Homozygous Resistance Research Articles

Гибрид подсолнечника Протектор с повышенной устойчивостью к заразихе расы G и вирулентным расам ложной мучнистой росы

Hybrid Protector has both high homozygous resistance to race G of broomrape and genetic resistance to the downy mildew pathogen (races 330, 334, 337, 710, 713, 730, 733, 734, 737) and is intended for cultivation in regions with the prevalence of virulent races of broomrape and downy mildew, as well as for organic farming technology. Protector is an early-maturing, high-yielding sunflower hybrid with a growing period of 104 days before harvesting, oil content of 48% and huskness of 25%. It is genetically close to the hybrid Fakel.

Metabolic Adaptations in Phosphine-Resistant Tribolium castaneum Driven by Mitochondrial Enzyme Variability and Gene Expression.

Phosphine fumigation is essential for controlling storage pests like Tribolium castaneum, but its frequent application has resulted in resistance, primarily due to mutations in the Dihydrolipoamide dehydrogenase (DLD) gene associated with the rph2 allele. This study demonstrates that the Patiala population exhibits homozygous resistance variations across populations, contrasting with the susceptibility observed in laboratory cultures. Our assessment of mitochondrial DLD and Cytochrome c oxidase (COX) activities showed significantly elevated levels in the Patiala population, with increases of approximately sevenfold for DLD and 6.92-fold for COX, indicating mitochondrial adaptations for enhanced energy production. Kinetic analyses of DLD in the resistant Patiala population showed a higher Vmax (0.005 mmol/min) and a significantly increased Km (16.66 mM), indicating variations in maximal enzyme activity and substrate affinity. Furthermore, resistant T. castaneum populations displayed substantial upregulation of DLD and COX gene expression, with DLD expression increasing by 10.53-fold and COX expression peaking at 102.57-fold in Patiala. Pearson correlation analysis indicated strong positive correlations (r > 0.8) between enzymatic activity and gene expression for both DLD and COX, suggesting a coordinated role in resistance mechanisms. The PCA biplot illustrated distribution patterns of enzymatic activity and gene expression among field-resistant populations, highlighting the association between increased resistance and elevated enzymatic activity and gene expression levels. Therefore, the upregulation of DLD and COX activities in resistant populations underscores their critical roles in counteracting phosphine, reflecting metabolic reprogramming for improved energy production under stress.

Introgression of OsAP47 by marker-assisted selection enhanced resistance against southern rice black-streaked dwarf virus disease

Southern rice black-streaked dwarf virus disease (SRBSDVD) is the most destructive viral disease in rice. In order to breeding resistant cultivars, Insertion-Deletion (InDel) markers were developed linked to OsAP47, the first isolated major resistance gene against SRBSDVD. Marker-assisted selection (MAS) was conducted to introduce this gene into the commercial variety. A rice line carrying homozygous resistance allele of OsAP47 was selected and named Kanghei No. 201 (KH201). Evaluated by artificial inoculation, KH201 showed significantly higher resistance than the recurrent parent Suxiu No.867 (SX867). And no significant differences were detected for KH201 in the yield-related components, including spikelets per panicle (SPP), ripened grains per panicle (RGPP), 1000-grain weight (TGW) and panicles per square meter (PPSM), leading to stable theoretical yield. The results indicated that introgression of OsAP47 improved rice resistance and can avoid yield losses produced by SRBSDVD. KH201 was demonstrated as a resistance material that could be used in rice breeding.

