Resistance-breaking Isolates Research Articles

Inter- and Intra-Pathogen Interactions Emanating from Coinfection with Different Fungal and Viral strains in Canola Cultivars with Differing Host Resistances.

Few recent investigations examine coinfection interactions between fungal and viral plant pathogens. Here, we investigated coinfections between Leptosphaeria maculans and turnip mosaic virus (TuMV) in canola (Brassica napus). Different combinations of L. maculans isolate P11 and resistance breaking isolates L. maculans UWA192 and TuMV 12.1, were inoculated to three cultivars with differing pathogen resistances/susceptibilities. They were inoculated first to entire or half cotyledons 10-12 days after emergence, and second to opposite entire or half cotyledons on the same day (day 0) or 3 or 7 days afterwards. The parameters measured were L. maculans cotyledon disease index (%CDI), and TuMV systemically infected leaf symptom intensity (SI) and virus concentration (VC). Except when both day 0 inoculations were with isolate UWA192, %CDI values were supressed strongly or only weakly when isolates P11 and/or UWA192 were inoculated to plants with L. maculans single gene resistance (SGR) or polygenic resistance, respectively. However, except when isolate P11 was inoculated first and UWA192 second, these values declined after inoculation day 0 when SGR was absent. TuMV infection suppressed %CDI values, although this decrease was usually smaller following day 0 half cotyledon inoculations. When TuMV temperature sensitive extreme resistance was present and both inoculations were with TuMV, SI and VC values diminished greatly. However, the extent of this decrease was reduced when second inoculations were with L. maculans. SI and VC values were also smaller when SGR was present and second inoculations were with L. maculans. When L. maculans resistance was lacking, SI and VC values were smaller when second inoculations to entire cotyledons were with L. maculans rather than TuMV. This also occurred after second half cotyledon inoculations with isolate P11 but not isolate UWA192. Therefore, diverse inter- or intra-pathogen interactions developed depending upon host resistance, isolate combination, cotyledon inoculation approach and second inoculation timing.

A Comparison Between Meloidogyne floridensis and M. incognita from California on Susceptible and Resistant Sweetpotato Cultivars.

The reproduction and ability to cause root-galling of a California isolate of the peach root-knot nematode Meloidogyne floridensis was evaluated on seven sweetpotato (Ipomea batatas) cultivars and compared with an M. incognita race 3 and an M. incognita Mi-gene resistance-breaking isolate. The susceptible tomato (Solanum lycopersicum) cultivar Daniela and the Mi-gene-carrying resistant cultivar Celebrity were included as controls. Repeated trials were done in pots in a nematode-quarantine greenhouse at the University of California, Riverside. The three Meloidogyne isolates reproduced equally well on susceptible tomato. On Mi-gene resistant tomato, the reproduction and root-galling by M. floridensis was intermediate between the avirulent M. incognita race 3 and the resistance-breaking M. incognita isolate. The sweetpotato cultivars 'Beauregard' and 'Diane' were excellent hosts for all three Meloidogyne isolates. Cultivars Bellevue, Burgundy, and Covington were resistant to these isolates. The cultivars Bonita and Murasaki-29 were hosts for the M. floridensis and the resistance-breaking M. incognita isolate, which allowed an increase in nematode levels, but they were poor hosts, resulting in a decrease in nematode levels for the M. incognita race 3 isolate. The study showed that M. floridensis can reproduce on tomato and some sweetpotato cultivars that are considered resistant to M. incognita.

