We investigated whether lines of transgenic tomato (Solanum lycopersicum) expressing the Bs2 resistance gene from pepper, a close relative of tomato, demonstrate improved resistance to bacterial spot ...disease caused by Xanthomonas species in replicated multi-year field trials under commercial type growing conditions. We report that the presence of the Bs2 gene in the highly susceptible VF 36 background reduced disease to extremely low levels, and VF 36-Bs2 plants displayed the lowest disease severity amongst all tomato varieties tested, including commercial and breeding lines with host resistance. Yields of marketable fruit from transgenic lines were typically 2.5 times that of the non-transformed parent line, but varied between 1.5 and 11.5 fold depending on weather conditions and disease pressure. Trials were conducted without application of any copper-based bactericides, presently in wide use despite negative impacts on the environment. This is the first demonstration of effective field resistance in a transgenic genotype based on a plant R gene and provides an opportunity for control of a devastating pathogen while eliminating ineffective copper pesticides.
Both hypersensitive and quantitative forms of resistance to the bacterial spot pathogens (Xanthomonas spp.) occur in pepper and tomato. Five resistance genes involved in hypersensitivity in pepper ...and four in tomato have been identified so far. The corresponding pathogen avirulence genes have been cloned and characterized, and features, including a propensity for accumulating mutations and at times, loss of plasmid-borne avirulence genes, are known to occur. The frequency of these changes affects race composition among pathogen populations and determines the durability of the corresponding plant resistance. At least four different species of Xanthomonas are known to cause bacterial spot, and these can differ in specific avirulence gene content. Quantitative or multigenic resistance has also more recently been researched and appears to be more durable than the hypersensitive resistance. Two recessive genes have been identified that yield a nonhypersensitive form of resistance in pepper and together can provide strong resistance. More emphasis is being given to transfer of quantitative trait resistance to commercial cultivars of both tomato and pepper.
Bacterial spot is a destructive disease of tomato in Florida that prior to the early 1990s was caused by Xanthomonas euvesicatoria. X. perforans was first identified in Florida in 1991 and by 2006 ...was the only xanthomonad associated with bacterial spot disease in tomato. The ability of an X. perforans strain to outcompete X. euvesicatoria both in vitro and in vivo was at least in part associated with the production of three bacteriocins designated Bcn-A, Bcn-B, and Bcn-C. The objective of this study was to characterize the genetic determinants of these bacteriocins. Bcn-A activity was confined to one locus consisting of five ORFs of which three (ORFA, ORF2 and ORF4) were required for bacteriocin activity. The fifth ORF is predicted to encode an immunity protein to Bcn-A based on in vitro and in vivo assays. The first ORF encodes Bcn-A, a 1,398 amino acid protein, which bioinformatic analysis predicts to be a member of the RHS family of toxins. Based on results of homology modeling, we hypothesize that the amino terminus of Bcn-A interacts with a protein in the outer membrane of X. euvesicatoria. The carboxy terminus of the protein may interact with an as yet unknown protein(s) and puncture the X. euvesicatoria membrane, thereby delivering the accessory proteins into the target and causing cell death. Bcn-A appears to be activated upon secretion based on cell fractionation assays. The other two loci were each shown to be single ORFs encoding Bcn-B and Bcn-C. Both gene products possess homology toward known proteases. Proteinase activity for both Bcn-B and Bcn-C was confirmed using a milk agar assay. Bcn-B is predicted to be an ArgC-like serine protease, which was confirmed by PMSF inhibition of proteolytic activity, whereas Bcn-C has greater than 50% amino acid sequence identity to two zinc metalloproteases.
Transcription activator-like effector (TALE) proteins of the plant pathogenic bacterial genus Xanthomonas bind to and transcriptionally activate host susceptibility genes, promoting disease. Plant ...immune systems have taken advantage of this mechanism by evolving TALE binding sites upstream of resistance (R) genes. For example, the pepper Bs3 and rice Xa27 genes are hypersensitive reaction plant R genes that are transcriptionally activated by corresponding TALEs. Both R genes have a hallmark expression pattern in which their transcripts are detectable only in the presence and not the absence of the corresponding TALE. By transcriptome profiling using next-generation sequencing (RNA-seq), we tested whether we could avoid laborious positional cloning for the isolation of TALE-induced R genes. In a proof-of-principle experiment, RNA-seq was used to identify a candidate for Bs4C , an R gene from pepper that mediates recognition of the Xanthomonas TALE protein AvrBs4. We identified one major Bs4C candidate transcript by RNA-seq that was expressed exclusively in the presence of AvrBs4. Complementation studies confirmed that the candidate corresponds to the Bs4C gene and that an AvrBs4 binding site in the Bs4C promoter directs its transcriptional activation. Comparison of Bs4C with a nonfunctional allele that is unable to recognize AvrBs4 revealed a 2-bp polymorphism within the TALE binding site of the Bs4C promoter. Bs4C encodes a structurally unique R protein and Bs4C -like genes that are present in many solanaceous genomes seem to be as tightly regulated as pepper Bs4C . These findings demonstrate that TALE-specific R genes can be cloned from large-genome crops with a highly efficient RNA-seq approach.
