Detailed analysis of the microdiversity of Prochlorococcus populations along a North-South Atlantic Ocean transect

In order to understand how environmental factors shape the diversity of Prochlorococcus in the Atlantic Ocean, we have elucidated the microdiversity along a north–south transect. The polymerase chain reaction-restriction fragment length polymorphism analysis of the genetic diversity of rpoC1 gene fragments of Prochlorococcus at 12 sampling sites revealed a latitudinal pattern in Prochlorococcus RFLP-type diversity in the samples collected from two depths. At the depth to which 14% of surface irradiance penetrated, HLII clones dominated the stations closest to the equator. The percentage of HLI clones increased with distance from the equator and LL clones were found only at the most northern and southern stations. In contrast, deeper (1% light depth) water samples did not show any overall trend in Prochlorococcus diversity or clade dominance. Multivariate statistical analyses indicated that Prochlorococcus diversity was linked to water temperature (partially an effect of latitude) and depth (which was linked to light penetration and turbidity). Phylogenetic analysis of the sequences obtained from the 423 different environmental RFLP-types detected in this study indicated that the HLII and HLI populations were composed of a wide range of genetically different clones, while the LL Prochlorococcus clade was less diverse, although half of the samples screened in this study derived from the 1% light depth.     

High vertical and low horizontal diversity of Prochlorococcus ecotypes in the Mediterranean Sea in summer

Natural populations of the marine cyanobacterium Prochlorococcus exist as two main ecotypes, inhabiting different layers of the ocean's photic zone. These so-called high light- (HL-) and low light (LL-) adapted ecotypes are both physiologically and genetically distinct. HL strains can be separated into two major clades (HLI and HLII), whereas LL strains are more diverse. Here, we used several molecular techniques to study the genetic diversity of natural Prochlorococcus populations during the Prosope cruise in the Mediterranean Sea in the summer of 1999. Using a dot blot hybridization technique, we found that HLI was the dominant HL group and was confined to the upper mixed layer. In contrast, LL ecotypes were only found below the thermocline. Secondly, a restriction fragment length polymorphism analysis of PCR-amplified pcb genes (encoding the major light-harvesting proteins of Prochlorococcus) suggested that there were at least four genetically different ecotypes, occupying distinct but overlapping light niches in the photic zone. At comparable depths, similar banding patterns were observed throughout the sampled area, suggesting a horizontal homogenization of ecotypes. Nevertheless, environmental pcb gene sequences retrieved from different depths at two stations proved all different at the nucleotide level, suggesting a large genetic microdiversity within those ecotypes.     

The phylogenetic and ecological context of cultured and whole genome-sequenced planktonic bacteria from the coastal NW Mediterranean Sea

Microbial isolates are useful models for physiological and ecological studies and can also be used to reassemble genomes from metagenomic analyses. However, the phylogenetic diversity that can be found among cultured marine bacteria may vary significantly depending on the isolation. Therefore, this study describes a set of 136 bacterial isolates obtained by traditional isolation techniques from the Blanes Bay Microbial Observatory, of which seven strains have had the whole genome sequenced. The complete set was compared to a series of environmental sequences obtained by culture-independent techniques (60 DGGE sequences and 303 clone library sequences) previously obtained by molecular methods. In this way, each isolate was placed in both its “ecological” (time of year, nutrient limitation, chlorophyll and temperature values) context or setting, and its “phylogenetic” landscape (i.e. similar organisms that were found by culture-independent techniques, when they were relevant, and when they appeared). Nearly all isolates belonged to the Gammaproteobacteria, Alphaproteobacteria, or the Bacteroidetes (70, 40 and 20 isolates, respectively). Rarefaction analyses showed similar diversity patterns for sequences from isolates and molecular approaches, except for Alphaproteobacteria where cultivation retrieved a higher diversity per unit effort. Approximately 30% of the environmental clones and isolates formed microdiversity clusters constrained at 99% 16S rRNA gene sequence identity, but the pattern was different in Bacteroidetes (less microdiversity) than in the other main groups. Seventeen cases (12.5%) of nearly complete (98–100%) rRNA sequence identity between isolates and environmental sequences were found: nine in the Alphaproteobacteria, five in the Gammaproteobacteria, and three in the Bacteroidetes, indicating that cultivation could be used to obtain at least some organisms representative of the various taxa detected by molecular methods. Collectively, these results illustrated the largely unexplored potential of culturing on standard media for complementing the study of microbial diversity by culture-independent techniques and for obtaining phylogenetically distinct model organisms from natural seawater.     