Disease Progress and Detection of a California Resistance-Breaking Strain of Tomato Spotted Wilt Virus in Tomato with LAMP and CRISPR-Cas12a Assays

Use of tomato cultivars with the Sw-5 resistance gene cluster has led to the occurrence of resistance-breaking (RB) tomato spotted wilt virus (TSWV) strains globally, including California and, recently, North Carolina and Texas. We documented disease on tomato infected with either an RB strain from California (CA-RB) or a wild type (CA-WT) strain of TSWV on tomato with (cultivar Mountain Merit) or without (cultivar Mountain Fresh Plus) the Sw-5b resistance gene and detected virus incidence over time using microneedle RNA extractions and LAMP. We developed a LAMP/Cas12a assay for detection of the CA-C118Y mutation in a CA-RB strain and tested the assay with field samples. Disease in the susceptible cultivar was less severe with CA-RB than with the CA-WT strain. In contrast, the resistant cultivar had little disease when inoculated with the CA-WT strain but exhibited stunting of greater than 50% when inoculated with the CA-RB strain. In the susceptible tomatoes, the detection rates over time by LAMP reaction were higher in CA-WT than in CA-RB-inoculated plants. In resistant tomato, CA-RB remained detectable by TSWV LAMP over 14 days, whereas the WT strain was undetectable. A two-step LAMP/Cas12a assay differentiated the two strains in 1 h. Our methods were validated with samples from TSWV-infected North Carolina fields. A phylogeny of NSm gene sequences that included North Carolina field samples revealed two independent origins of the North Carolina RB isolates. The LAMP/Cas12 assay showed excellent detection of the CA-C118Y mutation. The TSWV LAMP/Cas12a assay is adaptable for in-field applications on either a smart phone platform or heat block. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

Inheritance of Partial Resistance to Isolate VdLs17 of Verticillium dahliae Within Lactuca spp.

Lettuce (Lactuca sativa L.) production is greatly threatened by Verticillium wilt, which is caused by three pathogenic races (races 1, 2, and 3) of the soilborne fungus Verticillium dahliae. Race 1 is predominant, and resistant varieties that provide full protection against it are commercially available. However, heavily relying on race 1-resistant cultivars could shift the population towards resistance-breaking isolates and impact the durability of plant resistance. This study determined the inheritance of partial resistance to isolate VdLs17 of V. dahliae within Lactuca spp. using 258 F2:3 progeny generated from a cross between two partially resistant accessions, 11G99 (L. serriola) and PI 171674 (L. sativa). Eight experiments were performed under greenhouse and growth room conditions across 3 years using a randomized complete block design, and segregation analysis was conducted to determine the inheritance pattern. The results indicate that partial resistance to isolate VdLs17 of V. dahliae is conditioned by a two-major-gene genetic model with additive-dominance-epistatic effects. Transgressive segregants were infrequent but observed in both directions, indicating that favorable and adverse alleles are dispersed in both parents. Combining favorable alleles of these two partially resistant parents appears to be challenging because of epistatic effects and a significant role of environment in disease severity. The probability of capturing favorable additive genes could be maximized by generating and evaluating a large population and making selections at late generations. This study provides valuable insights into the inheritance pattern of partial resistance to isolate VdLs17 of V. dahliae that will be helpful in designing efficient breeding strategies in lettuce.[Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

A review of sources of resistance to turnip yellows virus (TuYV) in Brassica species.

Turnip yellows virus (TuYV; previously known as beet western yellows virus) causes major diseases of Brassica species worldwide resulting in severe yield-losses in arable and vegetable crops. It has also been shown to reduce the quality of vegetables, particularly cabbage where it causes tip burn. Incidences of 100% have been recorded in commercial crops of winter oilseed rape (Brassica napus) and vegetable crops (particularly Brassica oleracea) in Europe. This review summarises the known sources of resistance to TuYV in B. napus (AACC genome), Brassica rapa (AA genome) and B. oleracea (CC genome). It also proposes names for the quantitative trait loci (QTLs) responsible for the resistances, Turnip Yellows virus Resistance (TuYR), that have been mapped to at least the chromosome level in the different Brassica species. There is currently only one known source of resistance deployed commercially (TuYR1). This resistance is said to have originated in B. rapa and was introgressed into the A genome of oilseed rape via hybridisation with B. oleracea to produce allotetraploid (AACC) plants that were then backcrossed into oilseed rape. It has been utilised in the majority of known TuYV-resistant oilseed rape varieties. This has placed significant selection pressure for resistance-breaking mutations arising in TuYV. Further QTLs for resistance to TuYV (TuYR2-TuYR9) have been mapped in the genomes of B. napus, B. rapa and B. oleracea and are described here. QTLs from the latter two species have been introgressed into allotetraploid plants, providing for the first time, combined resistance from both the A and the C genomes for deployment in oilseed rape. Introgression of these new resistances into commercial oilseed rape and vegetable brassicas can be accelerated using the molecular markers that have been developed. The deployment of these resistances should lessen selection pressure for resistance-breaking isolates of TuYV and thereby prolong the effectiveness of each other and extant resistance.