Four phenotypic xanthomonad groups have been identified that are pathogenic to pepper, tomato, or both hosts. These include groups A and C which are found in
Xanthomonas axonopodis pv.
vesicatoria, ...group B found in X.
vesicatoria, and group D found in
‘X. gardneri’. We present DNA:DNA hybridization data in which
X. axonopodis pv.
vesicatoria group A and C strains have less than 70% DNA relatedness with each other, with the type strain of
X. axonopodis, and with the currently classified species within
Xanthomonas and, therefore, should be removed from this species and given species status. We present information that the A strains most closely resemble the strains originally isolated by Doidge in 1921. In an attempt to avoid confusion in nomenclature as stated in Principle 1 of the
Bacteriological Code, we propose that the A strains of
X. axonopodis pv.
vesicatoria be renamed as
X. euvesicatoria (ATCC11633
T = NCPPB2968
T = ICMP 109
T = ICMP 98
T). Use of the
euvesicatoria epithet should be reserved for strains originally identified by Doidge, which she designated
Bacterium vesicatorium (Ann. Appl. Biol. 7: 407–430, 1921) in the original description when she referred to those strains as being feebly amylolytic. The name
X. perforans sp. nov. is proposed for the C group of strains previously designated as
X. axonopodis pv.
vesicatoria (ATCC BAA-983
T = NCPPB 4321
T). We also propose that
‘X. gardneri’, which has less than 70% DNA relatedness with any of the
Xanthomonas species and which has never had taxonomic status, be named
X. gardneri (ATCC 19865
T = NCPPB 881
T) to reflect the specific epithet proposed by Sutic 17 in 1957.
Bacterial spot caused by
is a major disease of pepper (
L.) in warm and humid production environments. Use of genetically resistant cultivars is an effective approach to manage bacterial spot. Two ...recessive resistance genes,
and
, confer non-race-specific resistance against bacterial spot. The objective of our study was to map these two loci in the pepper genome. We used a genotyping-by-sequencing approach to initially map the position of the two resistances. Segregating populations for
and
were developed by crossing susceptible Early CalWonder (ECW) with near-isogenic lines ECW50R (
introgression) or ECW60R (
introgression). Following fine-mapping,
was delimited to a ~535 Kbp interval on chromosome 3, and
to a ~666 Kbp interval in chromosome 6. We identified 14 and 8 candidate resistance genes for
and
, respectively, based on predicted protein coding polymorphisms between ECW and the corresponding resistant parent. This research enhances marker-assisted selection of
and
in breeding programs and is a crucial step towards elucidating the molecular mechanisms underlying the resistances.
Despite evidence supporting pre-exposure prophylaxis (PrEP) efficacy, there are concerns regarding the feasibility of widespread PrEP implementation among men who have sex with men (MSM). To inform ...the development of targeted PrEP delivery guidelines, sexual risk trajectories among HIV-negative MSM were characterized.
At semiannual visits from 2003 to 2011, HIV-negative MSM (N = 419) participating in the Multicenter AIDS Cohort Study provided data on sexual risk behaviors (SRBs) since their last visit. Based on their reported behaviors, participants were assigned a SRB score at each visit as follows: 0 = no insertive or receptive anal intercourse, 1 = no unprotected insertive or receptive anal intercourse, 2 = only unprotected insertive anal intercourse, 3 = unprotected receptive anal intercourse with 1 HIV-negative partner, 4 = condom serosorting, 5 = condom seropositioning, and 6 = no seroadaptive behaviors. Group-based trajectory modeling was used to examine SRB scores (<4 vs. ≥4) and identify groups with distinct sexual risk trajectories.
Three sexual risk trajectory groups were identified: low-risk (n = 264; 63.0%), moderate-risk (n = 96; 22.9%; mean duration of consecutive high-risk intervals ∼1 year), and high-risk (n = 59; 14.1%; mean duration of consecutive high-risk intervals ∼2 years). Compared to low-risk group membership, high-risk group membership was associated with younger age (in years) adjusted odds ratio (AOR) = 0.92, 95% confidence interval (CI): 0.88 to 0.96, being White (AOR = 3.67, 95% CI: 1.48 to 9.11), earning an income ≥$20,000 (AOR = 4.98, 95% CI: 2.13 to 11.64), distress/depression symptoms (Center for Epidemiologic Studies Depression Scale ≥ 16) (AOR = 2.36, 95% CI: 1.14 to 4.92), and substance use (AOR = 2.00, 95% CI: 1.01 to 3.97).