Distribution and frequency of ace-1 and kdr mutations of Culex pipiens subgroup in the Republic of Korea

Mosquitoes in the Culex pipiens subgroup are the primary vectors of the West Nile virus. Two members, Culex pallens and Culex pipiens f. molestus, are present in the Republic of Korea (ROK). Because the Culex pipiens subgroup occurs in large amounts, often near human habitation, it is frequently exposed to various insecticides, which is probably responsible for the rapid evolution of insecticide resistance traits. Experiments related to insecticide resistance in the Culex pipiens subgroup conducted in the ROK have been performed without discrimination below the species level. This study categorized Culex pipiens mosquitoes subgroup from 13 regions in the ROK into Culex pallens and Culex pipiens f. molestus, and target site genotypes for acetylcholinesterase-1 (ace-1) and voltage-gated sodium channel (vgsc) genes were identified for each taxon. Screening for ace-1 did not identify a resistant allele (G119S) in Cx. pipiens f. molestus, and heterozygous resistance (AGC/GGC) was identified in one Cx. pallens collected in Mokpo. In vgsc, knockdown resistance (kdr) mutations [TTT(L1014F) and TCA(L1014S)] were present in both taxa, with Cx. pipiens f. molestus having homozygous resistance (TTT/TTT): 44%, heterozygous resistance (TTT/TTA): 28%, and homozygous susceptibility (TTA/TTA): 28%, whereas Cx. pallens showed homozygous resistance (TTT/TTT or TCA/TCA): 26%, heterozygous resistance (TTT/TTA, TTT/TCA, or TCA/TTA): 26%, and homozygous susceptibility (TTA/TTA): 48%. Furthermore, the unique vgsc allele was present in both Cx. pipiens f. molestus and Cx. pallens. This was the first experiment to analyze the Culex pipiens subgroup living in the ROK below the species level, and its results could be used in the future for more detailed mosquito control.

Distribution of Anopheles gambiae thioester-containing protein 1 alleles along malaria transmission gradients in The Gambia

BackgroundThioester-containing protein 1 (TEP1) is a highly polymorphic gene playing an important role in mosquito immunity to parasite development and associated with Anopheles gambiae vectorial competence. Allelic variations in TEP1 could render mosquito either susceptible or resistant to parasite infection. Despite reports of TEP1 genetic variations in An. gambiae, the correlation between TEP1 allelic variants and transmission patterns in malaria endemic settings remains unclear.MethodsTEP1 allelic variants were characterized by PCR from archived genomic DNA of > 1000 An. gambiae mosquitoes collected at 3 time points between 2009 and 2019 from eastern Gambia, where malaria transmission remains moderately high, and western regions with low transmission.ResultsEight common TEP1 allelic variants were identified at varying frequencies in An. gambiae from both transmission settings. These comprised the wild type TEP1, homozygous susceptible genotype, TEP1s; homozygous resistance genotypes: TEP1rA and TEP1rB, and the heterozygous resistance genotypes: TEP1srA, TEP1srB, TEP1rArB and TEP1srArB. There was no significant disproportionate distribution of the TEP1 alleles by transmission setting and the temporal distribution of alleles was also consistent across the transmission settings. TEP1s was the most common in all vector species in both settings (allele frequencies: East = 21.4–68.4%. West = 23.5–67.2%). In Anopheles arabiensis, the frequency of wild type TEP1 and susceptible TEP1s was significantly higher in low transmission setting than in high transmission setting (TEP1: Z = − 4.831, P < 0.0001; TEP1s: Z = − 2.073, P = 0.038).ConclusionsThe distribution of TEP1 allele variants does not distinctly correlate with malaria endemicity pattern in The Gambia. Further studies are needed to understand the link between genetic variations in vector population and transmission pattern in the study settings. Future studies on the implication for targeting TEP1 gene for vector control strategy such as gene drive systems in this settings is also recommended.

Effect of Soil Temperature on Reproduction of Root-knot Nematodes in Flue-cured Tobacco with Homozygous Rk1 and/or Rk2 Resistance Genes.