A novel tomato spotted wilt virus isolate encoding a noncanonical NSm C118F substitution associated with Sw-5 tomato gene resistance breaking.

The nonstructural protein NSm of tomato spotted wilt virus (TSWV) has been identified as the avirulence determinant of the tomato single dominant Sw-5 resistance gene. Although Sw-5 effectiveness has been shown for most TSWV isolates, the emergence of resistance-breaking (RB) isolates has been observed. It is strongly associated with two point mutations (C118Y or T120N) in the NSm viral protein. TSWV-like symptoms were observed in tomato crop cultivars (+Sw-5) in the Baja California peninsula, Mexico, and molecular methods confirmed the presence of TSWV. Sequence analysis of the NSm 118-120 motif and three-dimensional protein modelling exhibited a noncanonical C118F substitution in seven isolates, suggesting that this substitution could emulate the C118Y-related RB phenotype. Furthermore, phylogenetic and molecular analysis of the full-length genome (TSWV-MX) revealed its reassortment-related evolution and confirmed that putative RB-related features are restricted to the NSm protein. Biological and mutational NSm 118 residue assays in tomato (+Sw-5) confirmed the RB nature of TSWV-MX isolate, and the F118 residue plays a critical role in the RB phenotype. The discovery of a novel TSWV-RB Mexican isolate with the presence of C118F substitution highlights a not previously described viral adaptation in the genus Orthotospovirus, and hence, the necessity of further crop monitoring to alert the establishment of novel RB isolates in cultivated tomatoes.

Biological and molecular characterization of a resistance-breaking isolate of citrus tristeza virus from Uruguay and its effects on Poncirus trifoliata growth performance

Resistance-breaking (RB) isolates of citrus tristeza virus (CTV) can replicate and move systemically in Poncirus trifoliata, a rootstock widely used for management of decline caused by CTV and other purposes. In Uruguay, severe CTV isolates are prevalent, and an RB isolate (designated as RB-UY1) was identified. In order to predict the implications of this genotype circulating in citrus crops grafted on trifoliate rootstocks, the aim of this work was to determine the biological and molecular characteristics of this isolate, the efficiency of its transmission by Toxoptera citricida, and its effects on plant growth performance of P. trifoliata. Our results show that RB-UY1 can be classified as a mild isolate, that it is phylogenetically associated with the RB1 group, and that it is efficiently transmitted by T. citrida. They also suggest that the RB-UY1 isolate should not affect the performance of citrus crops grafted on trifoliate rootstocks, although some growth parameters of P. trifoliata seedlings were affected four years after inoculation.

Tomato Mi-gene Resistance-Breaking Populations of Meloidogyne Show Variable Reproduction on Susceptible and Resistant Crop Cultivars.

