Genetic diversity in Monilinia laxa populations in stone fruit species in Hungary

The objectives of this study were firstly, to determine the genetic diversity of Monilinia laxa isolates from Hungary, using the PCR-based inter-simple sequence repeat (ISSR) and randomly amplified polymorphic DNA (RAPD) technique; secondly, to prepare genetic diversity groups based on the dendrograms; and finally, to select some relevant isolates to study their fungicide sensitivity. 55 and 77 random amplified polymorphic ISSR and RAPD markers, of which 23 and 18 were polymorphic and 32 and 59 monomorphic, respectively, were used to assess the genetic diversity and to study the structure of M. laxa populations in Hungary. 27 isolates out of 57 ones were confirmed as M. laxa from several orchards (subpopulations) in three geographical regions, in various inoculum sources and in various hosts, were used. 10 fungicides and 12 isolates selected from genetic diversity groups based on the ISSR dendrograms were used to determine the fungicide sensitivity of the selected isolates. The analysis of population structure revealed that genetic diversity within locations, inoculum sources and host (H _S ) accounted for 99 % of the total genetic diversity (H _T ), while genetic diversity among locations, inoculum sources and host represented only 1 %. The relative magnitude of gene differentiation between subpopulations (G _ST ) and the estimate of the number of migrants per generation (Nm) averaged 0.005–0.009 and 53.9–99.2, respectively, for both ISSR and RAPD data set. The results obtained in dendrograms were in accordance with the gene diversity analysis. Grouping of isolates in the dendrograms was irrespective of whether they came from the same or different geographical locations. There was no relationship between clustering among isolates from inoculum sources and hosts. In the fungicide sensitivity tests, five isolates out of 12 were partly insensitive to boscalid+piraclostrobin, cyprodinil, fenhexamid or prochloraz. Obtained results in genetic diversity of M. laxa populations are discussed together with implications for the management of brown rot.     

Genetic diversity among clonal isolates of Karenia brevis as measured with microsatellite markers

Karenia brevis is the major harmful bloom-forming dinoflagellate in the Gulf of Mexico yet little is known about the intraspecific genetic diversity of this species. Here we describe nine new microsatellite markers and, combined with nine previously described microsatellites, use them to genotype 40 cultured isolates of K. brevis. Genetic diversity identified from cultured isolates was compared with the genetic diversity identified from two field samples to assess how well the current cultures represent the field population. Thirty-nine unique haplotypes were identified from 40 cultured isolates of K. brevis using 18 microsatellite markers. Genetic diversity was similar between cultured isolates and the two field samples. The success of 18 microsatellite markers to distinguish individual isolates supports the use of microsatellites as a genetic tool for diagnostic identification of cultured isolates of K. brevis.     

Distributions of Salmonella Subtypes Differ between Two U.S. Produce-Growing Regions

Salmonella accounts for approximately 50% of produce-associated outbreaks in the United States, several of which have been traced back to contamination in the produce production environment. To quantify Salmonella diversity and aid in identification of Salmonella contamination sources, we characterized Salmonella isolates from two geographically diverse produce-growing regions in the United States. Initially, we characterized the Salmonella serotype and subtype diversity associated with 1,677 samples collected from 33 produce farms in New York State (NYS). Among these 1,677 samples, 74 were Salmonella positive, yielding 80 unique isolates (from 147 total isolates), which represented 14 serovars and 23 different pulsed-field gel electrophoresis (PFGE) types. To explore regional Salmonella diversity associated with production environments, we collected a smaller set of samples (n = 65) from South Florida (SFL) production environments and compared the Salmonella diversity associated with these samples with the diversity found among NYS production environments. Among these 65 samples, 23 were Salmonella positive, yielding 32 unique isolates (from 81 total isolates), which represented 11 serovars and 17 different PFGE types. The most common serovars isolated in NYS were Salmonella enterica serovars Newport, Cerro, and Thompson, while common serovars isolated in SFL were Salmonella serovars Saphra and Newport and S. enterica subsp. diarizonae serovar 50:r:z. High PFGE type diversity (Simpson''s diversity index, 0.90 ± 0.02) was observed among Salmonella isolates across both regions; only three PFGE types were shared between the two regions. The probability of three or fewer shared PFGE types was <0.000001; therefore, Salmonella isolates were considerably different between the two sampled regions. These findings suggest the potential for PFGE-based source tracking of Salmonella in production environments.     

