1”, “2” and “1.2” according to Pelham’s classification [14], overcoming the respective resistance of the Tm-1, Tm-2= and Tm-1+Tm-2 genes [14, 15, 16, 20]. In tolerant to tobamovirus= es tomato some “0” strains are able to cause symptomless systemic infection while “1” strains induce mosaic symptoms. Cultivars carrying Tm-1 gene are spread in our country and are used in the production= of middle early and late tomato [20]. <o:p></o:p> In Bulgaria, ToMV strains from groups “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0R= 21;</st1:metricconverter> and “<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”= </st1:metricconverter> were found in yield tomato but strains of tobacco mosaic virus (TMV) were n= ot isolated. The latter was occasionally established in green houses. However,= in pepper TMV strains were frequently occurred besides ToMV ones and could even predominate [20]. The actual lack of TMV strains in tomato could be explain= ed on one hand with the fact that TMV strains easily mutated in ToMV, as found= in many investigations and on the other hand ToMV had a faster rate of multiplication or a greater invasiveness in tomato plants compared with TMV [26]. In the nature when TMV was found in tomato, it was mixed with ToMV [1= 6, 20]. <o:p></o:p> In the present work the obtaini= ng of a new vaccine ToMV strain is described which could be used for protectio= n of tomato with Tm-1 gene against pathogenic virus strains from group “<s= t1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter>. <o:p></o= :p> <o:p> </o:p> Materials and Method= s<o:p></o:p> <o:p> </o:p> Virus strains. </spa= n>Objects of the study we= re TMV strain D52 - symptomless for tomato and pepper and mild for tobacco as well= as its derivative isolates obtained after passages in tolerant tomato.<o:p></o= :p> Twenty one TMV and ToMV strains were included in the investigation. The ToMV vaccine strains were: t= he Bulgarian B-5 from group “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> after Pelham (National Bank for Industrial microorganisms and Cell Cultures 2024) and the Russian V-69 from group “<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1̶= 1;</st1:metricconverter> (NBIMCC 2114) [21, 24]. The used pathogenic strains were: ТМV -= U1 (3405), TGM (3294), B (2022), МА (2077) and Sm (2125), the latt= er being from group “<st1:metricconverter ProductID=3D"1”" w:st=3D= "on">1”</st1:metricconverter>; ТоМV strains from group “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> – E= NP (2074), SD (2076), Lat-0 (2064), GdK (2075), GM-0 (2078), YM-0 (2348), No29 (3297), ToGM (3296); ТоМV strains from group “<st1:= metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> – N= H-69, SPS, GeBl (2112), So69 (2111), GM-1 (2349) and YM-1 (2348). The plants from which TMV and ToMV “<st1:metricconverter ProductID=3D"0”" w:st= =3D"on">0”</st1:metricconverter> strains have been isolated are shown in Table 2. The strains from group = 220;<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> originate= from tomato. Strains TGM, B, GM-0, No29, ToGM, YM-0, GM-1 and YM-1 are from Bulg= aria [19]. The authors express much gratitude to Prof. Dr. A. Th. B Ra= st, Prof. Dr. H. Laterrot and Prof. Dr. F. Garcia-Arenal for the supplying of other tobamoviruses.<o:p></o:p> Test plants.<= /b> The following cultivars were used: tomato cv. Ideal susceptible to tobamoviruses and cv. Mobaci carrying Tm-1 tolerance gene to this virus gro= up; tobacco cv. Samsun N’N’ (N’ gene from Nicotiana sylvestris) and cv. Sams= un NN (N gene from N. glutinosa, localizing the tobamoviruses). The plants were grown in autoclaved soil and sterile metal pots. Seeds obtained from plants proved as virusless were use= d.<o:p></o:p> Virus propagation and purification. Tobacco plants cv. Samsun N’N’ in 5-6 leaf stage were inoculated with TMV strains. The multiplication of ToMV “<st1:metricconverter ProductID=3D"0”" w= :st=3D"on">0”</st1:metricconverter> strains was done in tomato cv. Ideal while strains from group “<st1:m= etricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> were prop= agated in tomato cv. Mobaci and plants in cotyledon-first real leaf stage were infected. The mosaic leaves were collected 21 days past inoculation (dpi) w= ith pathogenic strains. The plants inoculated with vaccine strains remained symptomless and all the leaves were harvested. Purified virus preparations = were obtained by the method of Gooding and Hebert [5]. The virus concentration w= as spectrophotometrically determined. Tobacco cv. Samsun NN was used for production of single lesion isolates. <o:p></o:p> Passages in tolerant tomato. Tomato cv. Mobaci was inoculated with Тi= 2;V D52 preparation (0.1 mg/ml), the plants were grown 45 days and the symptoms and the systemic virus distribution were recorded. All plants were symptomless and a purified preparation named D52-I was made from them. Toma= to plants were again mass inoculated with this isolate and they remained symptomless. A preparation D52-II was similarly obtained. The procedure was multifold repeated and isolates D52-III, D52-IV and D52-V were purified. 40 plants were inoculated at every passage.<o:p></o:p> As in these experiments it was exceptionally important to exclude the contamination, besides the mentioned above measures all pots were enclosed = with polyethylene. In this way the touch was avoided between the inoculated with different viruses plants, as well as the contact during watering.<o:p></o:p= > Electrophor= etic mobility of virus particles. The electrophoretic mobility (Rf) of tobamovirus virions was determined in 0.7 % agarose horizontal gel, in <st1:metricconve= rter ProductID=3D"40 mM" w:st=3D"on">40 mM</st1:metricconverter> Tris-acetate bu= ffer, pH 7.8, containing <st1:metricconverter ProductID=3D"2 mM" w:st=3D"on">2 mM</s= t1:metricconverter> EDTA. The virus preparations were applied in concentration of 10 mg/ml. The electrophoresis was carried out at 60 V for 2.5 hours. The gels were stained with 0.2 % amido black in <st1:metricconverter ProductID=3D"0.1 M" w:st=3D"= on">0.1 M</st1:metricconverter> sodium acetate, containing 10 % glycerol [4]. The = electrophoregrams were dried and stained additionally with Coomassie Brilliant Blue R-250 [27= ].<o:p></o:p> The Rf of the samples was accounted according to the equation Rf =3D= S/F, where F was the distance from the start to the front and S was the sample distance. This value was calculated to Rf of the type strain U1 accepted as 100 %. The standard deviation of three repetitions was determined. <o:= p></o:p> <h1 style=3D'text-indent:1.0cm;line-height:normal;mso-pagination:none'>Immunoelectrophoretic analysis. Counter immunoelectrophoresis was performed in 0.9% agarose gel in <st1:metricconve= rter ProductID=3D"0.075 M" w:st=3D"on">0.075 M</st1:metricconverter> veronal buf= fer, pH 8.6, at 150 V, 10 oC for two hours. The virus preparations = were applied in concentration of 5 mg/ml (25 <span lang=3DEN-US style=3D'font-family:Symbol;mso-ascii-font-family:"Times New Roman";mso-han= si-font-family: "Times New Roman";mso-char-type:symbol;mso-symbol-font-family:Symbol; font-weight:normal;mso-bidi-font-weight:bold'>mg per start). The antiserum against ToMV in dilution of 1:4 was used. The gels were stained w= ith Coomassie Brilliant Blue R-250 [27].<o:p></o:p></h1> Biological properties. Ten plants of each toma= to cvs Ideal, Mobaci and tobacco cv. Samsun N’N’ were inoculated with purified preparations of strain D52 and isolates D52-I, D52-II, D52-III, D5= 2-IV and D52-V. The symptoms on the inoculated leaves were recorded on the 7<sup= >th and 10th dpi and on the young leaves on the 40th day.= The tobamovirus presence was proved by DAS-ELISA.<o:p></o:p> Vaccine properties. Two experiments for comparative testing of D52 and D52-V vaccine properties were carried out. In the first experiment, tomato plants cv. Ideal were vaccinated with B-5, V-69, D52 and D52-V and after 7 days they were challenge inoculated with TMV and ToMV pathogenic str= ains from group “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0= ”</st1:metricconverter>. In the second case, tomato plants cv. Mobaci were vaccinated with B-5, V-69 and D52-V and after 7 days they were infected with group “<= st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> pathogenic strains. The following variants were used as controls: untreated plants (healthy control); plants inoculated only with the respective vaccine strain and plants infected only with the respective pathogenic strain. The number = of mosaic plants was accounted 30 days after the mosaic symptoms appearance in= the plants infected only with pathogenic strains. 40 plants were used in each variant.<o:p></o:p> <o:p> </o:p> Results and Discussion<o:p></o:p= > <o:p> </o:p> TMV strain No. 52 was isol= ated when the tobamovirus strain variability in commercial tobacco in Bulgaria w= as studied. This strain does not differ in electrophoretic mobility and serological properties from the typical TMV strains but it induces mild gre= en mosaic in tomato, pepper and tobacco [17, 19]. A significant percentage (12 %) symptomless plants was found after single lesion infection of tomato (Tm+) with No. 52. After a purposive selection a TMV strain D52= was obtained that was symptomless for tomato and pepper and mild for tobacco. In spite of the mult= ifold passages of strain D52 in tomato (Tm+), ToMV virions were not established in its preparations (testing on tobacco cv. Samsun N’N’).<o:p></o:= p> <o:p> </o:p> Passages and biological properti= es of the obtained tobamovirus isolates<o:p></o:p> <o:p> </o:p> Five consecutive passages of st= rain D52 in tolerant to tobamoviruses tomato (Tm-1) was performed. The initial infection was made with a preparation obtained from tobacco cv. Samsum N’N’ in order to exclude the opportunity of ToMV particles pres= ence in the started inoculum. <o:p= ></o:p> All the tomato plants remained symptomless during a period of 45 days although the virus presence in them = was proved by DAS-ELISA. Preparations of isolates D52-I, D52-II, D52-III, D52-IV and D52-V were purified. When studying the biological properties of the isolates it was found that they produced necrotic local lesions on the Sams= un N’N’ inoculated leaves without systemic infection. On the contr= ary, TMV strain D52 infected systemically this cultivar and did not cause necrot= ic local lesions. Therefore, as early as at the first passage TMV D52 was repl= aced by ToMV. However, the five consecutive passages did not induce other change= s in the isolates biological properties (Table 1). The only visible result was gradually increasing of the virus yield in the respective isolates. The D52= -V yield obtained from tomato cv. Mobaci (Tm-1) was approximately equal to tha= t of D52 from tomato cv. Ideal (Tm+). <o:p></o:p> <o:p> </o:p> Tab= le 1. Symptoms* induced by strain D52 and its isolates obtained after passagesin tolerant to tobamoviruses tomato (Tm-1).