Screening for the sociodemographic and behavioral factors described above may facilitate targeted PrEP delivery during high-risk periods among MSM.
A novel hypersensitive resistance (HR) in Capsicum baccatum var. pendulum against the bacterial spot of pepper pathogen, Xanthomonas gardneri, was introgressed into C. annuum cv. Early Calwonder ...(ECW) to create the near-isogenic line designated as ECW-70R. A corresponding avirulence gene avrBs7, in X. gardneri elicited a strong HR in ECW-70R. A homolog of avrBs7, avrBs1.1, was found in X. euvesicatoria 85-10, which showed delayed HR on ECW-70R leaves. Genetic analysis confirmed the presence of a single dominant resistance gene, Bs7, corresponding to the two avr genes. Both AvrBs7 and AvrBs1.1 share a consensus protein tyrosine phosphatase (PTP) active site domain and can dephosphorylate para-nitrophenyl phosphate. Mutation of Cys(265) to Ser in the PTP domain and subsequent loss of enzymatic activity and HR activity indicated the importance of the PTP domain in the recognition of the Avr protein by the Bs7 gene transcripts. Superpositioning of AvrBs7 and AvrBs1.1 homology models indicated variation in the geometry of the loops adjacent to the active sites. These predicted structural differences might be responsible for the differences in HR timing due to differential activation of the resistance gene. Mutating the PTP domain of AvrBs1.1 to match that of AvrBs7 failed to activate HR on ECW-70R, indicating the possibility of differential substrate specificities between AvrBs1.1 and AvrBs7.
The RXopJ4 resistance locus from the wild accession Solanum pennellii (Sp) LA716 confers resistance to bacterial spot disease of tomato (S. lycopersicum, Sl) caused by Xanthomonas perforans (Xp). ...RXopJ4 resistance depends on recognition of the pathogen type III effector protein XopJ4. We used a collection of Sp introgression lines (ILs) to narrow the RXopJ4 locus to a 4.2-Mb segment on the long arm of chromosome 6, encompassed by the ILs 6-2 and 6-2-2. We then adapted or developed a collection of 14 molecular markers to map on a segregating F₂ population from a cross between the susceptible parent Sl FL8000 and the resistant parent RXopJ4 8000 OC₇. In the F₂ population, a 190-kb segment between the markers J350 and J352 cosegregated with resistance. This fine mapping will enable both the identification of candidate genes and the detection of resistant plants using cosegregating markers. The RXopJ4 resistance gene(s), in combination with other recently characterized genes and a quantitative trait locus (QTL) for bacterial spot disease resistance, will likely be an effective tool for the development of durable resistance in cultivated tomato.
We provide the first conclusive evidence that Xanthomonas axonopodis pv. citri Asiatic strain (Xac-A) and, in particular, Xac-Aw, a unique citrus canker A strain isolated from Key lime in Wellington, ...Florida, induces a hypersensitive reaction (HR) in grapefruit leaves. Using the heterologous tomato pathogen X. perforans, as a recipient of the Xac-Aw genomic library, we identified a 1599-bp open reading frame responsible for HR in grapefruit, but not Key lime, and designated it avrGf1. Xac-AwΔavrGf1 produced typical, although visibly reduced, citrus canker symptoms (i.e. raised pustules) in grapefruit and typical canker symptoms in Key lime. We also determined that the X. perforans transconjugant carrying an Xac-Aw hrpG elicited HR in grapefruit and Key lime leaves, and that xopA in X. perforans was partly responsible for HR. Xac-A transconjugants carrying the X. perforans xopA were reduced in ability to grow in grapefruit leaves relative to wild-type Xac-A. The X. perforans xopA appears to be a host-limiting factor. An avrBs3 homologue, which contained 18.5 repeats and induced HR in tomato, was designated avrTaw. This gene, when expressed in a pustule-minus Xac-Aw, did not complement pustule formation; however, pthAw, a functional pthA homologue, complemented the mutant strain to produce typical pustules in Key lime, but markedly reduced pustules in grapefruit. Both avrBs3 homologues, when expressed in a typical Xac-A strain, resulted in typical citrus canker pustules in grapefruit, indicating that neither homologue suppressed pustule size in grapefruit. Xac-Aw contains other unidentified factors that suppress development in grapefruit.
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