Sixteen Meloidogyne isolates from tomato fields in California grown with resistant cultivars were multiplied on resistant tomato in a greenhouse. Of these resistance-breaking isolates, one was identified as M. javanica, and all others as M. incognita. The reproduction of the M. javanica isolate and four M. incognita isolates on six resistant tomato cultivars and on susceptible and resistant cultivars of pepper, sweetpotato, green bean, cotton, and cowpea was evaluated and compared to an avirulent M. incognita population in greenhouse pot trials. On resistant tomato cultivars, there were minor but significant differences between the resistance-breaking Meloidogyne isolates and between the different tomato cultivars. Of the other resistant crop cultivars, pepper was resistant to all isolates and green bean to all M. incognita isolates, while cotton and cowpea allowed reproduction of one of the resistance-breaking M. incognita isolates. The resistant sweetpotato cv. Bonita behaved like resistant tomato, allowing reproduction of all five resistance-breaking isolates but not of the avirulent M. incognita. Our results showed that variability exists among resistance-breaking Meloidogyne isolates, and that isolates overcoming resistance in tomato may also be virulent on resistant sweetpotato.

Identification and genetic inheritance of a new source of broad-spectrum extreme resistance to turnip mosaic virus (TuMV) in Brassica rapa

Brassica rapa line K185 was identified with broad-spectrum extreme resistance to turnip mosaic virus (TuMV). The resistance was effective against TuMV isolates UK 1, CZE 1, GBR 6, POL 1 and CDN 1 (representing major pathotypes 1, 3 and 4 and major resistance-breaking isolates) following mechanical inoculation. F1 plants from a cross between K185 and the rapid-cycling B. rapa ssp. trilocularis line R-o-18 (uniformly susceptible to all the above TuMV isolates), were resistant following challenge with TuMV isolates CDN 1 and GBR 6, indicating the involvement of dominant gene(s). F2 plants derived from a single F1, CDN 1- and GBR 6-resistant plant segregated for resistance to TuMV isolate GBR 6. The segregation ratio of resistant to susceptible plants was consistent with at least two loci controlling resistance and with both loci having dominant alleles for resistance. Other sources of broad-spectrum resistance to TuMV have been shown to involve the recessive allele retr01. The broad-spectrum resistance in K185 clearly involves different alleles to retr01 and provides the opportunity to introgress an alternative form of TuMV resistance into commercial B. rapa lines and thereby reduce the selection pressure for retr01 resistance breaking.

Specificity of Resistance and Tolerance to Cucumber Vein Yellowing Virus in Melon Accessions and Resistance Breaking with a Single Mutation in VPg.

Cucumber vein yellowing virus (CVYV) is an emerging virus on cucurbits in the Mediterranean Basin, against which few resistance sources are available, particularly in melon. The melon accession PI 164323 displays complete resistance to isolate CVYV-Esp, and accession HSD 2458 presents a tolerance, i.e., very mild symptoms despite virus accumulation in inoculated plants. The resistance is controlled by a dominant allele Cvy-11, while the tolerance is controlled by a recessive allele cvy-2, independent from Cvy-11. Before introducing the resistance or tolerance in commercial cultivars through a long breeding process, it is important to estimate their specificity and durability. Upon inoculation with eight molecularly diverse CVYV isolates, the resistance was found to be isolate-specific because many CVYV isolates induced necrosis on PI 164323, whereas the tolerance presented a broader range. A resistance-breaking isolate inducing severe mosaic on PI 164323 was obtained. This isolate differed from the parental strain by a single amino acid change in the VPg coding region. An infectious CVYV cDNA clone was obtained, and the effect of the mutation in the VPg cistron on resistance to PI 164323 was confirmed by reverse genetics. This represents the first determinant for resistance-breaking in an ipomovirus. Our results indicate that the use of the Cvy-11 allele alone will not provide durable resistance to CVYV and that, if used in the field, it should be combined with other control methods such as cultural practices and pyramiding of resistance genes to achieve long-lasting resistance against CVYV.

Determination of key residues in tospoviral NSm required for Sw-5b recognition, their potential ability to overcome resistance, and the effective resistance provided by improved Sw-5b mutants.