<o:p></o:p> <div align=3Dcenter> <table class=3DTableGrid border=3D1 cellspacing=3D0 cellpadding=3D0 width= =3D"85%" style=3D'width:85.0%;border-collapse:collapse;border:none;mso-border-alt:s= olid windowtext .5pt; mso-yfti-tbllook:480;mso-padding-alt:0cm 5.4pt 0cm 5.4pt;mso-border-inside= h: .5pt solid windowtext;mso-border-insidev:.5pt solid windowtext'> <tr style=3D'mso-yfti-irow:0;mso-yfti-firstrow:yes;height:34.0pt'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;mso-b= order-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt;height:34.0pt'> Strain/isolate<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border:solid windowtext 1.0pt;borde= r-left: none;mso-border-left-alt:solid windowtext .5pt;mso-border-alt:solid windo= wtext .5pt; padding:0cm 5.4pt 0cm 5.4pt;height:34.0pt'> Tomato cv. Ideal (Tm+)= <o:p></o:p> </td> <td width=3D132 style=3D'width:98.85pt;border:solid windowtext 1.0pt;bord= er-left: none;mso-border-left-alt:solid windowtext .5pt;mso-border-alt:solid windo= wtext .5pt; padding:0cm 5.4pt 0cm 5.4pt;height:34.0pt'> Tomato cv. Mobaci (Tm-= 1)<o:p></o:p> </td> <td width=3D123 style=3D'width:92.15pt;border:solid windowtext 1.0pt;bord= er-left: none;mso-border-left-alt:solid windowtext .5pt;mso-border-alt:solid windo= wtext .5pt; padding:0cm 5.4pt 0cm 5.4pt;height:34.0pt'> Tobacco cv. Samsum N<s= pan style=3D'color:fuchsia'>’NR= 17;<o:p></o:p> </td> <td width=3D85 style=3D'width:63.8pt;border:solid windowtext 1.0pt;border= -left: none;mso-border-left-alt:solid windowtext .5pt;mso-border-alt:solid windo= wtext .5pt; padding:0cm 5.4pt 0cm 5.4pt;height:34.0pt'> Virus<o:p></o:p></span= > </td> </tr> <tr style=3D'mso-yfti-irow:1'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;borde= r-top: none;mso-border-top-alt:solid windowtext .5pt;mso-border-alt:solid window= text .5pt; padding:0cm 5.4pt 0cm 5.4pt'> D52<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D132 style=3D'width:98.85pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D123 style=3D'width:92.15pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> MGM<o:p></o:p><= /p> </td> <td width=3D85 style=3D'width:63.8pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .5pt;mso-border-left-alt:solid windowtext .5pt;mso-borde= r-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> TMV<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:2'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;borde= r-top: none;mso-border-top-alt:solid windowtext .5pt;mso-border-alt:solid window= text .5pt; padding:0cm 5.4pt 0cm 5.4pt'> D52-I<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D132 style=3D'width:98.85pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D123 style=3D'width:92.15pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> NLL<o:p></o:p><= /p> </td> <td width=3D85 style=3D'width:63.8pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .5pt;mso-border-left-alt:solid windowtext .5pt;mso-borde= r-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> ToMV<o:p></o:p>= </td> </tr> <tr style=3D'mso-yfti-irow:3'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;borde= r-top: none;mso-border-top-alt:solid windowtext .5pt;mso-border-alt:solid window= text .5pt; padding:0cm 5.4pt 0cm 5.4pt'> D52-II<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D132 style=3D'width:98.85pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D123 style=3D'width:92.15pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> NLL<o:p></o:p><= /p> </td> <td width=3D85 style=3D'width:63.8pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .5pt;mso-border-left-alt:solid windowtext .5pt;mso-borde= r-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> ToMV<o:p></o:p>= </td> </tr> <tr style=3D'mso-yfti-irow:4'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;borde= r-top: none;mso-border-top-alt:solid windowtext .5pt;mso-border-alt:solid window= text .5pt; padding:0cm 5.4pt 0cm 5.4pt'> D52-III<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D132 style=3D'width:98.85pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D123 style=3D'width:92.15pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> NLL<o:p></o:p><= /p> </td> <td width=3D85 style=3D'width:63.8pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .5pt;mso-border-left-alt:solid windowtext .5pt;mso-borde= r-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> ToMV<o:p></o:p>= </td> </tr> <tr style=3D'mso-yfti-irow:5'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;borde= r-top: none;mso-border-top-alt:solid windowtext .5pt;mso-border-alt:solid window= text .5pt; padding:0cm 5.4pt 0cm 5.4pt'> D52-IV<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D132 style=3D'width:98.85pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D123 style=3D'width:92.15pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> NLL<o:p></o:p><= /p> </td> <td width=3D85 style=3D'width:63.8pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .5pt;mso-border-left-alt:solid windowtext .5pt;mso-borde= r-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> ToMV<o:p></o:p>= </td> </tr> <tr style=3D'mso-yfti-irow:6;mso-yfti-lastrow:yes'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;borde= r-top: none;mso-border-top-alt:solid windowtext .5pt;mso-border-alt:solid window= text .5pt; padding:0cm 5.4pt 0cm 5.4pt'> D52-V<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D132 style=3D'width:98.85pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D123 style=3D'width:92.15pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> NLL<o:p></o:p><= /p> </td> <td width=3D85 style=3D'width:63.8pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .5pt;mso-border-left-alt:solid windowtext .5pt;mso-borde= r-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> ToMV<o:p></o:p>= </td> </tr> </table> </div> *Symptoms: mild green mosaic (M= GM), necrotic local lesions (NLL), sympromless (SL).<o:p></o:p> <o:p> </o:p> Electrophoretic mobi= lity of the new obtained isolates<o:p></o:p> <o:p> </o:p> It is known that TMV strains di= ffer significantly in their electrophoretic mobility from ToMV ones [8, 23, 29] and this characteristic could be used for distinguish= ing both viruses [17]. <o:p></o:p> The strain D52 Rf was determine= d as well as the Rf of its new obtained isolates. Strains U1 for ТМV, GM-0 and GM-1, respectively for ToMV groups “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> and ̶= 0;<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter>, were use= d as control viruses (Fig. 1). It could be seen that there was not statistically proved difference between the mobility of ToMV “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> and ̶= 0;<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> stains bu= t this difference was significant between TMV and ToMV. The D52 Rf was equal to th= at of U1 and all isolates obtained after the passages moved as ToMV strains in= the gel. <o:p></o:p> The electrophoretic pattern of = the initial strain and the control ones as well as the lines of some obtained a= fter passages isolates are shown in Fig. 2. <o:p></o:p> <o:p> </o:p> <![if !vml]><img width=3D595 height=3D256 src=3D"JCC0549NB.files/image002.gif" v:shapes=3D"_x0000_i1025"><![endif]></= p> Fig. </= span>1.&= nbsp;Electrophoretic mobility of str= ain TMV 52 and the isolates obtained after serial passages in tolerant tomato. Control strains: for TMV – U1, for ToMV group “<st1:metricconve= rter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> – G= M-0 and ToMV group “1”– GM-1. Data are the means of three replications. The standard deviation is presented.<o:p></o:p> <o:p> </o:p> <![if !vml]><img width=3D605 height=3D325 src=3D"JCC0549NB.files/image004.gif" v:shapes=3D"_x0000_i1026"><![endif]><s= pan lang=3DEN-US style=3D'font-family:HebarU;mso-ansi-language:EN-US'><o:p></o:= p> Fig. 2. Electrophoretic analysis of virus particles of strain TMV 52 and the isolates obtained after serial passages = in tolerant tomato. Control strains: for TMV - U1, for ToMV group "0"= ; - GM-0 and ToMV group "1" - GM-1. <o:p></o:p> <o:p> </o:p> Serological properti= es of the new obtained isolates<o:p></o:p> <o:p> </o:p> The change = in the serological relationship of the obtained after passages in tolerant tomato isolates and the initial TMV strain D52 was followed (Fig. 3). The precipitation line of the preparation obtained after the first passage form= ed spur towards TMV D52. The same was the result for the isolates after the ne= xt passages. On the other hand, D52-I, D52-II, D52-III, D52-IV and D52-V react= ed as serologically identical to the control ToMV strain GM-<st1:metricconvert= er ProductID=3D"1. A" w:st=3D"on">1. A</st1:metricconverter> fusion of the precipitation lines was observed. This data shows a change of TMV D52 in To= MV as early as at the first passage in tomato with Tm-1 gene. <o:p></o:p></spa= n> <o:p> </o:p> <![if !vml]><img width=3D640 height=3D130 src=3D"JCC0549NB.files/image006.gif" v:shapes=3D"_x0000_i1027"><![endif]><s= pan lang=3DEN-US style=3D'font-size:11.0pt;font-family:HebarU;mso-ansi-language= :EN-US; font-weight:normal'><o:p></o:p> <o:p> </o:p> Fig. 3= . Serological relationship of strain TMV 52 and the isolates obtained after serial passag= es in tolerant tomato. Control strain for ToMV group "1" - GM-1.