Diversity of Synechococcus and Prochlorococcus populations determined from DNA sequences of the N-regulatory gene ntcA

The cyanobacteria Synechococcus and Prochlorococcus are abundant primary producers in the nitrogen-poor waters of the Gulf of Aqaba, northern Red Sea. Expression of the nitrogen regulatory gene ntcA is a useful indicator for determining the N-status of cyanobacteria, and preliminary work with this gene suggests that it may also serve as a useful biodiversity marker. Here we investigated the genotypic diversity of ntcA among the full spectrum of cultured Synechococcus and Prochlorococcus lineages and assessed cyanobacterial genotypic composition in environmental samples from the Gulf of Aqaba. The high level of ntcA diversification established this gene as an excellent biodiversity marker capable of distinguishing between numerous clades within each genus with high resolution. An unexpected large diversity was observed among Synechococcus populations, including the detection of four novel clades for which culture representatives have yet to be isolated. In addition, extensive microdiversity within a number of Synechococcus clades was revealed. Temporal differences in the detection of the various Synechococcus clades suggest seasonal fluctuations in the genotypic make-up of Synechococcus populations. In contrast, virtually all Prochlorococcus sequences fell within a single high-light adapted clade that was detected year round. We suggest that the limited genotypic diversity among Prochlorococcus in combination with a limited capacity for acclimation to environmental changes resulting from its small genome size led to the dramatic rise and demise of Prochlorococcus populations over the yearly cycle in the Gulf of Aqaba.     

Resolution of Prochlorococcus and Synechococcus ecotypes by using 16S-23S ribosomal DNA internal transcribed spacer sequences

Cultured isolates of the marine cyanobacteria Prochlorococcus and Synechococcus vary widely in their pigment compositions and growth responses to light and nutrients, yet show greater than 96% identity in their 16S ribosomal DNA (rDNA) sequences. In order to better define the genetic variation that accompanies their physiological diversity, sequences for the 16S-23S rDNA internal transcribed spacer (ITS) region were determined in 32 Prochlorococcus isolates and 25 Synechococcus isolates from around the globe. Each strain examined yielded one ITS sequence that contained two tRNA genes. Dramatic variations in the length and G+C content of the spacer were observed among the strains, particularly among Prochlorococcus strains. Secondary-structure models of the ITS were predicted in order to facilitate alignment of the sequences for phylogenetic analyses. The previously observed division of Prochlorococcus into two ecotypes (called high and low-B/A after their differences in chlorophyll content) were supported, as was the subdivision of the high-B/A ecotype into four genetically distinct clades. ITS-based phylogenies partitioned marine cluster A Synechococcus into six clades, three of which can be associated with a particular phenotype (motility, chromatic adaptation, and lack of phycourobilin). The pattern of sequence divergence within and between clades is suggestive of a mode of evolution driven by adaptive sweeps and implies that each clade represents an ecologically distinct population. Furthermore, many of the clades consist of strains isolated from disparate regions of the world's oceans, implying that they are geographically widely distributed. These results provide further evidence that natural populations of Prochlorococcus and Synechococcus consist of multiple coexisting ecotypes, genetically closely related but physiologically distinct, which may vary in relative abundance with changing environmental conditions.     