Sw‐5b is an effective resistance gene used widely in tomato to control tomato spotted wilt virus (TSWV), which causes severe losses in crops worldwide. Sw‐5b confers resistance by recognizing a 21‐amino‐acid peptide region of the viral movement protein NSm (NSm21, amino acids 115–135). However, C118Y or T120N mutation within this peptide region of NSm has given rise to field resistance‐breaking (RB) TSWV isolates. To investigate the potential ability of TSWV to break Sw‐5b‐mediated resistance, we mutagenized each amino acid on NSm21 and determined which amino acid mutations would evade Sw‐5b recognition. Among all alanine‐scan mutants, NSmP119A, NSmW121A, NSmD122A, NSmR124A, and NSmQ126A failed to induce a hypersensitive response (HR) when coexpressed with Sw‐5b in Nicotiana benthamiana leaves. TSWV with the NSmP119A, NSmW121A, or NSmQ126A mutation was defective in viral cell‐to‐cell movement and systemic infection, while TSWV carrying the NSmD122A or NSmR124A mutation was not only able to infect wild‐type N. benthamiana plants systemically but also able to break Sw‐5b‐mediated resistance and establish systemic infection on Sw‐5b‐transgenic N. benthamiana plants. Two improved mutants, Sw‐5bL33P/K319E/R927A and Sw‐5bL33P/K319E/R927Q , which we recently engineered and which provide effective resistance against field RB isolates carrying NSmC118Y or NSmT120N mutations, recognized all NSm21 alanine‐substitution mutants and conferred effective resistance against new experimental RB TSWV with the NSmD122A or NSmR124A mutation. Collectively, we determined the key residues of NSm for Sw‐5b recognition, investigated their potential RB ability, and demonstrated that the improved Sw‐5b mutants could provide effective resistance to both field and potential RB TSWV isolates.

Full Genome Evolutionary Studies of Wheat Streak Mosaic-Associated Viruses Using High-Throughput Sequencing.

Wheat streak mosaic (WSM), a viral disease affecting cereals and grasses, causes substantial losses in crop yields. Wheat streak mosaic virus (WSMV) is the main causal agent of the complex, but mixed infections with Triticum mosaic virus (TriMV) and High plains wheat mosaic emaravirus (HPWMoV) were reported as well. Although resistant varieties are effective for the disease control, a WSMV resistance-breaking isolate and several potential resistance-breaking isolates have been reported, suggesting that viral populations are genetically diverse. Previous phylogenetic studies of WSMV were conducted by focusing only on the virus coat protein (CP) sequence, while there is no such study for either TriMV or HPWMoV. Here, we studied the genetic variation and evolutionary mechanisms of natural populations of WSM-associated viruses mainly in Kansas fields and fields in some other parts of the Great Plains using high-throughput RNA sequencing. In total, 28 historic and field samples were used for total RNA sequencing to obtain full genome sequences of WSM-associated viruses. Field survey results showed WSMV as the predominant virus followed by mixed infections of WSMV + TriMV. Phylogenetic analyses of the full genome sequences demonstrated that WSMV Kansas isolates are widely distributed in sub-clades. In contrast, phylogenetic analyses for TriMV isolates showed no significant diversity. Recombination was identified as the major evolutionary force of WSMV and TriMV variation in KS fields, and positive selection was detected in some encoding genomic regions in the genome of both viruses. Furthermore, the full genome sequence of a second Kansas HPWMoV isolate was reported. Here, we also identified previously unknown WSMV isolates in the Great Plains sharing clades and high nucleotide sequence similarities with Central Europe isolates. The findings of this study will provide more insights into the genetic structure of WSM-associated viruses and, in turn, help in improving strategies for disease management.