<o:p= ></o:p> <o:p> </o:p> A method for group determinatio= n of symptomless strains by screening pathogenic revertants in the strain popula= tions and their classification according to Pelham was proposed by Suchov et al. [24]. The authors applied this approach for group determination of the vacc= ine V-<st1:metricconverter ProductID=3D"69, in" w:st=3D"on">69, in</st1:metricc= onverter> whose population about 1 % pathogenic virus particles from group ̶= 0;<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> was prove= d. The presence only of “<st1:metricconverter ProductID=3D"1”" w:st=3D= "on">1”</st1:metricconverter> pathogenic particles indicates that the vaccine strain is from the same gro= up. 250 tomato plants cv. Ideal were single lesion inoculated with isolate D52-V and all of them remained symptomless. This result required searching of new= way for the pathotype determination of the isolate. <o:p></o:p> <o:p> </o:p> Vaccine properties of strain D52 and isolate D52-V<o:p></o:p> <o:p> </o:p> The effectiveness of the protec= tive action of a given vaccine strain depends mainly on two factors. The first o= ne is availability of a good adaptation of the strain to the plants that it prote= cts. Usually the vaccine strains are searched in the virus populations spread in= the respective crop [24] or, if they have been isolated from other culture, the multifold consecutive passages in the plants, for which a vaccine aims to be obtained, improve these properties [20]. ToMV strains that have originated = from tomato and pepper and possess good vaccine effect to the respective culture= can not protect the tobacco against pathogenic TMV and ToMV strains [12, 18]. T= he second factor is the relationship degree between the vaccine and the pathog= enic strains. As their relationship is closer so the interferention between them= is more strongly expresses i.e. the vaccine effectiveness against these strain= s is higher [16, 18]. <o:p></o:p><= /span> On this basis we supposed that = the pathotype of the isolates obtained after passages of strain D52 could be determined comparing their protective properties with ones of other known vaccine from groups “<st1:metricconverter ProductID=3D"0”" w:st= =3D"on">0”</st1:metricconverter> and “<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”= </st1:metricconverter>. At the same time, as D52 and D52-V are symptomless for tomato and pathogenic virions were not found in their preparations, it was interesting to study t= heir protective activity. <o:p></o:p> The results obtained after comparing the vaccine properties of D52 and D52-V with two well known vacci= nes B-5 and V-<st1:metricconverter ProductID=3D"69 in" w:st=3D"on">69 in</st1:m= etricconverter> tomato cv. Ideal are presented in Table <st1:metricconverter ProductID=3D"2= . A" w:st=3D"on">2. A</st1:metricconverter> susceptible to tobamoviruses tomato cultivar was used because strain D52 could not infect tolerant tomato (Tm-1= ). All challenge strains were from group “<st1:metricconverter ProductID= =3D"0”" w:st=3D"on">0”</st1:metricconverter>. They are isolated from different cultures of family Solanaceae a= nd belong to TMV or ToMV. <o:p></o:p> <o:p> </= o:p> Table 2. Vaccine propert= ies of B-5, V-69, D-52 and D52-V in tomato cv. Ideal (Tm+) against pathogenic tobamovirus strains.<o:p></o:p> <div align=3Dcenter> <table class=3DTableNormal border=3D0 cellspacing=3D0 cellpadding=3D0 width= =3D"80%" style=3D'width:80.0%;border-collapse:collapse;mso-padding-alt:0cm 5.35pt 0= cm 5.35pt'> <tr style=3D'mso-yfti-irow:0;mso-yfti-firstrow:yes'> <td width=3D277 colspan=3D3 style=3D'width:208.05pt;border:solid windowte= xt 1.0pt; border-right:none;mso-border-top-alt:solid windowtext .75pt;mso-border-le= ft-alt: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> Pathogenic strains<o:p= ></o:p> </td> <td width=3D321 colspan=3D5 style=3D'width:241.0pt;border:solid windowtex= t 1.0pt; mso-border-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> Vaccine strains<o:p></= o:p> </td> </tr> <tr style=3D'mso-yfti-irow:1'> <td width=3D76 style=3D'width:57.1pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-top-alt: solid windowtext .75pt;mso-border-top-alt:solid windowtext .75pt;mso-bord= er-left-alt: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> Viruses<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-top-alt: solid windowtext .75pt;mso-border-top-alt:solid windowtext .75pt;mso-bord= er-left-alt: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> Strains<o:p></o:p></sp= an> </td> <td width=3D113 style=3D'width:3.0cm;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-top-alt: solid windowtext .75pt;mso-border-top-alt:solid windowtext .75pt;mso-bord= er-left-alt: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> Isolated from<o:p></o:= p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-top-alt: solid windowtext .75pt;mso-border-top-alt:solid windowtext .75pt;mso-bord= er-left-alt: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> -<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border:solid windowtext 1.0pt;border= -top: none;mso-border-top-alt:solid windowtext .75pt;mso-border-alt:solid windo= wtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> B-5<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-bottom:solid wind= owtext 1.0pt; mso-border-top-alt:solid windowtext .75pt;mso-border-top-alt:solid window= text .75pt; mso-border-bottom-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35p= t'> V-69<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:solid windowtext 1.0pt;border= -top: none;mso-border-top-alt:solid windowtext .75pt;mso-border-alt:solid windo= wtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> D52<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .75pt;mso-border-top-alt:solid windowtext .75pt;mso-bord= er-bottom-alt: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> D52-V<o:p></o:p></span= > </td> </tr> <tr style=3D'mso-yfti-irow:2'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> TMV<o:p></o:p><= /p> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> U1<o:p></o:p></= p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tobacco<o:p></o:p></sp= an> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100*<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 97.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 72.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 90<o:p></o:p></= p> </td> </tr> <tr style=3D'mso-yfti-irow:3'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> TGM<o:p></o:p><= /p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tobacco<o:p></o:p></sp= an> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 92.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 85<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 97.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:4'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> B<o:p></o:p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> pepper<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 97.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:5'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> MA<o:p></o:p></= p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 80<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 95<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:6'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> ToMV<o:p></o:p>= </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> ToGM<o:p></o:p>= </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tobacco<o:p></o:p></sp= an> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 97.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 95<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 95<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:7'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> No29<o:p></o:p>= </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> pepper<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 92.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 75<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 85<o:p></o:p></= p> </td> </tr> <tr style=3D'mso-yfti-irow:8'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> GM-0<o:p></o:p>= </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 72.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:9'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> ENP<o:p></o:p><= /p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 85<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 95<o:p></o:p></= p> </td> </tr> <tr style=3D'mso-yfti-irow:10'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> SD<o:p></o:p></= p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 87.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 97.5<o:p></o:p>= </td> </tr> <tr style=3D'mso-yfti-irow:11'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> YM-0<o:p></o:p>= </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 95<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 87.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:12'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> Lat-0<o:p></o:p></span= > </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 92.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 70<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:13'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> GdK<o:p></o:p><= /p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 95<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 97.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:14;mso-yfti-lastrow:yes'> <td width=3D76 style=3D'width:57.1pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-left-al= t: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> -<o:p></o:p> </td> <td width=3D88 style=3D'width:65.9pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-left-al= t: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> -<o:p></o:p> </td> <td width=3D113 style=3D'width:3.0cm;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-left-al= t: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> -<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-left-al= t: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border:solid windowtext 1.0pt;border= -top: none;mso-border-left-alt:solid windowtext .75pt;mso-border-bottom-alt:sol= id windowtext .75pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-bottom:solid wind= owtext 1.0pt; mso-border-bottom-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35p= t'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border:solid windowtext 1.0pt;border= -top: none;mso-border-left-alt:solid windowtext .75pt;mso-border-bottom-alt:sol= id windowtext .75pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-bot= tom-alt: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> </tr> </table> </div> *Number of symptomless p= lants / total number of tested plants x 100 (%).