Genome sequence of Xanthomonas fuscans subsp. fuscans strain 4834-R reveals that flagellar motility is not a general feature of xanthomonads

Abstract\ud \ud \ud \ud Background\ud Xanthomonads are plant-associated bacteria responsible for diseases on economically important crops. Xanthomonas fuscans subsp. fuscans (Xff) is one of the causal agents of common bacterial blight of bean. In this study, the complete genome sequence of strain Xff 4834-R was determined and compared to other Xanthomonas genome sequences.\ud \ud \ud \ud Results\ud Comparative genomics analyses revealed core characteristics shared between Xff 4834-R and other xanthomonads including chemotaxis elements, two-component systems, TonB-dependent transporters, secretion systems (from T1SS to T6SS) and multiple effectors. For instance a repertoire of 29 Type 3 Effectors (T3Es) with two Transcription Activator-Like Effectors was predicted. Mobile elements were associated with major modifications in the genome structure and gene content in comparison to other Xanthomonas genomes. Notably, a deletion of 33 kbp affects flagellum biosynthesis in Xff 4834-R. The presence of a complete flagellar cluster was assessed in a collection of more than 300 strains representing different species and pathovars of Xanthomonas. Five percent of the tested strains presented a deletion in the flagellar cluster and were non-motile. Moreover, half of the Xff strains isolated from the same epidemic than 4834-R was non-motile and this ratio was conserved in the strains colonizing the next bean seed generations.\ud \ud \ud \ud Conclusions\ud This work describes the first genome of a Xanthomonas strain pathogenic on bean and reports the existence of non-motile xanthomonads belonging to different species and pathovars. Isolation of such Xff variants from a natural epidemic may suggest that flagellar motility is not a key function for in planta fitness.AI is funded by a PhD grant from INRA-SPE and region Pays de la Loire, France. EG was funded by a PhD grant from the French Ministry of National Education and Research and French Guyana. SC, EG, MA, EL and LDN are funded by the LABEX TULIP (ANR-10-LABX-41), LSG is funded by ANR-2010-GENM-013 Xanthomix

Comparative analysis of the XopN family in the genus Xanthomonas.

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The “repertoire for repertoire” hypothesis: Repertoires of type three effectors may explain host specificity in Xanthomonas

International audienceHost range of many plant pathogenic bacteria is restricted to one or a few host plant species reflecting a tight adaptation of bacteria to specific hosts. Two hypotheses could explain host specificity: either it can be explained by the phylogenetic position of the strains, or, alternatively, by the association of virulence genes enabling a pathological convergence of phylogenetically distant strains. In this latter hypothesis, host specificity would result from the interaction between repertoires of bacterial virulence genes and repertoires of genes involved in host surveillance systems and defences (a “repertoire for repertoire” hypothesis). To challenge these two hypotheses, we selected 132 Xanthomonas axonopodis strains representative of 18 different pathovars which display different host range. First, the phylogenetic position of each strain was determined by sequencing the housekeeping gene rpoD. This study showed that many pathovars of Xanthomonas axonopodis are polyphyletic. We thus investigated the distribution of 37 T3E genes in these strains by both PCR and hybridization methods. Our study revealed that T3E repertoires are highly variable between these strains, both in terms of T3E presence and of size of repertoires. T3E repertoires comprise both core and flexible gene suites that likely have distinct roles in pathogenicity and different evolutionary histories. Our results showed a correspondence between composition of T3E repertoires and pathovars of Xanthomonas axonopodis. For polyphyletic pathovars, this strongly suggests that T3E genes explain a pathological convergence of phylogenetically distant strains. Finally, our results support the “repertoire for repertoire” hypothesis as the molecular basis of host specificity for plant pathogenic bacteria

Correction: A «Repertoire for Repertoire» Hypothesis: Repertoires of Type Three Effectors are Candidate Determinants of Host Specificity in Xanthomonas