Most commercial flue-cured tobacco cultivars contain the Rk1 resistance gene, which provides resistance to races 1 and 3 of Meloidogyne incognita and race 1 of M. arenaria. A number of cultivars now possess a second root-knot resistance gene, Rk2. High soil temperatures have been associated with a breakdown of root-knot resistance genes in a number of crops. Three greenhouse trials were performed from 2014 to 2015 investigate the effect of high soil temperature on the efficacy of Rk1 and/or Rk2 genes in reducing parasitism by a population of M. incognita race 3. Trials were arranged in randomized complete block design in open-top growth chambers set at 25°, 30°, and 35°C. Plants were inoculated with 3,000 eggs and data were collected 35 days post-inoculation. Galling, numbers of egg masses and eggs, and reproductive index were compared across cultivar entries. Nematode reproduction was reduced at 25°C and 30°C on entries possessing Rk1 and Rk1Rk2 compared to the susceptible entry and the entry possessing only Rk2. However, there were often no significant differences in reproduction at 35°C between entries with Rk1 and/or Rk2 compared to the susceptible control, indicating an increase of root-knot nematode parasitism on resistant entries at higher temperatures. Although seasonal differences in nematode reproduction were observed among experiments, relative differences among tobacco genotypes remained generally consistent.

Fine mapping and candidate gene analysis of CRA3.7 conferring clubroot resistance in Brassica rapa.

In this study, we fine-mapped a clubroot resistance gene CRA3.7 in Chinese cabbage and developed its closely linked marker syau-InDel3008 for marker-assisted selection in CR cultivars breeding. Chinese cabbage is an important leafy vegetable rich in many nutrients widely grown in China. Clubroot disease caused by an obligate biotrophic pathogen Plasmodiophora brassicae was rapidly spread and challenged to Chinese cabbage production. A clubroot resistance (CR) gene, CRA3.7, was mapped on chromosome A03 of Brassica rapa. A Chinese cabbage line 'CR510', which harbor homozygous resistance locus CRA3.7 was selected from a BC4F3 family. 'CR510' was crossed with a clubroot susceptible Chinese cabbage inbred line '59-1'. Total 51 recombinant plants were identified from an F2 population including 3000 individuals. These recombinants were selfed and the clubroot resistance of F2/3 families was evaluated. Finally, a clubroot resistance gene CRA3.7 was fine-mapped to an interval of approximately 386 kb between marker syau-InDel3024 and syau-InDel3008. According to the reference genome, total 54 genes including five encoding the TIR-NBS-LRR proteins was annotated in the fine-mapped region. Further, nine candidate's gene expression in parental lines at 7, 14 and 21 days after inoculation of P. brassicae were evaluated. Bra019376, Bra019401, Bra019403 and Bra019410 are highly expressed in 'CR510' than '59-1'. Gene sequence of Bra019410 from 'CR510' was cloned and identified different from CRa. Therefore, Bra019376, Bra019401, Bra019403 and Bra019410 are the most likely candidates for CRA3.7. Our research provides a valuable germplasm resource against P. brassicae Pb3 and CRA3.7 closely linked marker for marker-assisted selection in CR cultivars breeding.

Impacts of differential mosquito control treatment regimens on insecticide susceptibility status of Aedes aegypti (Diptera: Culicidae)

Aedes aegypti (Linnaeus) is an invasive mosquito species and notable vector of several pathogens in the USA. Their cryptic and anthropophilic nature puts this species in close association with humans, where they can also be a nuisance. Mosquito control programs are the front line of defense for protecting the community from nuisance-biting and disease. However, the occurrence and prevalence of insecticide resistance in mosquitoes is a well-documented phenomenon that directly impacts the efficacy of insecticide applications. In Florida specifically, widespread resistance in Ae. aegypti has created a need for operational strategies that combat and, ideally, reverse resistance. Laboratory studies and the association between fitness costs and insecticide resistance indicate that this reversion is possible under the right conditions. For a 2.5-year period, the impact of varying operational treatment regimens on insecticide resistance in Ae. aegypti is evaluated using kdr genotyping and the CDC bottle bioassay. In an organophosphate treatment area, a decrease in frequency of a double homozygous resistant genotype was observed. CDC bottle bioassays did not reveal any clear trends in the data to indicate a reversion to insecticide susceptibility. However, the changes in genotype could indicate the first step back to insecticide susceptibility. This study provides preliminary data that has implications for resistance management in mosquito control operations.Article HighlightsPresented study represents the first field study to evaluate how changing insecticide pressures on resistant mosquitoes affects insecticide resistance.A decrease in the double homozygous resistance genotype was observed in Ae. aegypti from the organophosphate treatment area after a change in use patterns.Identifiable trends in phenotypic resistance as a result of the treatment regimens were not observed.