<o:p></o:p> <sp= an lang=3DEN-US style=3D'mso-bidi-font-size:12.0pt;font-family:"Times New Roma= n"; mso-ansi-language:EN-US;font-weight:normal'><o:p> </o:p> The plants infected only with t= he one of the fourth vaccine as well as the control untreated plants remained symptomless to the experiment end. Up to 15th dpi all plants infected only with pathogenic strains developed mosaic symptoms. The percen= tage of symptomless plants inoculated initially with vaccine and after that with pathogenic strain varied in quite wide range. Only single mosaic plants were observed in a part of the variants where the protective properties of both vaccines and D52-V were tested. In analogous variants of TMV D52, the lowest portion of symptomless plants was found. <o:p></o:p> D52-V had very good vaccine activity and the symptomless plants percentage varied from 85 to 100 %= . In variants when the challenge strains were TMV, D52-V showed better vaccine properties than the others, with exception of the variant B-5+U1 where it w= as smaller and of the variant V-69+B where the values were equal. There were l= east symptomless plants in the combination D-52-V+ No.29, in which the pathogenic strain was TMV isolated from pepper. <o:p></o:p> From the obtained results it co= uld be concluded that D52 and D52-V protected tomato against pathogenic TMV and ToMV strains originated from different cultures, as TMV D52 gave way to ToM= V strains V-69 and B-<st1:metricconverter ProductID=3D"5 in" w:st=3D"on">5 in</st1:me= tricconverter> vaccine properties while ToMV D52-V equalized with them. This fact complete= ly correlates with the literature data that the effectiveness of the protection depends on whether the strain is isolated from the plant species it have to= protect, which presume also a good adaptation to it [9, 12, 18].<o:p></o:p></= p> The “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> strains p= ropagate difficult in tolerant to tobamoviruses tomato and accumulate in low concentrations, and in some cases they can not multiply in them at all [22, 28]. Hence, these viruses could not possess protective properties against pathogenic “<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1= ”</st1:metricconverter> strains in tolerant tomato. In order to be proved that D52-V is from group “<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st= 1:metricconverter>, its vaccine action was compared with that of strain V-69 which was from the same group as well as with strain B-5 that multiplied systemically in toler= ant tomato but was from group “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> [24]. The experiment was carried= out with the tolerant tomato cv. Mobaci and pathogenic “<st1:metricconver= ter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> strains w= ere only used (Fig. 4). The percentage of the symptomless plants was 92.5-100 %= for V-69 and 95-100 % for D52-V. The differences were small and the deviat= ions were in favor to the one as well as to the second vaccine. In the same time, only single symptomless plants were established in the variant vaccinated w= ith B-5 although the strain exhibited excellent protective effect in susceptible tomato cultivars.<o:p></o:p> <o:p> </o:p> <![if !vml]><img width=3D579 height=3D272 src=3D"JCC0549NB.files/image008.gif" v:shapes=3D"_x0000_i1028"><![endif]></= p> F= ig. 4= . Vaccine properties of V-69 and 52-V in tolerant to tobamoviruses tomato cv. Mobaci (Tm-1). The vaccine strain B-5 from group “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> is presen= ted for comparison.<o:p></o:p> <o:p> </o:p> These results allow considering that isolate D52-V is from group “<st1:metricconverter ProductID=3D"1= ”" w:st=3D"on">1”</st1:metricconverter> and could be successfully applie= d as vaccine in susceptible and tolerant to ToMV tomato.<o:p></o:p> It is known that the capsid pro= tein is responsible for the appearance of hypersensitive reaction in tobacco with N’ gene resulting in death of the infected cells [1, 2, 3, 11, 25]. Consequently, it could be supposed t= hat all isolates obtained after the passages in tolerant tomato consist of viri= ons, in which there is mutation(s) in the capsid protein. It simultaneously lead= s to changes in the Rf and serological properties of the virus as well as to necrotic local lesion appearance on the inoculated leaf of tobacco with N’ gene.<o:p></o:p> The ability of normal virus propagation in tomato with tolerance Tm-1 gene is connected with amino acid substitutions in the RNA-dependent RNA polymerases [6, 13, 28]. Probably isolate D52-V consists of virions possessing the respective mutation as it = is from group “<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1= ”</st1:metricconverter>. It is established that atenuated and protective properties of ToMV strains = are related with mutations in the virus RNA encoding the same proteins [10]. Therefore, it could be assumed that D52 and D52-V distinguish from the typi= cal pathogenic strains from groups “<st1:metricconverter ProductID=3D"0&#= 8221;" w:st=3D"on">0”</st1:metricconverter> and ‘1” in amino acid alterations again in the RNA-dependent RNA polymerases. The occurred change= s in the respective genes as a result of the passages in tolerant tomato do not concern these regions coding for attenuated properties of D52. The replacin= g of TMV D52 with ToMV isolate D52-I after the first passage and the subsequent mutations selection, resulting in the multiplication adaptation of the virus population in tolerant tomato, leads also in improving the vaccine properti= es of D52-V in comparison with D52. <o:p></o:p> The obtained isolate D52-V protecting excellent susceptible and tolerant tomato against pathogenic tobamovirus strains is v= ery stable because pathogenic virions are not found in its preparations. In this characteristic it exceeds known and widely used in practice vaccines [16, 2= 4]. At the same time, it is symptomless for pepper, a culture often growing in adjacency of tomato. That is why, the vaccinated with D52-V tomato could no= t be virus source for tobacco mosaic disease in pepper cultivars susceptible to tobamoviruses. Consequently, the objections against using vaccine strains do not referred to D52-V that makes it appropriate for application in the production of green house and field tomato. <o:p></o:p> <o:p> </o:p> References<o:p></o:p= > <o:p> </o:p> <![if !supportLists]>1. <![endif]>Culver, J. N., W.O. Dawson, 1991. Mol. Plant-Microbe Interact., 4, 458-463. <o:= p></o:p> <![if !supportLists]><s= pan style=3D'mso-list:Ignore'>2.&n= bsp; <![endif]>Culver, J.N., G. C. Lindbeck, W.O. Dawson, 1991. Ann. Rev. Phytopathol., 29, 193-217. <o:p></o:p> <![if !supportLists]>3. <![endif]>Culver, J.N., G. Stu= bbs, W.O. Dawson, 1994. J. Mol. Biol., = 242, 130-138.<o:p></o:p> <![if !supportLists]>4. <![endif]>Garcia-Lugue, I., M.= T. Serra, E. Alonso, B. Wicke, M. L. Ferrero, J. R. Diaz-Ruiz, 1990. J. Phytopathology, 129, 1-8. <o:p></o:p> <![if !supportLists]>5. <![endif]>Gooding, G. V., T. T. Hebert, 1967. Phytopathology, 61, 943-944.<o:p></o:p> <![if !s= upportLists]>6. <![endif]>Hamamoto, H., Y. 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CMI/AAB Descriptions = of plant viruses, No. 151.<o:= p></o:p> </div> </body> </html> ------=_NextPart_01C5DDFE.C36A4340 Content-Location: file:///C:/236AEE02/JCC0549NB.files/header.htm Content-Transfer-Encoding: quoted-printable Content-Type: text/html; charset="us-ascii" <html xmlns:v=3D"urn:schemas-microsoft-com:vml" xmlns:o=3D"urn:schemas-microsoft-com:office:office" xmlns:w=3D"urn:schemas-microsoft-com:office:word" xmlns:st1=3D"urn:schemas-microsoft-com:office:smarttags" xmlns=3D"http://www.w3.org/TR/REC-html40"> <head> <meta http-equiv=3DContent-Type content=3D"text/html; charset=3Dus-ascii"> <meta name=3DProgId content=3DWord.Document> <meta name=3DGenerator content=3D"Microsoft Word 11"> <meta name=3DOriginator content=3D"Microsoft Word 11"> <link id=3DMain-File rel=3DMain-File href=3D"../JCC0549NB.htm"> <![if IE]> <base href=3D"file:///C:\236AEE02\JCC0549NB.files\header.htm" id=3D"webarch_temp_base_tag"> <![endif]><o:SmartTagType 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MIME-Version: 1.0 Content-Type: multipart/related; boundary="----=_NextPart_01C5DDFE.C36A4340" Този документ е еднофайлова Web страница, известна също като файл за Web архив. Ако виждате това съобщение, вашият браузър или редактор не поддържа файлове за Web архиви. Изтеглете браузър, който поддържа Web архиви, например Microsoft Internet Explorer. ------=_NextPart_01C5DDFE.C36A4340 Content-Location: file:///C:/236AEE02/JCC0549NB.htm Content-Transfer-Encoding: quoted-printable Content-Type: text/html; charset="us-ascii" Резултати </ti= tle> <o:SmartTagType namespaceuri=3D"urn:schemas-microsoft-com:office:smarttags" name=3D"metricconverter"/>  <style>  </style>  </head> <body lang=3DBG link=3Dblue vlink=3Dpurple style=3D'tab-interval:36.0pt'> <div class=3DSection1> JOURNAL OF CULTURE COLLECTIONS<o:p></o:p> Volume 4, 2004-2005, pp. </span= >68-77<o:p></o:p></= i> <o:p> </o:p> <o:p> </o:p> <o:p> </o:p> Vaccine strain of tomato mosaic virus obtained after serial passages of attenuated strain of tobacco mosaic virus in tolerant to tobamoviruses tomato<o:p></o:p> <= o:p> </o:p> E= lisaveta Stoimenova1* and Angela Yordanova2<o:p></o:p><= /p> <o:p> </o:p> 1Institute of Genetics, Bulgarian Aca= demy of Sciences, 1113 Sofia, Bulgaria</spa= n>;<o:p></o:p></= span> 2<s= pan lang=3DEN-US style=3D'mso-bidi-font-size:12.0pt;font-family:"Times New Roma= n"; letter-spacing:-.1pt;mso-ansi-language:EN-US;font-weight:normal'>National B= ank for Industrial Microorganisms and Cell Cultures, 1113 Sofia, P.O.Box 239, Bulgaria<o:p></o:p> <o:p> </o:p> <o:p> </o:p> Summary<o:p></o:p></= span> <o:p> </o:p></s= pan> An attenuated strain of tobacco mosaic virus= </spa= n>(TMV<= span style=3D'mso-bidi-font-size:12.0pt;font-family:"Times New Roman";color:blac= k; font-weight:normal'>) <s= pan lang=3DEN-US style=3D'mso-bidi-font-size:12.0pt;font-family:"Times New Roma= n"; color:black;mso-ansi-language:EN-US;font-weight:normal'>isolated from tobac= co and symptopmless for tomato and pepper was put in fivefold passage in toler= ant to tobamoviruses tomato. The changes in the strain population after the pas= sages were studied by determination of the virion electrophoretic mobility, serological and biological properties of purified preparations of the obtai= ned isolates. All performed experiments pro= ved that as early as in the first passage TMV was replaced by a tomato mosaic virus (<s= pan lang=3DEN-US style=3D'mso-bidi-font-size:12.0pt;font-family:"Times New Roma= n"; color:black;mso-ansi-language:EN-US;font-weight:normal'>ToMV).