Serological, molecular and phylogenetic analysis of Rice Yellow Mottle Virus (RYMV) isolates collected in Southern-Benin

The rice yellow mottle is the best known viral disease of rice in Africa. It causes significant economic losses in farmer’s fields. The serological properties of Rice Yellow Mottle Virus (RYMV) isolates, collected in Southern Benin, were assessed by immunological tests with polyclonal and monoclonal antibodies (Pabs and MAbs). The isolates CP (Capsid protein) portions, obtained by RT-PCR, were sequenced and their amino-acids sequences were analyzed and used for phylogenetic analysis. Three different RYMV pathogenic groups, including three resistance breaking (RB) isolates (Be20, Be21 and Be27) which over came allele rymv1-5gene, were identified. Two serotypes Ser1/strain S1 and Ser2/ strain S2 were also distinguished. The molecular properties of the isolates CP gene and the phylogenetic characteristics indicated that the Southern Benin RYMV strain is heterogeneous. The strain S1-Benin linked to the West Central African lineage(S1-WCA) is related to strain from Togo and Niger, whereas, the strain S2 Benin is the stumps of the West African lineage (S1-WA) and is related to Mali, Burkina Faso and Ivory Coast RYMV strains. These results reported the virus pathogenicity level and showed the relationships between RYMV strains in all Dahomey gap countries and in West and Central Africa. Keywords: serotypes, RYMV strains, RB isolates and phylogenetic analysis

Stepwise artificial evolution of an Sw-5b immune receptor extends its resistance spectrum against resistance-breaking isolates of Tomato spotted wilt virus.

SummaryPlants use intracellular nucleotide‐binding leucine‐rich repeat immune receptors (NLRs) to recognize pathogen‐encoded effectors and initiate immune responses. Tomato spotted wilt virus (TSWV), which has been found to infect >1000 plant species, is among the most destructive plant viruses worldwide. The Sw‐5b is the most effective and widely used resistance gene in tomato breeding to control TSWV. However, broad application of tomato cultivars carrying Sw‐5b has resulted in an emergence of resistance‐breaking (RB) TSWV. Therefore, new effective genes are urgently needed to prevent further RB TSWV outbreaks. In this study, we conducted artificial evolution to select Sw‐5b mutants that could extend the resistance spectrum against TSWV RB isolates. Unlike regular NLRs, Sw‐5b detects viral elicitor NSm using both the N‐terminal Solanaceae‐specific domain (SD) and the C‐terminal LRR domain in a two‐step recognition process. Our attempts to select gain‐of‐function mutants by random mutagenesis involving either the SD or the LRR of Sw‐5b failed; therefore, we adopted a stepwise strategy, first introducing a NSmRB‐responsive mutation at the R927 residue in the LRR, followed by random mutagenesis involving the Sw‐5b SD domain. Using this strategy, we obtained Sw‐5bL33P/K319E/R927A and Sw‐5bL33P/K319E/R927Q mutants, which are effective against TSWV RB carrying the NSmC118Y or NSmT120N mutation, and against other American‐type tospoviruses. Thus, we were able to extend the resistance spectrum of Sw‐5b; the selected Sw‐5b mutants will provide new gene resources to control RB TSWV.

Identification of new sources of resistance to resistance-breaking isolates of tomato spotted wilt virus.

The tomato as both a fresh consumption and industrial product is one of the most profitable vegetables and has a large cultivation area in the world. Parallel to intense production activities, Tomato Spotted Wilt Virus (TSWV), like viral diseases, results in significant economic losses every year. Use of resistant cultivars is the most efficient and environmental-friendly method of fighting against these diseases. This study was conducted to develop new tomato genetic resources resistant to TSWV because of the Sw-5 resistance breaking (RB) isolates that were determined in tomato cultivation areas. In this study, a total of 40 tomato materials including 15 lines, 9 commercial varieties and 16 wild genotypes were by tested with molecular and biological testing methods. Mechanical inoculation method was used for biological testing and SCAR marker was used in molecular analysis. S. penellii, S. chmielewski, S. habrochaites, S. peruvianum and S. sitiens, LA0716, LA1028, LA1777, LA2744 and LA4110 genotypes were found as resistant against breaking isolates of Tomato Spotted Wilt Virus. These genotypes may be a good resistance source for the future breeding studies in tomato.

Susceptibility of Flordaguard peach rootstock to a resistant-breaking population of Meloidogyne floridensis and two populations of Meloidogyne arenaria.