Taxonomic resolution, ecotypes and the biogeography of Prochlorococcus

In order to expand our understanding of the diversity and biogeography of Prochlorococcus ribotypes, we PCR-amplified, cloned and sequenced the 16S/23S rRNA ITS region from sites in the Atlantic and Pacific oceans. Ninety-three per cent of the ITS sequences could be assigned to existing Prochlorococcus clades, although many novel subclades were detected. We assigned the sequences to operational taxonomic units using a graduated scale of sequence identity from 80% to 99.5% and correlated Prochlorococcus diversity with respect to environmental variables and dispersal time between the sites. Dispersal time was estimated using a global ocean circulation model. The significance of specific environmental variables was dependent on the degree of sequence identity used to define a taxon: light correlates with broad-scale diversity (90% cut-off), temperature with intermediate scale (95%) whereas no correlation with phosphate was observed. Community structure was correlated with dispersal time between sample sites only when taxa were defined using the finest sequence similarity cut-off. Surprisingly, the concentration of nitrate, which cannot be used as N source by the Prochlorococcus strains in culture, explains some variation in community structure for some definitions of taxa. This study suggests that the spatial distribution of Prochlorococcus ecotypes is shaped by a hierarchy of environmental factors as well dispersal limitation.     

Molecular analysis in the conservation of sturgeons and paddlefish

Sturgeon and paddlefish populations worldwide have declined because of anthropogenic influences. The structure and magnitude of genetic diversity of natural populations serves to buffer these fishes against environmental variation and should be maintained. Modern molecular biological techniques provide the ability to sensitively characterize and quantify the extent of genetic variation in natural populations. We provide a summary of those problems in sturgeon population biology that are amenable to investigation with DNA approaches, and their applications to date. These have included genetic identification and discrimination of taxa, identification of hybrids, stock identification, mixed-stock analysis, and estimation of gene flow and homing fidelity. To date, almost all studies have been restricted to North American fauna. Improvements to these technologies, including nondestructive sampling, should permit more widespread application of molecular approaches to problems of acipenseriform conservation. We suggest that the use of more sensitive molecular tools such as analyses of hypervariable repetitive and non-coding single copy nuclear DNA may assist management even in those taxa which exhibit overall low levels of genetic diversity.     

Synechococcus diversity in the California current as seen by RNA polymerase (rpoC1) gene sequences of isolated strains.

Because they are ubiquitous in a range of aquatic environments and culture methods are relatively advanced, cyanobacteria may be useful models for understanding the extent of evolutionary adaptation of prokaryotes in general to environmental gradients. The roles of environmental variables such as light and nutrients in influencing cyanobacterial genetic diversity are still poorly characterized, however. In this study, a total of 15 Synechococcus strains were isolated from the oligotrophic edge of the California Current from two depths (5 and 95 m) with large differences in light intensity, light quality, and nutrient concentrations. RNA polymerase gene (rpoC1) fragment sequences of the strains revealed two major genetic lineages, distinct from other marine or freshwater cyanobacterial isolates or groups seen in shotgun-cloned sequences from the oligotrophic Atlantic Ocean. The California Current low-phycourobilin (CCLPUB) group represented by six isolates in a single lineage was less diverse than the California Current high-phycourobilin (CCHPUB) group with nine isolates in three relatively divergent lineages. The former was found to be the closest known genetic group to Prochlorococcus spp., a chlorophyll b-containing cyanobacterial group. Having an isolate from this group will be valuable for looking at the molecular changes necessary for the transition from the use of phycobiliproteins to chlorophyll b as light-harvesting pigments. Both of the CCHPUB and CCLPUB groups included strains obtained from surface (5 m) and deep (95 m) samples. Thus, contrary to expectations, there was no clear correlation between sampling depth and isolation of genetic groups, despite the large environmental gradients present. To our knowledge, this is the first demonstration with isolates that genetically divergent Synechococcus groups coexist in the same seawater sample.     