= After five serial passages the presence of virions cau= sing mosaic in susceptible and/or tolerant tomato were not found. The study for vaccine properties of the new obtained isolate ToMV D52-V showed that it belonged to Pelham’s group “<st1:metricconverter ProductID=3D"1= ”" w:st=3D"on">1”</st1:metricconverter> since it was effective in tomato carrying Tm-1 gene. Its protective action did not give way to that of analo= gous vaccines applied in the practice and hence strain D52-V could be successful= ly used for protection of susceptible and tolerant tomato against pathogenic tobamovirus strains. <sp= an style=3D'mso-bidi-font-size:12.0pt;font-family:"Times New Roman";color:blac= k; font-weight:normal'><o:p></o:p> <o:p> </o:p> Introduction<o:p></o= :p> <o:p> </o:p> In the 70s and 80s of the twent= ieth century vaccine strains of tomato mosaic virus (ToMV) have been widely used= for tomato protection against pathogenic tobamovirus strains. The application o= f this method in the practice proved its effectiveness however, some problems aros= e. The main objections were related to the presence of a little of pathogenic particles (up to 1 %) in the vaccine preparations and to the risk of appearance of new highly virulent virus strains. Apprehensions were stated about the vaccine strains translation upon other crops, for which these str= ains were pathogenic [7, 24]. The method application in tomato was restrict= ed as consequence of the new cultivars creation - tolerant (Tm-1) or resistant= to tobamoviruses ones (Tm-2 and Tm-22). However, the broad use of t= hese cultivars led to raising of ToMV strains from group “1”, “2” and “1.2” according to Pelham’s classification [14], overcoming the respective resistance of the Tm-1, Tm-2= and Tm-1+Tm-2 genes [14, 15, 16, 20]. In tolerant to tobamovirus= es tomato some “0” strains are able to cause symptomless systemic infection while “1” strains induce mosaic symptoms. Cultivars carrying Tm-1 gene are spread in our country and are used in the production= of middle early and late tomato [20]. <o:p></o:p> In Bulgaria, ToMV strains from groups “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0R= 21;</st1:metricconverter> and “<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”= </st1:metricconverter> were found in yield tomato but strains of tobacco mosaic virus (TMV) were n= ot isolated. The latter was occasionally established in green houses. However,= in pepper TMV strains were frequently occurred besides ToMV ones and could even predominate [20]. The actual lack of TMV strains in tomato could be explain= ed on one hand with the fact that TMV strains easily mutated in ToMV, as found= in many investigations and on the other hand ToMV had a faster rate of multiplication or a greater invasiveness in tomato plants compared with TMV [26]. In the nature when TMV was found in tomato, it was mixed with ToMV [1= 6, 20]. <o:p></o:p> In the present work the obtaini= ng of a new vaccine ToMV strain is described which could be used for protectio= n of tomato with Tm-1 gene against pathogenic virus strains from group “<s= t1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter>. <o:p></o= :p> <o:p> </o:p> Materials and Method= s<o:p></o:p> <o:p> </o:p> Virus strains. </spa= n>Objects of the study we= re TMV strain D52 - symptomless for tomato and pepper and mild for tobacco as well= as its derivative isolates obtained after passages in tolerant tomato.<o:p></o= :p> Twenty one TMV and ToMV strains were included in the investigation. The ToMV vaccine strains were: t= he Bulgarian B-5 from group “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> after Pelham (National Bank for Industrial microorganisms and Cell Cultures 2024) and the Russian V-69 from group “<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1̶= 1;</st1:metricconverter> (NBIMCC 2114) [21, 24]. The used pathogenic strains were: ТМV -= U1 (3405), TGM (3294), B (2022), МА (2077) and Sm (2125), the latt= er being from group “<st1:metricconverter ProductID=3D"1”" w:st=3D= "on">1”</st1:metricconverter>; ТоМV strains from group “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> – E= NP (2074), SD (2076), Lat-0 (2064), GdK (2075), GM-0 (2078), YM-0 (2348), No29 (3297), ToGM (3296); ТоМV strains from group “<st1:= metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> – N= H-69, SPS, GeBl (2112), So69 (2111), GM-1 (2349) and YM-1 (2348). The plants from which TMV and ToMV “<st1:metricconverter ProductID=3D"0”" w:st= =3D"on">0”</st1:metricconverter> strains have been isolated are shown in Table 2. The strains from group = 220;<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> originate= from tomato. Strains TGM, B, GM-0, No29, ToGM, YM-0, GM-1 and YM-1 are from Bulg= aria [19]. The authors express much gratitude to Prof. Dr. A. Th. B Ra= st, Prof. Dr. H. Laterrot and Prof. Dr. F. Garcia-Arenal for the supplying of other tobamoviruses.<o:p></o:p> Test plants.<= /b> The following cultivars were used: tomato cv. Ideal susceptible to tobamoviruses and cv. Mobaci carrying Tm-1 tolerance gene to this virus gro= up; tobacco cv. Samsun N’N’ (N’ gene from Nicotiana sylvestris) and cv. Sams= un NN (N gene from N. glutinosa, localizing the tobamoviruses). The plants were grown in autoclaved soil and sterile metal pots. Seeds obtained from plants proved as virusless were use= d.<o:p></o:p> Virus propagation and purification. Tobacco plants cv. Samsun N’N’ in 5-6 leaf stage were inoculated with TMV strains. The multiplication of ToMV “<st1:metricconverter ProductID=3D"0”" w= :st=3D"on">0”</st1:metricconverter> strains was done in tomato cv. Ideal while strains from group “<st1:m= etricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> were prop= agated in tomato cv. Mobaci and plants in cotyledon-first real leaf stage were infected. The mosaic leaves were collected 21 days past inoculation (dpi) w= ith pathogenic strains. The plants inoculated with vaccine strains remained symptomless and all the leaves were harvested. Purified virus preparations = were obtained by the method of Gooding and Hebert [5]. The virus concentration w= as spectrophotometrically determined. Tobacco cv. Samsun NN was used for production of single lesion isolates. <o:p></o:p> Passages in tolerant tomato. Tomato cv. Mobaci was inoculated with Тi= 2;V D52 preparation (0.1 mg/ml), the plants were grown 45 days and the symptoms and the systemic virus distribution were recorded. All plants were symptomless and a purified preparation named D52-I was made from them. Toma= to plants were again mass inoculated with this isolate and they remained symptomless. A preparation D52-II was similarly obtained. The procedure was multifold repeated and isolates D52-III, D52-IV and D52-V were purified. 40 plants were inoculated at every passage.<o:p></o:p> As in these experiments it was exceptionally important to exclude the contamination, besides the mentioned above measures all pots were enclosed = with polyethylene. In this way the touch was avoided between the inoculated with different viruses plants, as well as the contact during watering.<o:p></o:p= > Electrophor= etic mobility of virus particles. The electrophoretic mobility (Rf) of tobamovirus virions was determined in 0.7 % agarose horizontal gel, in <st1:metricconve= rter ProductID=3D"40 mM" w:st=3D"on">40 mM</st1:metricconverter> Tris-acetate bu= ffer, pH 7.8, containing <st1:metricconverter ProductID=3D"2 mM" w:st=3D"on">2 mM</s= t1:metricconverter> EDTA. The virus preparations were applied in concentration of 10 mg/ml. The electrophoresis was carried out at 60 V for 2.5 hours. The gels were stained with 0.2 % amido black in <st1:metricconverter ProductID=3D"0.1 M" w:st=3D"= on">0.1 M</st1:metricconverter> sodium acetate, containing 10 % glycerol [4]. The = electrophoregrams were dried and stained additionally with Coomassie Brilliant Blue R-250 [27= ].<o:p></o:p> The Rf of the samples was accounted according to the equation Rf =3D= S/F, where F was the distance from the start to the front and S was the sample distance. This value was calculated to Rf of the type strain U1 accepted as 100 %. The standard deviation of three repetitions was determined. <o:= p></o:p> <h1 style=3D'text-indent:1.0cm;line-height:normal;mso-pagination:none'>Immunoelectrophoretic analysis. Counter immunoelectrophoresis was performed in 0.9% agarose gel in <st1:metricconve= rter ProductID=3D"0.075 M" w:st=3D"on">0.075 M</st1:metricconverter> veronal buf= fer, pH 8.6, at 150 V, 10 oC for two hours. The virus preparations = were applied in concentration of 5 mg/ml (25 <span lang=3DEN-US style=3D'font-family:Symbol;mso-ascii-font-family:"Times New Roman";mso-han= si-font-family: "Times New Roman";mso-char-type:symbol;mso-symbol-font-family:Symbol; font-weight:normal;mso-bidi-font-weight:bold'>mg per start). The antiserum against ToMV in dilution of 1:4 was used. The gels were stained w= ith Coomassie Brilliant Blue R-250 [27].<o:p></o:p></h1> Biological properties. Ten plants of each toma= to cvs Ideal, Mobaci and tobacco cv. Samsun N’N’ were inoculated with purified preparations of strain D52 and isolates D52-I, D52-II, D52-III, D5= 2-IV and D52-V. The symptoms on the inoculated leaves were recorded on the 7<sup= >th and 10th dpi and on the young leaves on the 40th day.= The tobamovirus presence was proved by DAS-ELISA.<o:p></o:p> Vaccine properties. Two experiments for comparative testing of D52 and D52-V vaccine properties were carried out. In the first experiment, tomato plants cv. Ideal were vaccinated with B-5, V-69, D52 and D52-V and after 7 days they were challenge inoculated with TMV and ToMV pathogenic str= ains from group “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0= ”</st1:metricconverter>. In the second case, tomato plants cv. Mobaci were vaccinated with B-5, V-69 and D52-V and after 7 days they were infected with group “<= st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> pathogenic strains. The following variants were used as controls: untreated plants (healthy control); plants inoculated only with the respective vaccine strain and plants infected only with the respective pathogenic strain. The number = of mosaic plants was accounted 30 days after the mosaic symptoms appearance in= the plants infected only with pathogenic strains. 40 plants were used in each variant.<o:p></o:p> <o:p> </o:p> Results and Discussion<o:p></o:p= > <o:p> </o:p> TMV strain No. 52 was isol= ated when the tobamovirus strain variability in commercial tobacco in Bulgaria w= as studied. This strain does not differ in electrophoretic mobility and serological properties from the typical TMV strains but it induces mild gre= en mosaic in tomato, pepper and tobacco [17, 19]. A significant percentage (12 %) symptomless plants was found after single lesion infection of tomato (Tm+) with No. 52. After a purposive selection a TMV strain D52= was obtained that was symptomless for tomato and pepper and mild for tobacco. In spite of the mult= ifold passages of strain D52 in tomato (Tm+), ToMV virions were not established in its preparations (testing on tobacco cv. Samsun N’N’).