Cultivar Flordaguard is suggested as a root-knot nematode (RKN) resistant rootstock for Florida peaches, however, RKN disease has been observed on this rootstock in peach orchards. Our goal was to confirm whether the RKN resistance breaking isolates of M. floridensis and M. arenaria indeed could infect and reproduce on the peach rootstock cv. Flordaguard in both laboratory and field studies. Root galling occurred on all peach cultivars evaluated including Flordaguard, Flordaglo, Okinawa, and Lovell, in the presence of the RKN resistance-breaking isolates of M. floridensis (MfGnv14) and two M. arenaria isolates (Ma1 and Ma2). These rootstocks showed varying degrees of susceptibility (to a lesser extent in Okinawa) to these three RKN resistance-breaking isolates. The importance of nematode inoculum concentrations in differentiating between resistance and susceptible plants was demonstrated, and thus are an important factor to consider in nematode resistance breeding programs. In host differential tests the peach-originated isolates of M. floridensis and M. arenaria behaved similarly with the vegetable-originated isolates of M. floridensis on tomato, peanut, watermelon, and tobacco, but showed variable host responses on cotton and pepper. The two M. arenaria isolates from peach reproduced on pepper but not on peanut. To our knowledge this is the first report of M. arenaria race 3 infecting Flordaguard and pepper in Florida. Soil and root samples collected from cv. Flordaguard infected trees at two commercial peach orchards showed that M. floridensis and M. arenaria were established on the rootstock.

Resistance to the wheat curl mite and mite-transmitted viruses: challenges and future directions.

Wheat curl mite (WCM) is the only known arthropod vector of four wheat viruses, the most important of which is Wheat streak mosaic virus (WSMV). Host resistance to WCM and WSMV is limited to a small number of loci, most of which are introgressed from wild relatives and are often associated with linkage drag and temperature sensitivity. Reports of virulent WCM populations and potential resistance-breaking WSMV isolates highlight the need for more diverse sources of resistance. Genome sequencing will be critical to fully characterize the genetic diversity in WCM and WSMV populations to better understand the incidence of WCM-transmitted viruses and to evaluate the potential stability of resistance genes. Characterizing host resistance genes will help build a mechanistic understanding of wheat-WCM-WSMV interactions and inform strategies to identify and engineer more durable resistance sources.

Structural basis of nanobody recognition of grapevine fanleaf virus and of virus resistance loss

Grapevine fanleaf virus (GFLV) is a picorna-like plant virus transmitted by nematodes that affects vineyards worldwide. Nanobody (Nb)-mediated resistance against GFLV has been created recently, and shown to be highly effective in plants, including grapevine, but the underlying mechanism is unknown. Here we present the high-resolution cryo electron microscopy structure of the GFLV-Nb23 complex, which provides the basis for molecular recognition by the Nb. The structure reveals a composite binding site bridging over three domains of one capsid protein (CP) monomer. The structure provides a precise mapping of the Nb23 epitope on the GFLV capsid in which the antigen loop is accommodated through an induced-fit mechanism. Moreover, we uncover and characterize several resistance-breaking GFLV isolates with amino acids mapping within this epitope, including C-terminal extensions of the CP, which would sterically interfere with Nb binding. Escape variants with such extended CP fail to be transmitted by nematodes linking Nb-mediated resistance to vector transmission. Together, these data provide insights into the molecular mechanism of Nb23-mediated recognition of GFLV and of virus resistance loss.

Novel Diversity and Virulence Patterns Found in New Isolates of Cydia pomonella Granulovirus from China.