Phylogenetic characterization of bacteria in the gut of house flies (Musca domestica L.)

House flies (Musca domestica L.) are cosmopolitan, ubiquitous, synanthropic insects that serve as mechanical or biological vectors for various microorganisms. To fully assess the role of house flies in the epidemiology of human diseases, it is essential to understand the diversity of microbiota harbored by natural fly populations. This study aimed to identify the diversity of house fly gut bacteria by both culture-dependent and culture-independent approaches. A total of 102 bacterial strains were isolated from the gut of 65 house flies collected from various public places including a garden, public park, garbage/dump area, public toilet, hospital, restaurant/canteen, mutton shop/market, and house/human habitation. Molecular phylogenetic analyses placed these isolates into 22 different genera. The majority of bacteria identified were known potential pathogens of the genera Klebsiella, Aeromonas, Shigella, Morganella, Providencia, and Staphylococcus. Culture-independent methods involved the construction of a 16S rRNA gene clone library, and sequence analyses supported culture recovery results. However, additional bacterial taxa not determined via culture recovery were revealed using this methodology and included members of the classes Alphaproteobacteria, Deltaproteobacteria, and the phylum Bacteroidetes. Here, we show that the house fly gut is an environmental reservoir for a vast number of bacterial species, which may have impacts on vector potential and pathogen transmission.     

Application of a Novel <Emphasis Type="Italic">rpoC1</Emphasis>-RFLP Approach Reveals that Marine <Emphasis Type="Italic">Prochlorococcus</Emphasis> Populations in the Atlantic Gyres are Composed of Greater Microdiversity than Previously Described

To elucidate the degree of microdiversity within the genus Prochlorococcus, novel Prochlorococcus-specific polymerase chain reaction (PCR) primers were developed for the rpoC1 gene, which encodes the ribonucleic acid (RNA) polymerase core subunit. The size of the PCR fragment (925 bp) coupled with high sequence variation within the rpoC1 fragments (70–99% sequence similarity, 16S ribosomal RNA sequences show greater than 97% sequence similarity) meant that it was possible to distinguish Prochlorococcus strains by restriction fragment length polymorphism (RFLP) analysis. Clone libraries were constructed from environmental deoxyribonucleic acid samples from two stations, one in the northern and one in the southern oligotrophic gyre of the Atlantic Ocean. These were screened to determine the microdiversity of Prochlorococcus populations using this high-resolution high-throughput analysis approach. RFLP analysis of the clone libraries from the two gyre sites revealed that the two Prochlorococcus populations had a high degree of microdiversity with 40 and 52 different RFLP-type clones among the 143 clones tested for both the northern and southern gyres, respectively. Phylogenetic analysis of the nucleotide sequences of the RFLP types not only showed that it contained representatives of each of the currently recognized Prochlorococcus clades (based on the internal transcribed spacer region as molecular marker) but also led to the discovery of a previously unseen genetic microdiversity. This level of diversity was greater at the southern gyre site compared to the northern gyre site. Moreover, the high genetic resolution approach also revealed that there are two putative novel lineages within the HL I clade. Analyses of further samples by producing clone libraries from different geographic origins is likely to reveal further diversity and novel lineages within Prochlorococcus.     

Effects of Culturing on the Population Structure of a Hyperthermophilic Virus

The existence of a culturing bias has long been known when sampling organisms from the environment. This bias underestimates microbial diversity and does not accurately reflect the most ecologically relevant species. Until now no study has examined the effects of culture bias on viral populations. We have employed culture-independent methods to assess the diversity of Sulfolobus spindle–shaped viruses (SSVs) from extremely hyperthermal environments. This diversity is then compared to the viral diversity of cultured samples. We detected a clear culturing bias between environmental samples and cultured isolates. This is the first study identifying a culture bias in a viral population.     