BackgroundThe genetic basis of host specificity for animal and plant pathogenic bacteria remains poorly understood. For plant pathogenic bacteria, host range is restricted to one or a few host plant species reflecting a tight adaptation to specific hosts. Methodology/Principal Findings Two hypotheses can be formulated to explain host specificity: either it can be explained by the phylogenetic position of the strains, or by the association of virulence genes enabling a pathological convergence of phylogenically distant strains. In this latter hypothesis, host specificity would result from the interaction between repertoires of bacterial virulence genes and repertoires of genes involved in host defences. To challenge these two hypotheses, we selected 132 Xanthomonas axonopodis strains representative of 18 different pathovars which display different host range. First, the phylogenetic position of each strain was determined by sequencing the housekeeping gene rpoD. This study showed that many pathovars of Xanthomonas axonopodis are polyphyletic. Second, we investigated the distribution of 35 type III effector genes (T3Es) in these strains by both PCR and hybridization methods. Indeed, for pathogenic bacteria T3Es were shown to trigger and to subvert host defences. Our study revealed that T3E repertoires comprise core and variable gene suites that likely have distinct roles in pathogenicity and different evolutionary histories. Our results showed a correspondence between composition of T3E repertoires and pathovars of Xanthomonas axonopodis. For polyphyletic pathovars, this suggests that T3E genes might explain a pathological convergence of phylogenetically distant strains. We also identified several DNA rearrangements within T3E genes, some of which correlate with host specificity of strains. Conclusions/Significance These data provide insight into the potential role played by T3E genes for pathogenic bacteria and support a “repertoire for repertoire” hypothesis that may explain host specificity. Our work provides resources for functional and evolutionary studies aiming at understanding host specificity of pathogenic bacteria, functional redundancy between T3Es and the driving forces shaping T3E repertoires

The Type VI secretion system of Stenotrophomonas rhizophila CFBP13503 limits the transmission of Xanthomonas campestris pv. campestris 8004 from radish seeds to seedlings

Stenotrophomonas rhizophila CFBP13503 is a seed-borne commensal bacterial strain, which is efficiently transmitted to seedlings and can outcompete the phytopathogenic bacteria Xanthomonas campestris pv. campestris (Xcc8004). The type VI Secretion System (T6SS), an interference contact-dependent mechanism, is a critical component of interbacterial competition. The involvement of the T6SS of S. rhizophila CFBP13503 in the inhibition of Xcc8004 growth and seed-to-seedling transmission was assessed. The T6SS cluster of S. rhizophila CFBP13503 and nine putative effectors were identified. Deletion of two T6SS structural genes, hcp and tssB, abolished the competitive advantage of S. rhizophila against Xcc8004 in vitro. The population sizes of these two bacterial species were monitored in seedlings after inoculation of radish seeds with mixtures of Xcc8004 and either S. rhizophila wild type (wt) strain or isogenic hcp mutant. A significant decrease in the population size of Xcc8004 was observed during confrontation with the S. rhizophila wt in comparison to T6SS deletion mutants in germinated seeds and seedlings. We found that the T6SS distribution among 835 genomes of the Stenotrophomonas genus is scarce. In contrast, in all available S. rhizophila genomes, T6SS clusters are widespread and mainly belong to the T6SS group i4. In conclusion, the T6SS of S. rhizophila CFBP13503 is involved in the antibiosis against Xcc8004 and reduces seedling transmission of Xcc8004 in radish. The distribution of this T6SS cluster in the S. rhizophila complex could make it possible to exploit these strains as biocontrol agents against X. campestris pv. campestris

Evidences for selection on promoter region reveal a hidden aspect of the multifaceted evolution of the type three effector Avrbs2 in the Xanthomonas axonopodis species

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Evolutionary history of the plant pathogenic bacterium Xanthomonas axonopodis.

Deciphering mechanisms shaping bacterial diversity should help to build tools to predict the emergence of infectious diseases. Xanthomonads are plant pathogenic bacteria found worldwide. Xanthomonas axonopodis is a genetically heterogeneous species clustering, into six groups, strains that are collectively pathogenic on a large number of plants. However, each strain displays a narrow host range. We address the question of the nature of the evolutionary processes--geographical and ecological speciation--that shaped this diversity. We assembled a large collection of X. axonopodis strains that were isolated over a long period, over continents, and from various hosts. Based on the sequence analysis of seven housekeeping genes, we found that recombination occurred as frequently as point mutation in the evolutionary history of X. axonopodis. However, the impact of recombination was about three times greater than the impact of mutation on the diversity observed in the whole dataset. We then reconstructed the clonal genealogy of the strains using coalescent and genealogy approaches and we studied the diversification of the pathogen using a model of divergence with migration. The suggested scenario involves a first step of generalist diversification that spanned over the last 25,000 years. A second step of ecology-driven specialization occurred during the past two centuries. Eventually, secondary contacts between host-specialized strains probably occurred as a result of agricultural development and intensification, allowing genetic exchanges of virulence-associated genes. These transfers may have favored the emergence of novel pathotypes. Finally, we argue that the largest ecological entity within X. axonopodis is the pathovar