Cydia pomonella granulovirus (CpGV) is successfully used worldwide as a biocontrol agent of the codling moth (CM) (Cydia pomonella). The occurrence of CM populations with different modes of resistance against commercial CpGV preparations in Europe, as well as the invasiveness of CM in China, threatening major apple production areas there, requires the development of new control options. Utilizing the naturally occurring genetic diversity of CpGV can improve such control strategies. Here, we report the identification of seven new CpGV isolates that were collected from infected CM larvae in northwest China. Resistance testing using a discriminating CpGV concentration and the determination of the median lethal concentration (LC50) were performed to characterize their levels of virulence against susceptible and resistant CM larvae. The isolates were further screened for the presence of the 2 × 12-bp-repeat insertion in CpGV gene pe38 (open reading frame 24 [ORF24]), which was shown to be the target of type I resistance. It was found that three isolates, CpGV-JQ, -KS1, and -ZY2, could break type I resistance, although delayed mortality was observed in the infection process. All isolates followed the pe38 model of breaking type I resistance, except for CpGV-WW, which harbored the genetic factor but failed to overcome type I resistance. However, CpGV-WW was able to overcome type II and type III resistance. The bioassay results and sequencing data of pe38 support previous findings that pe38 is the major target for type I resistance. The new isolates show some distinct virulence characteristics when infection of different CM strains is considered.IMPORTANCE CpGV is a highly virulent pathogen of the codling moth (CM). It is registered and widely applied as a biocontrol agent in nearly all apple-growing countries worldwide. The emergence of CpGV resistance and the increasing lack of chemical control options require improvements to current control strategies. Natural CpGV isolates, as well as resistance-breaking isolates selected in resistant CM strains, have provided resources for improved resistance-breaking CpGV products. Here, we report novel CpGV isolates collected in China, which have new resistance-breaking capacities and may be an important asset for future application in the biological control of codling moths.

Phenotypic characterization of Tomato spotted wilt virus resistance breaking isolate in tomatoes

Domateste lekeli solgunluk virüsü (Tomato spotted wilt virus-TSWV), dünya genelinde ekonomik anlamda büyük kayıplar meydana getiren virüslerden biridir. Son zamanlara kadar domateslerde Sw-5 geni vasıtasıyla bu hastalık ile mücadele edilmekteydi. 2016-2019 yılları arasında Antalya ili ve ilçelerinde domates yetiştiriciliği yapılan seralarda Sw-5 geni barındıran çeşitler üzerinde TSWV’a ait simptomların geliştiği gözlemlenmiştir. Bu izolatların PCR çalışmaları ile bitkilerin Sw-5 geni içerdiği ve RT-PCR (Revers-Transkriptaz Polimeraz Zincir Reaksiyonu) çalışmaları ile de simptomların TSWV’a ait olduğu doğrulandıktan sonra izolatın ismi TSWVAntRB olarak belirlenmiştir. Yapılan gözlemlemeler sonucunda TSWVAntRB izolatının farklı simptomolojik karakterlere sahip olduğu tespit edilmiştir. Bu farklılıklarının belirlenebilmesi için TSWVAntRB izolatının, bitkiler üzerine mekanik inokulasyonu 5 tekerrürlü olacak şekilde gerçekleştirilmiştir. Mekanik inokulasyon çalışmalarında; Sw-5 geni içeren 4 adet ticari çeşit, bir adet dayanıklılık kaynağı olarak bilinen Solanum peruvianum ve bir adet hassas çeşit kullanılmıştır. Sw-5 dayanımını kırmayan (NRB) izolatında şimdiye kadar yaprakta sadece nekrotik lekelenmeler meydana gelirken, Sw-5 dayanımını kıran (RB) izolatında yaprak üzerinde de halkalı lekelenmelerin meydana geldiği belirlenmiştir. Aynı zamanda bazı çeşitler üzerinde ilk simptomlar meyve dönemine kadar gizli kaldığı ve meyve salkımında bazen bir veya iki meyve üzerinde simptomlar meydana getirdiği gözlemlenmiştir. Çalışmada simptom şiddetinin hava sıcaklığı ile ilişkili olduğu saptanmış ve TSWVAntRB izolatının simptomolojik özellikleri her açıdan incelenmiştir.