Coupling 16S-ITS rDNA clone libraries and automated ribosomal intergenic spacer analysis to show marine microbial diversity: development and application to a time series

We outline an approach to simultaneously assess multilevel microbial diversity patterns utilizing 16S-ITS rDNA clone libraries coupled with automated ribosomal intergenic spacer analysis (ARISA). Sequence data from 512 clones allowed estimation of ARISA fragment lengths associated with bacteria in a coastal marine environment. We matched 92% of ARISA peaks (each comprising >1% total amplified product) with corresponding lengths from clone libraries. These peaks with putative identification accounted for an average of 83% of total amplified community DNA. At 16S rDNA similarities <98%, most taxa displayed differences in ARISA fragment lengths >10 bp, readily detectable and suggesting ARISA resolution is near the 'species' level. Prochlorococcus abundance profiles from ARISA were strongly correlated (r2=0.86) to Prochlorococcus cell counts, indicating ARISA data are roughly proportional to actual cell abundance within a defined taxon. Analysis of ARISA profiles for 42 months elucidated patterns of microbial presence and abundance providing insights into community shifts and ecological niches for specific organisms, including a coupling of ecological patterns for taxa within the Prochlorococcus, the Gamma Proteobacteria and Actinobacteria. Clade-specific ARISA protocols were developed for the SAR11 and marine cyanobacteria to resolve ambiguous identifications and to perform focused studies. 16S-ITS data allowed high-resolution identification of organisms by ITS sequence analysis, and examination of microdiversity.     

Thermus thermophilus TMY isolated from silica scale taken from a geothermal power plant

Aims:  To identify an extreme thermophile, strain TMY, isolated from silica scale from the geothermal electric power plant and to examine microdiversity of Thermus thermophilus strains.
Materials and Results:  The isolated strain TMY was identified by morphological, biochemical and physiological tests. Phylogenetic comparison of the strain and other Thermus strains with 16S rDNA analysis, RAPD and ERIC-PCR fingerprinting were performed. Strain TMY was closely related to strain which was isolated from a hot spring in New Zealand and shown to belong to the Japanese Thermus cluster. However, there were considerable genetic differences between strain TMY and other Thermus species using DNA fingerprinting.
Conclusions:  Based on morphological, physiological and genetic properties, strain TMY could be a strain of T. thermophilus . The distinct properties of strain TMY suggest that microdiversity of T. thermophilus strains should be considered.
Significance and Impact of the Study:  The results of this study have demonstrated genetic diversity within T. thermophilus strains, which were previously masked by an almost identical 16S rDNA sequence. RAPD and ERIC-PCR could be potential methods for distinguishing between Thermus strains.     

Culture-dependent and culture-independent diversity within the obligate marine actinomycete genus Salinispora

Salinispora is the first obligate marine genus within the order Actinomycetales and a productive source of biologically active secondary metabolites. Despite a worldwide, tropical or subtropical distribution in marine sediments, only two Salinispora species have thus far been cultivated, suggesting limited species-level diversity. To further explore Salinispora diversity and distributions, the phylogenetic diversity of more than 350 strains isolated from sediments collected around the Bahamas was examined, including strains cultured using new enrichment methods. A culture-independent method, using a Salinispora-specific seminested PCR technique, was used to detect Salinispora from environmental DNA and estimate diversity. Overall, the 16S rRNA gene sequence diversity of cultured strains agreed well with that detected in the environmental clone libraries. Despite extensive effort, no new species level diversity was detected, and 97% of the 105 strains examined by restriction fragment length polymorphism belonged to one phylotype (S. arenicola). New intraspecific diversity was detected in the libraries, including an abundant new phylotype that has yet to be cultured, and a new depth record of 1,100 m was established for the genus. PCR-introduced error, primarily from Taq polymerase, significantly increased clone library sequence diversity and, if not masked from the analyses, would have led to an overestimation of total diversity. An environmental DNA extraction method specific for vegetative cells provided evidence for active actinomycete growth in marine sediments while indicating that a majority of sediment samples contained predominantly Salinispora spores at concentrations that could not be detected in environmental clone libraries. Challenges involved with the direct sequence-based detection of spore-forming microorganisms in environmental samples are discussed.     

