Comparative Genomics Identifies a Novel Conserved Protein, HpaT, in Proteobacterial Type III Secretion Systems that Do Not Possess the Putative Translocon Protein HrpF

Xanthomonas translucens is the causal agent of bacterial leaf streak, the most common bacterial disease of wheat and barley. To cause disease, most xanthomonads depend on a highly conserved type III secretion system, which translocates type III effectors into host plant cells. Mutagenesis of the conserved type III secretion gene hrcT confirmed that the X. translucens type III secretion system is required to cause disease on the host plant barley and to trigger a non-host hypersensitive response (HR) in pepper leaves. Type III effectors are delivered to the host cell by a surface appendage, the Hrp pilus, and a translocon protein complex that inserts into the plant cell plasma membrane. Homologs of the Xanthomonas HrpF protein, including PopF from Ralstonia solanacearum and NolX from rhizobia, are thought to act as a translocon protein. Comparative genomics revealed that X. translucens strains harbor a noncanonical hrp gene cluster, which rather shares features with type III secretion systems from Ralstonia solanacearum, Paraburkholderia andropogonis, Collimonas fungivorans, and Uliginosibacterium gangwonense than other Xanthomonas spp. Surprisingly, none of these bacteria, except R. solanacearum, encode a homolog of the HrpF translocon. Here, we aimed at identifying a candidate translocon from X. translucens. Notably, genomes from strains that lacked hrpF/popF/nolX instead encode another gene, called hpaT, adjacent to and co-regulated with the type III secretion system gene cluster. An insertional mutant in the X. translucens hpaT gene, which is the first gene of a two-gene operon, hpaT-hpaH, was non-pathogenic on barley and did not cause the HR or programmed cell death in non-host pepper similar to the hrcT mutant. The hpaT mutant phenotypes were partially complemented by either hpaT or the downstream gene, hpaH, which has been described as a facilitator of translocation in Xanthomonas oryzae. Interestingly, the hpaT mutant was also complemented by the hrpF gene from Xanthomonas euvesicatoria. These findings reveal that both HpaT and HpaH contribute to the injection of type III effectors into plant cells

Actinorhizal Signaling Molecules: Frankia Root Hair Deforming Factor Shares Properties With NIN Inducing Factor

Actinorhizal plants are able to establish a symbiotic relationship with Frankia bacteria leading to the formation of root nodules. The symbiotic interaction starts with the exchange of symbiotic signals in the soil between the plant and the bacteria. This molecular dialog involves signaling molecules that are responsible for the specific recognition of the plant host and its endosymbiont. Here we studied two factors potentially involved in signaling between Frankia casuarinae and its actinorhizal host Casuarina glauca: (1) the Root Hair Deforming Factor (CgRHDF) detected using a test based on the characteristic deformation of C. glauca root hairs inoculated with F. casuarinae and (2) a NIN activating factor (CgNINA) which is able to activate the expression of CgNIN, a symbiotic gene expressed during preinfection stages of root hair development. We showed that CgRHDF and CgNINA corresponded to small thermoresistant molecules. Both factors were also hydrophilic and resistant to a chitinase digestion indicating structural differences from rhizobial Nod factors (NFs) or mycorrhizal Myc-LCOs. We also investigated the presence of CgNINA and CgRHDF in 16 Frankia strains representative of Frankia diversity. High levels of root hair deformation (RHD) and activation of ProCgNIN were detected for Casuarina-infective strains from clade Ic and closely related strains from clade Ia unable to nodulate C. glauca. Lower levels were present for distantly related strains belonging to clade III. No CgRHDF or CgNINA could be detected for Frankia coriariae (Clade II) or for uninfective strains from clade IV

Functional analysis of type III secretion system and transcription activator-like effectors of Xanthomonas translucens, the causal agent of bacterial leaf streak on cereals

Xanthomonas translucens is the causal agent of bacterial leaf streak, the most common bacterial disease of small grain cereals. To cause disease, most xanthomonads depend on a conserved type III secretion system (Hrp system), which translocates type III effectors (T3E) into plant host cells. Comparative genomics of several X. translucens strains revealed that they contain a hrp gene cluster the genomic organization which is different to that found in other Xanthomonas species. Inoculation methods of barley plants were optimized in order to permit quantitative pathogenicity assays. Mutations were generated in two conserved genes of the X. translucens hrp cluster, affecting hrcT and hgiA (HrpG-induced gene). Both mutations caused drastic reductions in symptom development on the host plant barley and a loss of hypersensitive response in the non-host plant pepper. Evidence is provided that the HgiA protein is an analog of the putative translocon protein HrpF. A specific family of T3Es, called Transcription Activator-Like Effectors (TALEs), modulates plant gene expression. Artificial TALEs were established as a test system for the contribution of plant target genes to disease. TALEs from X. translucens were found to have distinct characteristics compared to other Xanthomonas TALEs. This study identified a conserved TALE in barley-pathogenic xanthomonads, which strongly contributes to disease development.(AGRO - Sciences agronomiques et ingénierie biologique) -- UCL, 201