<o:p></o:= p> <o:p> </o:p> Passages and biological properti= es of the obtained tobamovirus isolates<o:p></o:p> <o:p> </o:p> Five consecutive passages of st= rain D52 in tolerant to tobamoviruses tomato (Tm-1) was performed. The initial infection was made with a preparation obtained from tobacco cv. Samsum N’N’ in order to exclude the opportunity of ToMV particles pres= ence in the started inoculum. <o:p= ></o:p> All the tomato plants remained symptomless during a period of 45 days although the virus presence in them = was proved by DAS-ELISA. Preparations of isolates D52-I, D52-II, D52-III, D52-IV and D52-V were purified. When studying the biological properties of the isolates it was found that they produced necrotic local lesions on the Sams= un N’N’ inoculated leaves without systemic infection. On the contr= ary, TMV strain D52 infected systemically this cultivar and did not cause necrot= ic local lesions. Therefore, as early as at the first passage TMV D52 was repl= aced by ToMV. However, the five consecutive passages did not induce other change= s in the isolates biological properties (Table 1). The only visible result was gradually increasing of the virus yield in the respective isolates. The D52= -V yield obtained from tomato cv. Mobaci (Tm-1) was approximately equal to tha= t of D52 from tomato cv. Ideal (Tm+). <o:p></o:p> <o:p> </o:p> Tab= le 1. Symptoms* induced by strain D52 and its isolates obtained after passagesin tolerant to tobamoviruses tomato (Tm-1).<o:p></o:p> <div align=3Dcenter> <table class=3DTableGrid border=3D1 cellspacing=3D0 cellpadding=3D0 width= =3D"85%" style=3D'width:85.0%;border-collapse:collapse;border:none;mso-border-alt:s= olid windowtext .5pt; mso-yfti-tbllook:480;mso-padding-alt:0cm 5.4pt 0cm 5.4pt;mso-border-inside= h: .5pt solid windowtext;mso-border-insidev:.5pt solid windowtext'> <tr style=3D'mso-yfti-irow:0;mso-yfti-firstrow:yes;height:34.0pt'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;mso-b= order-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt;height:34.0pt'> Strain/isolate<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border:solid windowtext 1.0pt;borde= r-left: none;mso-border-left-alt:solid windowtext .5pt;mso-border-alt:solid windo= wtext .5pt; padding:0cm 5.4pt 0cm 5.4pt;height:34.0pt'> Tomato cv. Ideal (Tm+)= <o:p></o:p> </td> <td width=3D132 style=3D'width:98.85pt;border:solid windowtext 1.0pt;bord= er-left: none;mso-border-left-alt:solid windowtext .5pt;mso-border-alt:solid windo= wtext .5pt; padding:0cm 5.4pt 0cm 5.4pt;height:34.0pt'> Tomato cv. Mobaci (Tm-= 1)<o:p></o:p> </td> <td width=3D123 style=3D'width:92.15pt;border:solid windowtext 1.0pt;bord= er-left: none;mso-border-left-alt:solid windowtext .5pt;mso-border-alt:solid windo= wtext .5pt; padding:0cm 5.4pt 0cm 5.4pt;height:34.0pt'> Tobacco cv. Samsum N<s= pan style=3D'color:fuchsia'>’NR= 17;<o:p></o:p> </td> <td width=3D85 style=3D'width:63.8pt;border:solid windowtext 1.0pt;border= -left: none;mso-border-left-alt:solid windowtext .5pt;mso-border-alt:solid windo= wtext .5pt; padding:0cm 5.4pt 0cm 5.4pt;height:34.0pt'> Virus<o:p></o:p></span= > </td> </tr> <tr style=3D'mso-yfti-irow:1'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;borde= r-top: none;mso-border-top-alt:solid windowtext .5pt;mso-border-alt:solid window= text .5pt; padding:0cm 5.4pt 0cm 5.4pt'> D52<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D132 style=3D'width:98.85pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D123 style=3D'width:92.15pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> MGM<o:p></o:p><= /p> </td> <td width=3D85 style=3D'width:63.8pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .5pt;mso-border-left-alt:solid windowtext .5pt;mso-borde= r-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> TMV<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:2'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;borde= r-top: none;mso-border-top-alt:solid windowtext .5pt;mso-border-alt:solid window= text .5pt; padding:0cm 5.4pt 0cm 5.4pt'> D52-I<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D132 style=3D'width:98.85pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D123 style=3D'width:92.15pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> NLL<o:p></o:p><= /p> </td> <td width=3D85 style=3D'width:63.8pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .5pt;mso-border-left-alt:solid windowtext .5pt;mso-borde= r-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> ToMV<o:p></o:p>= </td> </tr> <tr style=3D'mso-yfti-irow:3'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;borde= r-top: none;mso-border-top-alt:solid windowtext .5pt;mso-border-alt:solid window= text .5pt; padding:0cm 5.4pt 0cm 5.4pt'> D52-II<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D132 style=3D'width:98.85pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D123 style=3D'width:92.15pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> NLL<o:p></o:p><= /p> </td> <td width=3D85 style=3D'width:63.8pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .5pt;mso-border-left-alt:solid windowtext .5pt;mso-borde= r-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> ToMV<o:p></o:p>= </td> </tr> <tr style=3D'mso-yfti-irow:4'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;borde= r-top: none;mso-border-top-alt:solid windowtext .5pt;mso-border-alt:solid window= text .5pt; padding:0cm 5.4pt 0cm 5.4pt'> D52-III<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D132 style=3D'width:98.85pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D123 style=3D'width:92.15pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> NLL<o:p></o:p><= /p> </td> <td width=3D85 style=3D'width:63.8pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .5pt;mso-border-left-alt:solid windowtext .5pt;mso-borde= r-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> ToMV<o:p></o:p>= </td> </tr> <tr style=3D'mso-yfti-irow:5'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;borde= r-top: none;mso-border-top-alt:solid windowtext .5pt;mso-border-alt:solid window= text .5pt; padding:0cm 5.4pt 0cm 5.4pt'> D52-IV<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D132 style=3D'width:98.85pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D123 style=3D'width:92.15pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> NLL<o:p></o:p><= /p> </td> <td width=3D85 style=3D'width:63.8pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .5pt;mso-border-left-alt:solid windowtext .5pt;mso-borde= r-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> ToMV<o:p></o:p>= </td> </tr> <tr style=3D'mso-yfti-irow:6;mso-yfti-lastrow:yes'> <td width=3D101 style=3D'width:75.7pt;border:solid windowtext 1.0pt;borde= r-top: none;mso-border-top-alt:solid windowtext .5pt;mso-border-alt:solid window= text .5pt; padding:0cm 5.4pt 0cm 5.4pt'> D52-V<o:p></o:p> </td> <td width=3D114 style=3D'width:85.4pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D132 style=3D'width:98.85pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> SL<o:p></o:p></= p> </td> <td width=3D123 style=3D'width:92.15pt;border-top:none;border-left:none; border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; mso-border-top-alt:solid windowtext .5pt;mso-border-left-alt:solid window= text .5pt; mso-border-alt:solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> NLL<o:p></o:p><= /p> </td> <td width=3D85 style=3D'width:63.8pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .5pt;mso-border-left-alt:solid windowtext .5pt;mso-borde= r-alt: solid windowtext .5pt;padding:0cm 5.4pt 0cm 5.4pt'> ToMV<o:p></o:p>= </td> </tr> </table> </div> *Symptoms: mild green mosaic (M= GM), necrotic local lesions (NLL), sympromless (SL).<o:p></o:p> <o:p> </o:p> Electrophoretic mobi= lity of the new obtained isolates<o:p></o:p> <o:p> </o:p> It is known that TMV strains di= ffer significantly in their electrophoretic mobility from ToMV ones [8, 23, 29] and this characteristic could be used for distinguish= ing both viruses [17]. <o:p></o:p> The strain D52 Rf was determine= d as well as the Rf of its new obtained isolates. Strains U1 for ТМV, GM-0 and GM-1, respectively for ToMV groups “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> and ̶= 0;<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter>, were use= d as control viruses (Fig. 1). It could be seen that there was not statistically proved difference between the mobility of ToMV “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> and ̶= 0;<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> stains bu= t this difference was significant between TMV and ToMV. The D52 Rf was equal to th= at of U1 and all isolates obtained after the passages moved as ToMV strains in= the gel. <o:p></o:p> The electrophoretic pattern of = the initial strain and the control ones as well as the lines of some obtained a= fter passages isolates are shown in Fig. 2. <o:p></o:p> <o:p> </o:p> <![if !vml]><img width=3D595 height=3D256 src=3D"JCC0549NB.files/image002.gif" v:shapes=3D"_x0000_i1025"><![endif]></= p> Fig. </= span>1.&= nbsp;Electrophoretic mobility of str= ain TMV 52 and the isolates obtained after serial passages in tolerant tomato. Control strains: for TMV – U1, for ToMV group “<st1:metricconve= rter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> – G= M-0 and ToMV group “1”– GM-1. Data are the means of three replications. The standard deviation is presented.<o:p></o:p> <o:p> </o:p> <![if !vml]><img width=3D605 height=3D325 src=3D"JCC0549NB.files/image004.gif" v:shapes=3D"_x0000_i1026"><![endif]><s= pan lang=3DEN-US style=3D'font-family:HebarU;mso-ansi-language:EN-US'><o:p></o:= p> Fig. 2. Electrophoretic analysis of virus particles of strain TMV 52 and the isolates obtained after serial passages = in tolerant tomato. Control strains: for TMV - U1, for ToMV group "0"= ; - GM-0 and ToMV group "1" - GM-1. <o:p></o:p> <o:p> </o:p> Serological properti= es of the new obtained isolates<o:p></o:p> <o:p> </o:p> The change = in the serological relationship of the obtained after passages in tolerant tomato isolates and the initial TMV strain D52 was followed (Fig. 3). The precipitation line of the preparation obtained after the first passage form= ed spur towards TMV D52. The same was the result for the isolates after the ne= xt passages. On the other hand, D52-I, D52-II, D52-III, D52-IV and D52-V react= ed as serologically identical to the control ToMV strain GM-<st1:metricconvert= er ProductID=3D"1. A" w:st=3D"on">1. A</st1:metricconverter> fusion of the precipitation lines was observed. This data shows a change of TMV D52 in To= MV as early as at the first passage in tomato with Tm-1 gene. <o:p></o:p></spa= n> <o:p> </o:p> <![if !vml]><img width=3D640 height=3D130 src=3D"JCC0549NB.files/image006.gif" v:shapes=3D"_x0000_i1027"><![endif]><s= pan lang=3DEN-US style=3D'font-size:11.0pt;font-family:HebarU;mso-ansi-language= :EN-US; font-weight:normal'><o:p></o:p> <o:p> </o:p> Fig. 3= . Serological relationship of strain TMV 52 and the isolates obtained after serial passag= es in tolerant tomato. Control strain for ToMV group "1" - GM-1.