QTL-Seq Analysis for Identification of Resistance Loci to Bacterial Canker in Tomato

Bacterial canker caused by Clavibacter michiganensis (Cm) is one of the most economically important vascular diseases causing unilateral leaf wilting, stem canker, a bird’s-eye lesion on fruit, and whole plant wilting in tomato. There is no commercially available cultivar with bacterial canker resistance, and genomics-assisted breeding can accelerate the development of cultivars with enhanced resistance. Solanum lycopersicum “Hawaii 7998” was found to show bacterial canker resistance. A Quantitative trait loci (QTL)-seq was performed to identify the resistance loci using 909 F2 individuals derived from a cross between S. lycopersicum “E6203” (susceptible) and “Hawaii 7998,” and a genomic region (37.24–41.15 Mb) associated with bacterial canker resistance on chromosome 6 (Rcm6) was found. To dissect the Rcm6 region, 12 markers were developed and several markers were associated with the resistance phenotypes. Among the markers, the Rcm6-9 genotype completely matched with the phenotype in the 47 cultivars. To further validate the Rcm6 as a resistance locus and the Rcm6-9 efficiency, subsequent analysis using F2 and F3 progenies was conducted. The progeny individuals with homozygous resistance allele at the Rcm6-9 showed significantly lower disease severity than those possessing homozygous susceptibility alleles. Genomes of five susceptible and two resistant cultivars were analyzed and previously known R-genes were selected to find candidate genes for Rcm6. Nucleotide-binding leucine-rich repeat, receptor-like kinase, and receptor-like protein were identified to have putative functional mutations and show differential expression upon the Cm infection. The DNA markers and candidate genes will facilitate marker-assisted breeding and provide genetic insight of bacterial canker resistance in tomato.

Marker-Assisted Development and Evaluation of Monogenic Lines of Rice cv. Kaohsiung 145 Carrying Blast Resistance Genes.

Rice blast is a serious threat to global rice production. Large-scale and long-term cultivation of rice varieties with a single blast resistance gene usually leads to breakdown of resistance. To effectively control rice blast in Taiwan, marker-assisted backcrossing was conducted to develop monogenic lines carrying different blast resistance genes in the genetic background of an elite japonica rice cultivar, Kaohsiung 145 (KH145). Eleven International Rice Research Institute (IRRI)-bred blast-resistant lines (IRBLs) showing broad-spectrum resistance to local Pyricularia oryzae isolates were used as resistance donors. Sequencing analysis revealed that the recurrent parent, KH145, does not carry known resistance alleles at the target Pi2/9, Pik, Pita, and Ptr loci. For each IRBL × KH145 cross, we screened 21 to 370 (average of 108) plants per generation from the BC1F1 to BC3F1/BC4F1 generation. A total of 1,499 BC3F2/BC4F2 lines carrying homozygous resistance alleles were selected and self-crossed for four to six successive generations. The derived lines were also evaluated for background genotype using genotyping by sequencing, for blast resistance under artificial inoculation and natural infection conditions, and for agronomic performance in multiple field trials. In Chiayi and Taitung blast nurseries in 2018 to 2020, Pi2, Pi9, and Ptr conferred high resistance, Pi20 and Pik-h moderate resistance, and Pi1, Pi7, Pik-p, and Pik susceptibility to leaf blast; only Pi2, Pi9, and Ptr conferred effective resistance against panicle blast. The monogenic lines showed agronomic traits, yield, and grain quality similar to those of KH145, suggesting the potential of growing a mixture of lines to achieve durable resistance in the field.