Frankia & Friends: Roles of Various Nodule Inhabitants in the Actinorhizal Symbiosis

Actinorhizal plants are woody dicotyledons from eight families that form symbiotic root nodules with the genus Frankia, nitrogen-fixing actinobacteria. Recent studies have found that actinorhizal nodules contain other bacterial inhabitants besides Frankia. The roles that members of the actinorhizal microbiome play are largely unknown. In this study, several bacterial strains were isolated from alder (Alnus) nodules growing at Adam’s Point in Durham, NH in spring 2018 and 2019. These isolates were tested for chemotactic/chemotropic properties and their impacts on alder seedlings and nodulation. A chemotaxis/chemotropism assay was developed to detect the response of these bacteria to actinorhizal root exudates. Additionally, sterile Alnus glutinosa seedlings grown under nitrogen-deficient conditions were inoculated with bacterial isolates alone or in co-culture with Frankia to assess impacts on plant health. For the chemotaxis assay, a few strains including Kocuria, Curtobacterium, Streptomyces, and Herbaspirillum, exhibited differences in motility or produced crystals depending on conditions. The Streptomyces isolate and one of the Kocuria strains exhibited attraction to root exudates from the actinorhizal plant Eleagnus angustifolia, and A. glutinosa to a lesser extent. Preliminary plant studies suggest Streptomyces strain 23 decreased nitrogen stress symptoms in A. glutinosa, and the Streptomyces and Bacillus isolates may play a role in promoting secondary root formation

Effects of the phenylurea herbicide diuron on natural riverine microbial communities in an experimental study.

International audienceThe effects of the phenylurea herbicide diuron (10 microgl(-1)) on natural riverine microbial communities were investigated using a three-week laboratory microcosm study. During the first six days, a latency period was observed both in the algal and the bacterial communities despite favorable abiotic conditions and independently of diuron exposure. From the second week, an intense algal bloom (chlorophyll a concentrations and cell abundances) was observed in the uncontaminated microcosms but not in the treated microcosms. The bloom stimulated the bacterial community and led to an increase in heterotrophic bacterial production ([3H]thymidine incorporation), activity (CTC reduction) and cell abundance. In parallel, shifts in bacterial community composition were recorded by polymerase chain reaction (PCR)-temporal temperature gradient gel electrophoresis (TTGE) analysis, whereas no major variation was detected using the fluorescent in situ hybridization (FISH) method. In the treated microcosms, the diuron acted not by damaging the initial communities but by inhibiting the algal bloom and indirectly maintaining constant bacterial conditions throughout the experiment. These inhibitory effects, which were recorded in terms of abundance, activity and diversity, suggest that exposure to diuron can decrease the recovery capacities of microbial communities and delay the resumption of an efficient microbial food web despite favorable environmental conditions

Five phylogenetically close rice SWEET genes confer TAL effector-mediated susceptibility to Xanthomonas oryzae pv. oryzae

Bacterial plant-pathogenic Xanthomonas strains translocate transcription activator-like (TAL) effectors into plant cells to function as specific transcription factors. Only a few plant target genes of TAL effectors have been identified, so far. Three plant SWEET genes encoding putative sugar transporters are known to be induced by TAL effectors from rice-pathogenic Xanthomonas oryzae pv. oryzae (Xoo). We predict and validate that expression of OsSWEET14 is induced by a novel TAL effector, Tal5, from an African Xoo strain. Artificial TAL effectors (ArtTALs) were constructed to individually target 20 SWEET orthologs in rice. They were used as designer virulence factors to study which rice SWEET genes can support Xoo virulence. The Tal5 target box differs from those of the already known TAL effectors TalC, AvrXa7 and PthXo3, which also induce expression of OsSWEET14, suggesting evolutionary convergence on key targets. ArtTALs efficiently complemented an Xoo talC mutant, demonstrating that specific induction of OsSWEET14 is the key target of TalC. ArtTALs that specifically target individual members of the rice SWEET family revealed three known and two novel SWEET genes to support bacterial virulence. Our results demonstrate that five phylogenetically close SWEET proteins, which presumably act as sucrose transporters, can support Xoo virulence. © 2013 No claim to French Government works. © 2013 New Phytologist Trust

Effects of diuron in microcosms on natural riverine bacterial community composition : new insight into phylogenetic approaches using PCR-TTGE analysis

International audienceWe recently demonstrated in a microcosm study using PCR-TTGE analysis that a realistic diuron exposure (10 μg/L) could directly and indirectly affect the diversity of a natural riverine bacterial community. Here we extended our first approach by identifying predominant bacterial phylotypes in diuron and control microcosms by sequencing and phylogenetic analysis of bands excised from the previously obtained PCR-TTGE gel. We found a sharp difference between phylotypes obtained from the two types of microcosm. Those that appeared or were maintained only in treated microcosms were mainly γ-Proteobacteria, especially the Pseudomonadaceae family, the Verrucomicrobia and the Gemmatimonadetes. In contrast, phylotypes that appeared only in control microcosms belonged to the Chlamydiae. Methodological aspects related to biases encountered after sequence retrieval from fingerprint gels are also discussed