Role of the acquisition of a type 3 secretion system in the emergence of novel pathogenic strains of Xanthomonas

International audienceCases of emergence of novel plant-pathogenic strains are regularly reported that reduce the yields of crops and trees. However, the molecular mechanisms underlying such emergence are still poorly understood. The acquisition by environmental non-pathogenic strains of novel virulence genes by horizontal gene transfer has been suggested as a driver for the emergence of novel pathogenic strains. In this study, we tested such an hypothesis by transferring a plasmid encoding the type 3 secretion system (T3SS) and four associated type 3 secreted proteins (T3SPs) to the non-pathogenic strains of Xanthomonas CFBP 7698 and CFBP 7700, which lack genes encoding T3SS and any previously known T3SPs. The resulting strains were phenotyped on Nicotiana benthamiana using chlorophyll fluorescence imaging and image analysis. Wild-type, non-pathogenic strains induced a hypersensitive response (HR)-like necrosis, whereas strains complemented with T3SS and T3SPs suppressed this response. Such suppression depends on a functional T3SS. Amongst the T3SPs encoded on the plasmid, Hpa2, Hpa1 and, to a lesser extent, XopF1 collectively participate in suppression. Monitoring of the population sizes in planta showed that the sole acquisition of a functional T3SS by non-pathogenic strains impairs growth inside leaf tissues. These results provide functional evidence that the acquisition via horizontal gene transfer of a T3SS and four T3SPs by environmental non-pathogenic strains is not sufficient to make strains pathogenic. In the absence of a canonical effector, the sole acquisition of a T3SS seems to be counter-selective, and further acquisition of type 3 effectors is probably needed to allow the emergence of novel pathogenic strains.© 2018 BSPP and John Wiley & Sons Ltd

Data from: Genome mining reveals the genus Xanthomonas to be a promising reservoir for new bioactive non-ribosomally synthesized peptides

Background: Various bacteria can use non-ribosomal peptide synthesis (NRPS) to produce peptides or other small molecules. Conserved features within the NRPS machinery allow the type, and sometimes even the structure, of the synthesized polypeptide to be predicted. Thus, bacterial genome mining via in silico analyses of NRPS genes offers an attractive opportunity to uncover new bioactive non-ribosomally synthesized peptides. Xanthomonas is a large genus of Gram-negative bacteria that cause disease in hundreds of plant species. To date, the only known small molecule synthesized by NRPS in this genus is albicidin produced by Xanthomonas albilineans. This study aims to estimate the biosynthetic potential of Xanthomonas spp. by in silico analyses of NRPS genes with unknown function recently identified in the sequenced genomes of X. albilineans and related species of Xanthomonas. Results: We performed in silico analyses of NRPS genes present in all published genome sequences of Xanthomonas spp., as well as in unpublished draft genome sequences of Xanthomonas oryzae pv. oryzae strain BAI3 and Xanthomonas spp. strain XaS3. These two latter strains, together with X. albilineans strain GPE PC73 and X. oryzae pv. oryzae strains X8-1A and X11-5A, possess novel NRPS gene clusters and share related NRPS-associated genes such as those required for the biosynthesis of non-proteinogenic amino acids or the secretion of peptides. In silico prediction of peptide structures according to NRPS architecture suggests eight different peptides, each specific to its producing strain. Interestingly, these eight peptides cannot be assigned to any known gene cluster or related to known compounds from natural product databases. PCR screening of a collection of 94 plant pathogenic bacteria indicates that these novel NRPS gene clusters are specific to the genus Xanthomonas and are also present in Xanthomonas translucens and X. oryzae pv. oryzicola. Further genome mining revealed other novel NRPS genes specific to X. oryzae pv. oryzicola or Xanthomonas sacchari. Conclusions: This study revealed the significant potential of the genus Xanthomonas to produce new non-ribosomally synthesized peptides. Interestingly, this biosynthetic potential seems to be specific to strains of Xanthomonas associated with monocotyledonous plants, suggesting a putative involvement of non-ribosomally synthesized peptides in plant-bacteria interactions