Genetic relationships among Italian and Mexican maize-rhizosphere Burkholderia cepacia complex (BCC) populations belonging to Burkholderia cenocepacia IIIB and BCC6 group

<p>Abstract</p> <p>Background</p> <p>A close association between maize roots and <it>Burkholderia cepacia </it>complex (BCC) bacteria has been observed in different locations globally. In this study we investigated by MultiLocus Restriction Typing (MLRT) the genetic diversity and relationships among <it>Burkholderia cenocepacia </it>IIIB and BCC6 populations associated with roots of maize plants cultivated in geographically distant countries (Italy and Mexico), in order to provide new insights into their population structure, evolution and ecology.</p> <p>Results</p> <p>The 31 <it>B. cenocepacia </it>IIIB and 65 BCC6 isolates gave rise to 29 and 39 different restriction types (RTs), respectively. Two pairs of isolates of <it>B. cenocepacia </it>IIIB and BCC6, recovered from both Italian and Mexican maize rhizospheres, were found to share the same RT. The eBURST (Based Upon Related Sequence Types) analysis of MLRT data grouped all the <it>B. cenocepacia </it>IIIB isolates into four clonal complexes, with the RT-4-complex including the 42% of them, while the majority of the BCC6 isolates (94%) were grouped into the RT-104-complex. These two main clonal complexes included RTs shared by both Italian and Mexican maize rhizospheres and a clear relationship between grouping and maize variety was also found. Grouping established by eBURST correlated well with the assessment using unweighted-pair group method with arithmetic mean (UPGMA). The standardized index of association values obtained in both <it>B. cenocepacia </it>IIIB and BCC6 suggests an epidemic population structure in which occasional clones emerge and spread.</p> <p>Conclusions</p> <p>Taken together our data demonstrate a wide dispersal of certain <it>B. cenocepacia </it>IIIB and BCC6 isolates in Mexican and Italian maize rhizospheres. Despite the clear relationship found between the geographic origin of isolates and grouping, identical RTs and closely related isolates were observed in geographically distant regions. Ecological factors and selective pressure may preferably promote some genotypes within each local microbial population, favouring the spread of a single clone above the rest of the recombinant population.</p

Selection of nitrogen-fixing deficient Burkholderia vietnamiensis strains by cystic fibrosis patients: involvement of nif gene deletions and auxotrophic mutations

Burkholderia vietnamiensis is the third most prevalent species of the Burkholderia cepacia complex (Bcc) found in cystic fibrosis (CF) patients. Its ability at fixing nitrogen makes it one of the main Bcc species showing strong filiations with environmental reservoirs. In this study, 83% (29 over 35) of the B. vietnamiensis CF isolates and 100% of the environmental ones (over 29) were found expressing the dinitrogenase complex (encoded by the nif cluster) which is essential in N 2 fixation. Among the deficient strains, two were found growing with ammonium chloride suggesting that they were defective in N 2 fixation, and four with amino acids supplements suggesting that they were harbouring auxotrophic mutations. To get insights about the genetic events that led to the emergence of the N 2 -fixing defective strains, a genetic analysis of B. vietnamiensis nitrogen-fixing property was undertaken. A 40-kb-long nif cluster and nif regulatory genes were identified within the B. vietnamiensis strain G4 genome sequence, and analysed. Transposon mutagenesis and nifH genetic marker exchanges showed the nif cluster and several other genes like gltB (encoding a subunit of the glutamate synthase) to play a key role in B. vietnamiensis ability at growing in nitrogen-free media. nif cluster DNA probings of restricted genomic DNA blots showed a full deletion of the nif cluster for one of the N 2 -fixing defective strain while the other one showed a genetic organization similar to the one of the G4 strain. For 17% of B. vietnamiensis clinical strains, CF lungs appeared to have favoured the selection of mutations or deletions leading to N 2 -fixing deficiencies.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/75167/1/j.1462-2920.2007.01240.x.pd

Plant-associated symbiotic Burkholderia species lack hallmark strategies required in mammalian pathogenesis

Burkholderia is a diverse and dynamic genus, containing pathogenic species as well as species that form complex interactions with plants. Pathogenic strains, such as B. pseudomallei and B. mallei, can cause serious disease in mammals, while other Burkholderia strains are opportunistic pathogens, infecting humans or animals with a compromised immune system. Although some of the opportunistic Burkholderia pathogens are known to promote plant growth and even fix nitrogen, the risk of infection to infants, the elderly, and people who are immunocompromised has not only resulted in a restriction on their use, but has also limited the application of non-pathogenic, symbiotic species, several of which nodulate legume roots or have positive effects on plant growth. However, recent phylogenetic analyses have demonstrated that Burkholderia species separate into distinct lineages, suggesting the possibility for safe use of certain symbiotic species in agricultural contexts. A number of environmental strains that promote plant growth or degrade xenobiotics are also included in the symbiotic lineage. Many of these species have the potential to enhance agriculture in areas where fertilizers are not readily available and may serve in the future as inocula for crops growing in soils impacted by climate change. Here we address the pathogenic potential of several of the symbiotic Burkholderia strains using bioinformatics and functional tests. A series of infection experiments using Caenorhabditis elegans and HeLa cells, as well as genomic characterization of pathogenic loci, show that the risk of opportunistic infection by symbiotic strains such as B. tuberum is extremely low

Limited Genetic Diversity in the Endophytic Sugarcane Bacterium Acetobacter diazotrophicus

Acetobacter diazotrophicus isolates that originated from different sugarcane cultivars growing in diverse geographic regions of Mexico and Brazil were shown to have limited genetic diversity. Measurements of polymorphism in the electrophoretic mobilities of metabolic enzymes revealed that the mean genetic diversity per enzyme locus (among the four electrophoretic types distinguished) was 0.064. The results of the genetic analysis indicate that the genetic structure of A. diazotrophicus is clonal, with one largely predominant clone. Plasmids were present in 20 of 24 isolates, and the molecular sizes of the plasmids ranged from 2.0 to 170 kb. Two plasmids (a 20- to 24-kb plasmid detected in all 20 plasmid-containing isolates and a 170-kb plasmid observed in 14 isolates) were highly conserved among the isolates examined. Regardless of the presence of plasmids, all of the isolates shared a common pattern of nif structural gene organization on the chromosome

Development of a real-time PCR method to quantify the PGPR strain Azospirillum lipoferum CRT1 on maize seedlings

International audienceAzospirillum lipoferum CRT1 is a promising phytostimulatory PGPR for maize, whose effect on the plant is cell density-dependent. A nested PCR method is available for detection of the strain but does not allow quantification. The objective was to develop a real-time PCR method for quantification of A. lipoferum CRT1 in the rhizosphere of maize seedlings. Primers were designed based on a strain-specific RFLP marker, and their specificity was verified under qualitative and quantitative PCR conditions based on successful CRT1 amplification and absence of cross-reaction with genomic DNA from various rhizosphere strains. Real-time PCR conditions were then optimized using DNA from inoculated or non-inoculated maize rhizosphere samples. The detection limit was 60 fg DNA (corresponding to 19 cells) with pure cultures and 4 × 104 CFU equivalents g−1 lyophilized sample consisting of mixture of rhizosphere soil and roots. Inoculant quantification was effective down to 104 CFU equivalents g−1. Assessment of CRT1 rhizosphere levels in a field trial was in accordance with estimates from semi-quantitative PCR targeting another locus. This real-time PCR method, which is now available for direct rhizosphere monitoring of A. lipoferum CRT1 in greenhouse and field experiments, could be used as a reference for developing quantification tools for other Azospirillum inoculants