Phenotypic and molecular characterization of Hessian fly resistance in diploid wheat, Aegilops tauschii

BackgroundThe Hessian fly (Mayetiola destructor), belonging to the gall midge family (Cecidomyiidae), is a devastating pest of wheat (Triticum aestivum) causing significant yield losses. Despite identification and characterization of numerous Hessian fly-responsive genes and associated biological pathways involved in wheat defense against this dipteran pest, their functional validation has been challenging. This is largely attributed to the large genome, polyploidy, repetitive DNA, and limited genetic resources in hexaploid wheat. The diploid progenitor Aegilops tauschii, D-genome donor of modern-day hexaploid wheat, offers an ideal surrogate eliminating the need to target all three homeologous chromosomes (A, B and D) individually, and thereby making the functional validation of candidate Hessian fly-responsive genes plausible. Furthermore, the well-annotated sequence of Ae. tauschii genome and availability of genetic resources amenable to manipulations makes the functional assays less tedious and time-consuming. However, prior to utilization of this diploid genome for downstream studies, it is imperative to characterize its physical and molecular responses to Hessian fly.ResultsIn this study we screened five Ae. tauschii accessions for their response to the Hessian fly biotypes L and vH13. Two lines were identified that exhibited a homozygous resistance response to feeding by both Hessian fly biotypes. Studies using physical measurements and neutral red staining showed that the resistant Ae. tauschii accessions resembled hexaploid wheat in their phenotypic responses to Hessian fly, that included similarities in larval developmental stages, leaf and plant growth, and cell wall permeability. Furthermore, molecular responses, characterized by gene expression profiling using quantitative real-time PCR, in select resistant Ae. tauschii lines also revealed similarities with resistant hexaploid wheat.ConclusionsPhenotypic and molecular characterization of Ae. tauschii to Hessian fly infestation revealed resistant accessions that shared similarities to hexaploid wheat. Resembling the resistant hexaploid wheat, the Ae. tauschii accessions mount an early defense strategy involving defense proteins including lectins, secondary metabolites and reactive oxygen species (ROS) radicals. Our results reveal the suitability of the diploid progenitor for use as an ideal tool for functional genomics research in deciphering the wheat-Hessian fly molecular interactions.

Preliminary report of knockdown resistance in Culex pipiens pallens and Aedes koreicus from Korea

AbstractPyrethroid insecticides have been effective and powerful for controlling mosquitoes. However, abuse of these insecticides increases the number of resistant mosquitoes. In this study, Culex pipiens pallens and Aedes koreicus were collected from an artificial reservoir in the vicinity of a populated area in Korea, which is also a migratory bird catchment area. To monitor resistance to pyrethroid insecticides in mosquitoes, genomic DNA from the collected mosquitoes was sequenced for the kdr mutation in the voltage‐gated sodium channel (VGSC) gene. As a result, three samples with homozygous resistance (17.6%) and one with heterozygous resistance (5.9%) were found among 17 Cx. pipiens pallens specimens. One of the samples had a unique sequence at the amplified VGSC region. Of the 15 Ae. koreicus, no insecticide resistant individuals were found. In Korea, this is the first report of kdr genetic traits in Ae. koreicus and Cx. pipiens pallens and of a unique VGSC allele in Cx. pipiens pallens. Further investigation is needed to monitor the kdr resistance of these species in Korea and to determine how the unique sequence found in Cx. pipiens pallens is related to insecticide resistance.

Registration of 17 Sorghum Pollinator Germplasm Lines Resistant to Acetolactate Synthase (ALS)–Inhibitor Herbicides