<o:p= ></o:p> <o:p> </o:p> A method for group determinatio= n of symptomless strains by screening pathogenic revertants in the strain popula= tions and their classification according to Pelham was proposed by Suchov et al. [24]. The authors applied this approach for group determination of the vacc= ine V-<st1:metricconverter ProductID=3D"69, in" w:st=3D"on">69, in</st1:metricc= onverter> whose population about 1 % pathogenic virus particles from group ̶= 0;<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> was prove= d. The presence only of “<st1:metricconverter ProductID=3D"1”" w:st=3D= "on">1”</st1:metricconverter> pathogenic particles indicates that the vaccine strain is from the same gro= up. 250 tomato plants cv. Ideal were single lesion inoculated with isolate D52-V and all of them remained symptomless. This result required searching of new= way for the pathotype determination of the isolate. <o:p></o:p> <o:p> </o:p> Vaccine properties of strain D52 and isolate D52-V<o:p></o:p> <o:p> </o:p> The effectiveness of the protec= tive action of a given vaccine strain depends mainly on two factors. The first o= ne is availability of a good adaptation of the strain to the plants that it prote= cts. Usually the vaccine strains are searched in the virus populations spread in= the respective crop [24] or, if they have been isolated from other culture, the multifold consecutive passages in the plants, for which a vaccine aims to be obtained, improve these properties [20]. ToMV strains that have originated = from tomato and pepper and possess good vaccine effect to the respective culture= can not protect the tobacco against pathogenic TMV and ToMV strains [12, 18]. T= he second factor is the relationship degree between the vaccine and the pathog= enic strains. As their relationship is closer so the interferention between them= is more strongly expresses i.e. the vaccine effectiveness against these strain= s is higher [16, 18]. <o:p></o:p><= /span> On this basis we supposed that = the pathotype of the isolates obtained after passages of strain D52 could be determined comparing their protective properties with ones of other known vaccine from groups “<st1:metricconverter ProductID=3D"0”" w:st= =3D"on">0”</st1:metricconverter> and “<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”= </st1:metricconverter>. At the same time, as D52 and D52-V are symptomless for tomato and pathogenic virions were not found in their preparations, it was interesting to study t= heir protective activity. <o:p></o:p> The results obtained after comparing the vaccine properties of D52 and D52-V with two well known vacci= nes B-5 and V-<st1:metricconverter ProductID=3D"69 in" w:st=3D"on">69 in</st1:m= etricconverter> tomato cv. Ideal are presented in Table <st1:metricconverter ProductID=3D"2= . A" w:st=3D"on">2. A</st1:metricconverter> susceptible to tobamoviruses tomato cultivar was used because strain D52 could not infect tolerant tomato (Tm-1= ). All challenge strains were from group “<st1:metricconverter ProductID= =3D"0”" w:st=3D"on">0”</st1:metricconverter>. They are isolated from different cultures of family Solanaceae a= nd belong to TMV or ToMV. <o:p></o:p> <o:p> </= o:p> Table 2. Vaccine propert= ies of B-5, V-69, D-52 and D52-V in tomato cv. Ideal (Tm+) against pathogenic tobamovirus strains.<o:p></o:p> <div align=3Dcenter> <table class=3DTableNormal border=3D0 cellspacing=3D0 cellpadding=3D0 width= =3D"80%" style=3D'width:80.0%;border-collapse:collapse;mso-padding-alt:0cm 5.35pt 0= cm 5.35pt'> <tr style=3D'mso-yfti-irow:0;mso-yfti-firstrow:yes'> <td width=3D277 colspan=3D3 style=3D'width:208.05pt;border:solid windowte= xt 1.0pt; border-right:none;mso-border-top-alt:solid windowtext .75pt;mso-border-le= ft-alt: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> Pathogenic strains<o:p= ></o:p> </td> <td width=3D321 colspan=3D5 style=3D'width:241.0pt;border:solid windowtex= t 1.0pt; mso-border-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> Vaccine strains<o:p></= o:p> </td> </tr> <tr style=3D'mso-yfti-irow:1'> <td width=3D76 style=3D'width:57.1pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-top-alt: solid windowtext .75pt;mso-border-top-alt:solid windowtext .75pt;mso-bord= er-left-alt: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> Viruses<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-top-alt: solid windowtext .75pt;mso-border-top-alt:solid windowtext .75pt;mso-bord= er-left-alt: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> Strains<o:p></o:p></sp= an> </td> <td width=3D113 style=3D'width:3.0cm;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-top-alt: solid windowtext .75pt;mso-border-top-alt:solid windowtext .75pt;mso-bord= er-left-alt: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> Isolated from<o:p></o:= p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-top-alt: solid windowtext .75pt;mso-border-top-alt:solid windowtext .75pt;mso-bord= er-left-alt: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> -<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border:solid windowtext 1.0pt;border= -top: none;mso-border-top-alt:solid windowtext .75pt;mso-border-alt:solid windo= wtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> B-5<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-bottom:solid wind= owtext 1.0pt; mso-border-top-alt:solid windowtext .75pt;mso-border-top-alt:solid window= text .75pt; mso-border-bottom-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35p= t'> V-69<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:solid windowtext 1.0pt;border= -top: none;mso-border-top-alt:solid windowtext .75pt;mso-border-alt:solid windo= wtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> D52<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-top= -alt: solid windowtext .75pt;mso-border-top-alt:solid windowtext .75pt;mso-bord= er-bottom-alt: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> D52-V<o:p></o:p></span= > </td> </tr> <tr style=3D'mso-yfti-irow:2'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> TMV<o:p></o:p><= /p> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> U1<o:p></o:p></= p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tobacco<o:p></o:p></sp= an> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100*<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 97.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 72.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 90<o:p></o:p></= p> </td> </tr> <tr style=3D'mso-yfti-irow:3'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> TGM<o:p></o:p><= /p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tobacco<o:p></o:p></sp= an> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 92.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 85<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 97.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:4'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> B<o:p></o:p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> pepper<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 97.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:5'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> MA<o:p></o:p></= p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 80<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 95<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:6'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> ToMV<o:p></o:p>= </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> ToGM<o:p></o:p>= </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tobacco<o:p></o:p></sp= an> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 97.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 95<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 95<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:7'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> No29<o:p></o:p>= </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> pepper<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 92.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 75<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 85<o:p></o:p></= p> </td> </tr> <tr style=3D'mso-yfti-irow:8'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> GM-0<o:p></o:p>= </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 72.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:9'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> ENP<o:p></o:p><= /p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 85<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 95<o:p></o:p></= p> </td> </tr> <tr style=3D'mso-yfti-irow:10'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> SD<o:p></o:p></= p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 87.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 97.5<o:p></o:p>= </td> </tr> <tr style=3D'mso-yfti-irow:11'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> YM-0<o:p></o:p>= </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 95<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 87.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:12'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> Lat-0<o:p></o:p></span= > </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 92.