Genetic diversity of Bradyrhizobium strains isolated from Arachis hypogaea

Rhizobia are used exclusively in agricultural systems for enhancing the ability of legumes to fix atmospheric nitrogen. Knowledge about the indigenous population is necessary for the selection and application of inoculant strains. In this study, we have assessed the genetic diversity of Bradyrhizobium strains isolated from the host plant, Arachis hypogaea along the coastline of Tamil Nadu. Different populations collected from varying environmental conditions were analysed for salt and pH tolerance. Genetic diversity among the strains was studied using RAPD markers and PCR-RFLP of 16S rDNA and nifD genes. The approaches used in this study yielded consistent results, which revealed a high degree of heterogeneity among strains and detection of two distinct genetic groups.     

Establishment of “The Gene Mine”: a resource for rapid identification of complex trait genes

Identification of genes underlying complex traits presents a challenge to which geneticists have responded with many diverse approaches. A common feature of these approaches is that different research groups must, on a case-by-case basis, replicate similar efforts in recruitment, genetic characterization, and analyses. To avoid this expensive “churning,” an alternative approach has been proposed: production of an experimental genetic reference population, the Collaborative Cross, in which both genetic diversity and mapping power are maximized. Since this population consists of inbred mouse strains, further advantages are that it is essentially infinitely reproducible; genetic characterization needs to be performed only once; and the founder strains’ genomes have been or will be sequenced, allowing imputation of allele sequences of all members of the reference population. Here we describe the establishment of such a genetic reference population, which we dub “The Gene Mine.” Over 1000 genetically distinct lines have been established, descended from eight diverse founder strains. Preliminary phenotypic ascertainment of these strains indicates unexpected variability arising from independent assortment of genetic variants. The Gene Mine will be a powerful resource for characterization of essentially any mouse phenotype that has a genetic basis.     

Genetic and phenotypic microdiversity of Ochrobactrum spp

The diversity of Ochrobactrum anthropi, Ochrobactrum intermedium, Ochrobactrum tritici and Ochrobactrum grignonense in agricultural soil and on the wheat rhizoplane was investigated. O. anthropi was isolated both from soil and from the rhizoplane, O. intermedium and grignonense only from bulk soil, and O. tritici only from the wheat rhizoplane. On the genetic level, the immunotrapped isolates and a number of strains from culture collection mainly of clinical origin were compared with rep-PCR profiling using BOX primers, and a subset of these isolates and strains using REP primers. The isolates clustered according to their species affiliation. There was no correlation between rep clusters of O. anthropi isolates and habitat (place of isolation). The genetic diversity of Ochrobactrum at the species level as well as microdiversity of O. anthropi (number of BOX groups) was higher in soil than on the rhizoplane. Similarity values from genetic rep-PCR profiles correlated positively with DNA-DNA reassociation percentages. Isolates with >80.7% similarity in BOX profile and >86.4% in rep profile clustered within the same species. Similarity analysis of rep-PCR profiles is hence an alternative to DNA-DNA hybridization as a genomic criterion for species delineation within the genus Ochrobactrum. We used the substrate utilization system BIOLOG-GN to compare the immunotrapped isolates on the phenetic level. For the isolates from bulk soil, substrate utilization versatility (number of utilized substrates) and substrate utilization capacity (mean conversion rate of substrates) were slightly but significantly higher than for the isolates from the rhizoplane. This trend was also seen using API 20E and 20NE systems. Plate counts of total bacteria and the number of immunotrapped Ochrobactrum isolates per gram dry weight were higher for the rhizoplane than for the soil samples. The results of genetic and phenotypic analyses indicated a 'rhizosphere effect'; the diversity and metabolic capacity of Ochrobactrum isolates were higher in bulk soil, and the population density was higher on the wheat rhizoplane.     