Seventeen sorghum [Sorghum bicolor (L.) Moench] pollinator lines carrying the homozygous resistance gene to the acetolactate synthase (ALS)‐inhibitor herbicides were developed and released by the Kansas Agricultural Experiment Station, Manhattan, KS in 2014. The resistance gene was introduced from shattercane, a wild relative of sorghum, first discovered in Kansas. These lines—PR11/12‐806 (Reg. No. GP‐867, PI 689022), PR11/12‐807 (Reg. No. GP‐868, PI 689023), PR11/12‐816 (Reg. No. GP‐869, PI 689024); PR11/12‐821 (Reg. No. GP‐870, PI 689025), PR11/12‐824 (Reg. No. GP‐871, PI 689026), PR11/12‐828 (Reg. No. GP‐872, PI 689027), PR11/12‐829 (Reg. No. GP‐873, PI 689028), PR11/12‐830 (Reg. No. GP‐874, PI 689029), PR11/12‐836 (Reg. No. GP‐875, PI 689030), PR11/12‐838 (Reg. No. GP‐876, PI 689031), PR11/12‐867 (Reg. No. GP‐877, PI 689032), PR11/12‐873 (Reg. No. GP‐878, PI 689033), PR11/12‐903 (Reg. No. GP‐879, PI 689034), PR11/12‐925 (Reg. No. GP‐880, PI 689035), PR11/12‐928 (Reg. No. GP‐881, PI 689036), PR11/12‐930 (Reg. No. GP‐882, PI 689037), and PR11/12‐948 (Reg. No. GP‐883, PI 689022)—are of typical combine height, are photoperiod insensitive, and reach half‐bloom stage in 60 to 70 d after planting under Manhattan, KS, conditions. The seed color ranges from bright red to tan white. All lines restore fertility in A1 cytoplasm and have good combining ability for yield. These lines can be useful sources for breeding herbicide‐resistant pollinator lines or for direct use as pollinator parent in the production of ALS inhibitor herbicide–resistant hybrid seed.

Feature of blast resistant near-isogenic lines using an elite maintainer line II-32B by marker-assisted selection

Rice plays a crucial role in food production worldwide as one of the most important cereal crops. Breeding blast disease resistant varieties is a long-term aim for sustainable rice production. II-32B is an elite maintainer line of II-32A which has been widely used as a hybrid rice parent in China. However, it is significantly susceptible to rice blast. In this study, multiple alleles, Pi-2 and Pi-9, and the chromosomal segment of blast resistance genes, Pi-1 and Pi-kh, were introgressed to receptor parents II-32B by marker-assisted backcross breeding. Eleven near-isogenic lines (NILs) with single Pi-gene and 12 pyramid lines containing double or three homozygous resistance genes were obtained. These improved lines were identified by inoculating Magnaporthe oryzae isolates in the field/greenhouses; the result showed that the resistance of pyramid lines containing three resistance genes was stronger than the lines with double or single gene, whereas the lines with double genes were stronger than the one with single-gene. Additionally, pyramiding of different Pi-genes was effective in obtaining the durable blast resistance in rice. This implies that gene pyramiding in rice can increase spectrum and strength of resistance to blast. Furthermore, three-line retention of sixteen improved NILs was kept well and the agronomic traits were consistence with II-32B. We conclude that gene pyramiding is an effective breeding approach for the improvement of rice varieties with durable resistance to blast in the future.

Identification of resistance in tomato against root knot nematode (Meloidogyne incognita) and comparison of molecular markers for Mi gene

Tomato production is limited by many biotic stresses of which root knot nematode (RKN, Meloidogyne incognita) is a major pest. The present study aimed to identify resistance sources in controlled conditions and compare molecular markers for efficient and rapid screening of M. incognita resistance. Among the ten genotypes evaluated, HAT-310 and HAT-311 were found immune to M. incognita infestation. Further, six crosses with these two resistant sources, (HAT-311 x Swarna Lalima, HAT-296 x HAT-311, EC-596747 x HAT-311, Swarna Lalima x HAT-310, EC-596743 x HAT-310 and Swarna Lalima x HAT-311), exhibited immune responses against M. incognita. Four molecular markers viz. JB-1, REX-1, PMi12 and Mi23 were employed in eighteen germplasm to characterise resistance and susceptibility of the genotypes against infestation by M. incognita. JB-1 yielded 420 bp in all the genotypes after digestion and hence could not be used to differentiate between nematode resistance and susceptibility. Marker PMi12 yielded additional DNA fragments in addition to the expected bands and did not give consistent results. REX-1 and Mi23 markers successfully differentiated between nematode resistant and susceptible genotypes. Moreover, Mi23 separated the homozygous and heterozygous resistance sources since the restriction enzyme analysis was not needed. The resistant genotypes identified from the present study may be used further in nematode resistance breeding programmes of tomato and the Mi23 marker can be used for rapid screening of the germplasm.