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 70<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:13'> <td width=3D76 style=3D'width:57.1pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> “<o:p></o:p></sp= an> </td> <td width=3D88 style=3D'width:65.9pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> GdK<o:p></o:p><= /p> </td> <td width=3D113 style=3D'width:3.0cm;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> tomato<o:p></o:p></spa= n> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-left:solid window= text 1.0pt; mso-border-left-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;padding:0cm 5.35pt 0cm 5.35pt'> 95<o:p></o:p></= p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:none;border-right:solid windowtext 1.0pt;mso-border-left-al= t: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 97.5<o:p></o:p>= </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-right:solid windo= wtext 1.0pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> </tr> <tr style=3D'mso-yfti-irow:14;mso-yfti-lastrow:yes'> <td width=3D76 style=3D'width:57.1pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-left-al= t: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> -<o:p></o:p> </td> <td width=3D88 style=3D'width:65.9pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-left-al= t: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> -<o:p></o:p> </td> <td width=3D113 style=3D'width:3.0cm;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-left-al= t: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> -<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:solid wi= ndowtext 1.0pt; border-bottom:solid windowtext 1.0pt;border-right:none;mso-border-left-al= t: solid windowtext .75pt;mso-border-bottom-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 0<o:p></o:p> </td> <td width=3D64 style=3D'width:48.2pt;border:solid windowtext 1.0pt;border= -top: none;mso-border-left-alt:solid windowtext .75pt;mso-border-bottom-alt:sol= id windowtext .75pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border:none;border-bottom:solid wind= owtext 1.0pt; mso-border-bottom-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35p= t'> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border:solid windowtext 1.0pt;border= -top: none;mso-border-left-alt:solid windowtext .75pt;mso-border-bottom-alt:sol= id windowtext .75pt; mso-border-right-alt:solid windowtext .75pt;padding:0cm 5.35pt 0cm 5.35pt= '> 100<o:p></o:p><= /p> </td> <td width=3D64 style=3D'width:48.2pt;border-top:none;border-left:none;bor= der-bottom: solid windowtext 1.0pt;border-right:solid windowtext 1.0pt;mso-border-bot= tom-alt: solid windowtext .75pt;mso-border-right-alt:solid windowtext .75pt; padding:0cm 5.35pt 0cm 5.35pt'> 100<o:p></o:p><= /p> </td> </tr> </table> </div> *Number of symptomless p= lants / total number of tested plants x 100 (%).<o:p></o:p> <sp= an lang=3DEN-US style=3D'mso-bidi-font-size:12.0pt;font-family:"Times New Roma= n"; mso-ansi-language:EN-US;font-weight:normal'><o:p> </o:p> The plants infected only with t= he one of the fourth vaccine as well as the control untreated plants remained symptomless to the experiment end. Up to 15th dpi all plants infected only with pathogenic strains developed mosaic symptoms. The percen= tage of symptomless plants inoculated initially with vaccine and after that with pathogenic strain varied in quite wide range. Only single mosaic plants were observed in a part of the variants where the protective properties of both vaccines and D52-V were tested. In analogous variants of TMV D52, the lowest portion of symptomless plants was found. <o:p></o:p> D52-V had very good vaccine activity and the symptomless plants percentage varied from 85 to 100 %= . In variants when the challenge strains were TMV, D52-V showed better vaccine properties than the others, with exception of the variant B-5+U1 where it w= as smaller and of the variant V-69+B where the values were equal. There were l= east symptomless plants in the combination D-52-V+ No.29, in which the pathogenic strain was TMV isolated from pepper. <o:p></o:p> From the obtained results it co= uld be concluded that D52 and D52-V protected tomato against pathogenic TMV and ToMV strains originated from different cultures, as TMV D52 gave way to ToM= V strains V-69 and B-<st1:metricconverter ProductID=3D"5 in" w:st=3D"on">5 in</st1:me= tricconverter> vaccine properties while ToMV D52-V equalized with them. This fact complete= ly correlates with the literature data that the effectiveness of the protection depends on whether the strain is isolated from the plant species it have to= protect, which presume also a good adaptation to it [9, 12, 18].<o:p></o:p></= p> The “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> strains p= ropagate difficult in tolerant to tobamoviruses tomato and accumulate in low concentrations, and in some cases they can not multiply in them at all [22, 28]. Hence, these viruses could not possess protective properties against pathogenic “<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1= ”</st1:metricconverter> strains in tolerant tomato. In order to be proved that D52-V is from group “<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1”</st= 1:metricconverter>, its vaccine action was compared with that of strain V-69 which was from the same group as well as with strain B-5 that multiplied systemically in toler= ant tomato but was from group “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> [24]. The experiment was carried= out with the tolerant tomato cv. Mobaci and pathogenic “<st1:metricconver= ter ProductID=3D"1”" w:st=3D"on">1”</st1:metricconverter> strains w= ere only used (Fig. 4). The percentage of the symptomless plants was 92.5-100 %= for V-69 and 95-100 % for D52-V. The differences were small and the deviat= ions were in favor to the one as well as to the second vaccine. In the same time, only single symptomless plants were established in the variant vaccinated w= ith B-5 although the strain exhibited excellent protective effect in susceptible tomato cultivars.<o:p></o:p> <o:p> </o:p> <![if !vml]><img width=3D579 height=3D272 src=3D"JCC0549NB.files/image008.gif" v:shapes=3D"_x0000_i1028"><![endif]></= p> F= ig. 4= . Vaccine properties of V-69 and 52-V in tolerant to tobamoviruses tomato cv. Mobaci (Tm-1). The vaccine strain B-5 from group “<st1:metricconverter ProductID=3D"0”" w:st=3D"on">0”</st1:metricconverter> is presen= ted for comparison.<o:p></o:p> <o:p> </o:p> These results allow considering that isolate D52-V is from group “<st1:metricconverter ProductID=3D"1= ”" w:st=3D"on">1”</st1:metricconverter> and could be successfully applie= d as vaccine in susceptible and tolerant to ToMV tomato.<o:p></o:p> It is known that the capsid pro= tein is responsible for the appearance of hypersensitive reaction in tobacco with N’ gene resulting in death of the infected cells [1, 2, 3, 11, 25]. Consequently, it could be supposed t= hat all isolates obtained after the passages in tolerant tomato consist of viri= ons, in which there is mutation(s) in the capsid protein. It simultaneously lead= s to changes in the Rf and serological properties of the virus as well as to necrotic local lesion appearance on the inoculated leaf of tobacco with N’ gene.<o:p></o:p> The ability of normal virus propagation in tomato with tolerance Tm-1 gene is connected with amino acid substitutions in the RNA-dependent RNA polymerases [6, 13, 28]. Probably isolate D52-V consists of virions possessing the respective mutation as it = is from group “<st1:metricconverter ProductID=3D"1”" w:st=3D"on">1= ”</st1:metricconverter>. It is established that atenuated and protective properties of ToMV strains = are related with mutations in the virus RNA encoding the same proteins [10]. Therefore, it could be assumed that D52 and D52-V distinguish from the typi= cal pathogenic strains from groups “<st1:metricconverter ProductID=3D"0&#= 8221;" w:st=3D"on">0”</st1:metricconverter> and ‘1” in amino acid alterations again in the RNA-dependent RNA polymerases. The occurred change= s in the respective genes as a result of the passages in tolerant tomato do not concern these regions coding for attenuated properties of D52. The replacin= g of TMV D52 with ToMV isolate D52-I after the first passage and the subsequent mutations selection, resulting in the multiplication adaptation of the virus population in tolerant tomato, leads also in improving the vaccine properti= es of D52-V in comparison with D52. <o:p></o:p> The obtained isolate D52-V protecting excellent susceptible and tolerant tomato against pathogenic tobamovirus strains is v= ery stable because pathogenic virions are not found in its preparations. In this characteristic it exceeds known and widely used in practice vaccines [16, 2= 4]. At the same time, it is symptomless for pepper, a culture often growing in adjacency of tomato. That is why, the vaccinated with D52-V tomato could no= t be virus source for tobacco mosaic disease in pepper cultivars susceptible to tobamoviruses. Consequently, the objections against using vaccine strains do not referred to D52-V that makes it appropriate for application in the production of green house and field tomato. <o:p></o:p> <o:p> </o:p> References<o:p></o:p= > <o:p> </o:p> <![if !supportLists]>1. <![endif]>Culver, J. N., W.O. Dawson, 1991. Mol. Plant-Microbe Interact., 4, 458-463. <o:= p></o:p> <![if !supportLists]><s= pan style=3D'mso-list:Ignore'>2.&n= bsp; <![endif]>Culver, J.N., G. C. Lindbeck, W.O. Dawson, 1991. Ann. Rev. Phytopathol., 29, 193-217. <o:p></o:p> <![if !supportLists]>3. <![endif]>Culver, J.N., G. Stu= bbs, W.O. Dawson, 1994. J. Mol. Biol., = 242, 130-138.<o:p></o:p> <![if !supportLists]>4. <![endif]>Garcia-Lugue, I., M.= T. Serra, E. Alonso, B. Wicke, M. L. Ferrero, J. R. Diaz-Ruiz, 1990. J. Phytopathology, 129, 1-8. <o:p></o:p> <![if !supportLists]>5. <![endif]>Gooding, G. V., T. T. Hebert, 1967. Phytopathology, 61, 943-944.<o:p></o:p> <![if !s= upportLists]>6. <![endif]>Hamamoto, H., Y. 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CMI/AAB Descriptions = of plant viruses, No. 151.<o:= p></o:p> </div> </body> </html> ------=_NextPart_01C5DDFE.C36A4340 Content-Location: file:///C:/236AEE02/JCC0549NB.files/header.htm Content-Transfer-Encoding: quoted-printable Content-Type: text/html; charset="us-ascii" <html xmlns:v=3D"urn:schemas-microsoft-com:vml" xmlns:o=3D"urn:schemas-microsoft-com:office:office" xmlns:w=3D"urn:schemas-microsoft-com:office:word" xmlns:st1=3D"urn:schemas-microsoft-com:office:smarttags" xmlns=3D"http://www.w3.org/TR/REC-html40"> <head> <meta http-equiv=3DContent-Type content=3D"text/html; charset=3Dus-ascii"> <meta name=3DProgId content=3DWord.Document> <meta name=3DGenerator content=3D"Microsoft Word 11"> <meta name=3DOriginator content=3D"Microsoft Word 11"> <link id=3DMain-File rel=3DMain-File href=3D"../JCC0549NB.htm"> <![if IE]> <base href=3D"file:///C:\236AEE02\JCC0549NB.files\header.htm" id=3D"webarch_temp_base_tag"> <![endif]><o:SmartTagType 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