Natural community structure of cyanobacteria in the South China Sea as revealed by rpoC1 gene sequence analysis

AIMS: To examine the community structure of cyanobacterial populations of the South China Sea on the surface and at depth of 80 m. METHODS AND RESULTS: Direct PCR amplification of RNA polymerase (rpoC1) genes from environmental DNAs extracted from seawater, and cloning of the fragments and sequence analysis were used. A great diversity of Prochlorococcus and Synechococcus were detected at the investigation site. Genetically related Prochlorococcus were found in both layers while Synechococcus were found only on the surface. Prochlorococcus were clustered with the known high-light adapted II genotypes, and further divided into seven groups. Synechococcus could be divided into two groups, and the second group could be further subdivided into several clades. CONCLUSION: The dominant genotype of Prochlorococcus was high-light adapted II genotype, and Synechococcus were distributed basically on the surface. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report about the cyanobacterial community structure in the South China Sea, and an important supplement to the current understanding of the relationship between genetic and ecological diversity and environments.     

91 Maintenance of genetic diversity in heterosigma akashiwo by temporal variability in its environment

Heterosigma akashiwo (Hada) Hada ex Y. Hara et Chihara (Chromophyta: Raphidophyceae) is a coastal HAB forming species that is not known to undergo sexual reproduction. Ribosomal ITS sequences for 36 strains of Heterosigma have been determined and found to be identical except for two strains that had one single nucleotide polymorphism each. Despite the apparent absence of meiosis and the ostensible homogeneity of genotypes (at least at the ITS locus), different strains of Heterosigma display a very wide range of phenotypes. This inconsistency suggests that greater genetic diversity exists in Heterosigma than is evident through sequence analysis of the ITS region. Data will be presented summarizing efforts to develop better genetic markers for examining intrapopulational diversity. Results of experiments examining the effects of nutrient concentration in batch culture on growth response have demonstrated that sufficient haplotype variability exists in established culture strains to allow Darwinian selection, even when these strains are "clonal". We propose that batch culture imposes a sequence of different selection pressures on a rapidly dividing population of cells, and this environmental variation promotes the maintenance of genetic diversity. Extrapolating the batch culture scenario to the much more highly dynamic marine coastal environment, it becomes apparent that the observed phenotypic variation between Heterosigma strains is largely a function of a diverse haplotype distribution, maintained, in a large part, by patchy resource availability on both temporal and spatial scales.     

Multiple Infection and Microdiversity among Helicobacter pylori Isolates in a Single Host in India

Helicobacter pylori is one of the most diverse bacterial species that chronically infects more than 70% of Indian population. Interestingly, data showing microdiversity of the H. pylori strains within a particular gastric niche remained scarce. To understand the extent of genetic diversity among H. pylori strains within a given host, 30 patients with gastro-duodenal problems were subjected to endoscopy and from each patient 10 single colonies were isolated. Characterization of each of these 10 single colonies by DNA fingerprinting as well as genotyping of several important genetic markers viz. cagA, vacA, iceA, vapD, cag PAI empty site, IS605, RFLP and two other genetic segments within cag PAI revealed that all of the 30 patients were infected with more than one strain and sometimes strains with 5 to 6 types of genetic variants. Analyses of certain genetic loci showed the microdiversity among the colonies from single patient, which may be due to the recombination events during long-term carriage of the pathogen. These results suggest that most of the patients have acquired H. pylori due to repeated exposure to this pathogen with different genetic make-up, which may increase the possibility of super infections. Genetic exchanges between these unrelated H. pylori strains may support certain H. pylori variant to grow better in a given host than the parental strain and thereby increasing the possibility for the severity of the infection.