Twenty Years of Tomato Breeding at EPSO-UMH: Transfer Resistance from Wild Types to Local Landraces—From the First Molecular Markers to Genotyping by Sequencing (GBS)

In 1998, the plant breeding team at the School of Engineering of Orihuela (EPSO), part of the Miguel Hernández University (UMH) in Elche, commenced a tomato breeding program. Marker-assisted selection and backcrossing were used to simultaneously introduce three genes (Tm-2a, Ty-1, and Sw-5) that confer resistance to relevant viruses, such as tomato mosaic virus (ToMV), tomato yellow curl virus (TYLCV), and tomato spotted wilt virus (TSWV), to traditional varieties of local tomatoes, specifically the “Muchamiel” and the “De la pera” types. After each backcross, cleaved amplified polymorphic sequence (CAPS) molecular markers were used to select the plants with the resistance genes of interest. A previously described marker was used for TSWV, and new markers were designed for ToMV, and TYLCV using available sequences in the National Center for Biotechnology Information (NCBI) database. In parallel to the breeding program, several molecular markers—Sequence Related Amplified Polymorphism (SRAP), Simple Sequence Repeats (SSRs), Amplified Fragment Length Polymorphisms (AFLPs), Single Nucleotide Polymorphisms (SNPs), and (GATA)4 probes—were used to study genetic variability, and to identify a collection of Spanish and Italian traditional tomato varieties. The results showed a limited genetic variability among cultivated tomato varieties. The breeding lines Muchamiel UMH 1200, and De la pera 1203 (both with homozygous resistance to the three viruses) were the first new varieties that were obtained. They were included in the Register of Protected Plant Varieties in 2013. Lines without a resistance to TYLCV were also developed, and protected in 2017. We have begun to use SNP massive genotyping for studies of genetic association, and for selecting plants with the Ty-1 gene with less linkage drag. Molecular markers have been extremely useful in identifying the different steps of the tomato breeding program at EPSO-UMH.

Identification, introgression, and molecular marker genetic analysis and selection of a highly effective novel oat crown rust resistance from diploid oat, Avena strigosa.

Oat crown rust is one of the most damaging diseases of oat. We identified a new source of resistance and developed KASP and TaqMan markers for selection in breeding programs. A new highly effective resistance to oat crown rust (Puccinia coronata f. sp. avenae) was identified in the diploid oat Avena strigosa PI 258731 and introgressed into hexaploid cultivated oat. Young plants with this resistance show moderate susceptibility, whereas older plant tissues and adult plants are resistant with no virulent isolates encountered in over 8years of testing. Resistance was incorporated into hexaploid oat by embryo rescue, colchicine chromosome doubling followed by backcrosses with a hexaploid parent, and selection for stable transmission of resistance. To mitigate flag leaf and panicle chlorosis/necrosis associated with the resistance, crosses were made with derived resistant lines to breeding lines of divergent parentage followed by selection. Subsequently, two F2 sister lines, termed MNBT1020-1 and MNBT1021-1, were identified in which the chlorosis/necrosis was reduced. These two lines performed well in replicated multi-location state trials in 2015 and 2016 out-yielding all cultivar entries. Segregating F2:3 plants resulting from crosses of MNBT lines to susceptible parents were genotyped with the oat 6K SNP array, and SNP loci with close linkage to the resistance were identified. KASP assays generated from linked SNPs showed accurate discrimination of the resistance in derivatives of the resistant MNBT lines crossed to susceptible breeding lines. A TaqMan marker was developed and correctly identified homozygous resistance in over 95% of 379 F4 plants when rust was scored in F4:5 plants in the field. Thus, a novel highly effective resistance and associated molecular markers are available for use in breeding, genetic analysis, and functional studies.

Major histocompatibility complex variation and the evolution of resistance to amphibian chytridiomycosis.

Chytridiomycosis, caused by the fungal pathogen Batrachochytrium dendrobatidis (Bd), has been implicated in population declines and species extinctions of amphibians around the world. Susceptibility to the disease varies both within and among species, most likely attributable to heritable immunogenetic variation. Analyses of transcriptional expression in hosts following their infection by Bd reveal complex responses. Species resistant to Bd generally show evidence of stronger innate and adaptive immune system responses. Major histocompatibility complex (MHC) class I and class II genes of some susceptible species are up-regulated following host infection by Bd, but resistant species show no comparable changes in transcriptional expression. Bd-resistant species share similar pocket conformations within the MHC-II antigen-binding groove. Among susceptible species, survivors of epizootics bear alleles encoding these conformations. Individuals with homozygous resistance alleles appear to benefit by enhanced resistance, especially in environmental conditions that promote pathogen virulence. Subjects that are repeatedly infected and subsequently cleared of Bd can develop an acquired immune response to the pathogen. Strong directional selection for MHC alleles that encode resistance to Bd may deplete genetic variation necessary to respond to other pathogens. Resistance to chytridiomycosis incurs life-history costs that require further study.

Transferring clubroot resistance from Chinese cabbage (Brassica rapa) to canola (B. napus)

Clubroot, caused by Plasmodiophora brassicae, is one of the most important diseases of Brassica species worldwide, including vegetable and oilseed crops. A dominant clubroot resistance gene from B. rapa (Chinese cabbage) was previously fine mapped and molecular markers were developed in Chinese cabbage that could be used for marker-assisted selection (MAS) in other Brassica crops. To transfer this clubroot resistance gene to B. napus (canola), an interspecific hybridization was made between B. napus (canola) and B. rapa (Chinese cabbage). Subsequently, the F1 was backcrossed to the canola recurrent parent for three generations to produce BC1, BC2 and BC3 progenies. Using these populations, simple sequence repeat (SSR) markers flanking the clubroot resistance gene were used to perform MAS in canola. These molecular markers were then evaluated in 13 different canola and rapeseed quality genotypes in B. napus and B. rapa. These markers exhibited high reliability in identifying clubroot resistance in this diverse set of Brassica genotypes. Clubroot resistance also co-segregated with the SSR markers flanking the clubroot resistance gene in the BC3 and BC3S1. The segregation ratio of resistant and susceptible individuals in the BC3 supported the expected 1:1 ratio for the segregation of a single Mendelian gene. BC3S1 families with homozygous clubroot resistance were developed during this process, and should be valuable sources of clubroot resistance in B. napus breeding activities.

Breeding Elite Lines of Apple Carrying Pyramided Homozygous Resistance Genes Against Apple Scab and Resistance Against Powdery Mildew and Fire Blight

The development of high-quality cultivars, with durable disease resistance, is a major objective of apple breeding. The selection procedures of modern breeding programs no longer rely exclusively on phenotypic criteria but include marker-assisted breeding (MAB). Currently, molecular markers linked to several resistance genes and quantitative trait loci (QTLs) are available. In this study, we focused on markers available for resistance breeding against the major diseases scab (Venturia inaequalis), powdery mildew (Podosphaera leucotricha), and fire blight (Erwinia amylovora). One approach proposed to achieve durable resistance is the pyramiding of functionally different resistance genes against the same pathogen. This approach can be complemented with the incorporation of resistance genes against other pathogens. The resulting resistant apple cultivars would contribute considerably to low-input, sustainable, fruit production. Furthermore, apple cultivars can be developed carrying homozygous allele sets of specific resistance genes, and these genotypes can be used as parents for further crosses. Due to the ensured inheritance of the resistance genes to the progeny, MAB for these genes will become superfluous. In this study, we developed elite apple plants which are homozygous for three different scab resistance genes, Rvi6, Rvi2, and Rvi4. Furthermore, these apple selections tested positive for a resistance gene against powdery mildew (Pl1 or Pl2), and the FBF7 QTL from ‘Fiesta’ for enhanced fire blight resistance. Selected progeny plants were tested for their fire blight resistance after artificial shoot inoculation and evaluated for tree and fruit characteristics.