Variability of United States Isolates of Macrophomina phaseolina Based on Simple Sequence Repeats and Cross Genus Transferability to Related Genera Within Botryosphaeriaceae

Twelve simple sequence repeat (SSRs) loci were used to evaluate genetic diversity of 109 isolates of Macrophomina phaseolina collected from different geographical regions and host species throughout the United States (US). Genetic diversity was assessed using Nei’s minimum genetic distance, and the usefulness of each locus was determined by calculating the polymorphism information content (PIC). A total of 98 alleles were detected and of these 31 were unique to individual genotypes. Eight of twelve loci were highly informative with PIC values greater than 0.50. The majority of pairwise comparisons of genetic distance were greater than 0.60 indicating moderate to high genetic diversity. Dendrograms based on the genetic dissimilarities were created for the 109 isolates of which 79 were from soybean. Some clustering by host and geography was noted, but, the dendrograms generally grouped isolates independent of host or geography. Additionally, sequencing of the internal transcribed spacer region (ITS) for 10 isolates revealed that all of these isolates were 99% similar. Three SSR loci from M. phaseolina were cross amplified in other genera in the Botryosphaeriaceae. This was the first study of genotyping and assessing genetic diversity of M. phaseolina isolates collected from a widespread host and geographic range across the US with SSRs. With an additional 34 loci publically available for M. phaseolina, the results indicate that previously developed SSRs from one species can be used in future population, ecological, and genetic studies of M. phaseolina and other genera within the Botryosphaeriaceae.     

Leishmania donovani: Genetic diversity of isolates from Sudan characterized by PCR-based RAPD

Drug unresponsiveness in patients with visceral leishmaniasis (VL) is a problem in many endemic areas. This study aimed to determine genetic diversity of Leishmania donovani isolates from a VL endemic area in Sudan as a possible explanation for drug unresponsiveness in some patients. Thirty clinically stibogluconate (SSG)-sensitive isolates were made SSG-unresponsive in vitro by gradually increasing SSG concentrations. The sensitive isolates and their SSG-unresponsive counterparts were typed using mini-circle kDNA and categorized using PCR-RAPD. All the isolates were typed as L. donovani, the resulting PCR-RAPD characterization of the SSG-sensitive isolates gave three distinct primary genotypes while, the SSG-unresponsive isolates showed only a single band. L. donovani isolates from eastern Sudan are diverse; this probably resulted from emergence of new L. donovani strains during epidemics due to the pressure of widespread use of antimonials.In this communication the possible role of isolates diversity in antimonial unresponsiveness and the in vitro changing PCR-RAPD band pattern in SSG-unresponsive strains were discussed.     

Molecular Analysis of Rhynchosporium secalis Population Collected from Barley Cultivars having Different Resistance Specificities

Molecular analysis was performed to detect genetic diversity in 106 Rhynchosporium secalis isolates collected from different regions of Canada using random amplified polymorphic DNA (RAPD) markers. The isolates collected from barley cultivars having different resistance specificity to R. secalis and grown in geographically distinct regions, exhibited reproducible variation for 2–3 polymorphic PCR products per decamer primer. Analysis of 1960 RAPD markers data obtained with five primers formed 5 groups with different genetic similarity. High genetic variation was observed in R. secalis isolates obtained from resistant and susceptible cultivars of barley. Isolates collected from susceptible cultivars showed a tendency to group together, whereas isolates from resistant cultivars were divergent. R. secalis isolates infecting different barley cultivars released as resistant to the barley scald formed a specific group with UPGMA, even though all these isolates were collected from the same epidemiological region. Analysis of 15 isolates collected from one resistant cultivar Duke formed three clusters with low bootstrap values indicating high genetic diversity among the isolates present on a single host cultivar.     

Genetic characterization of grapevine-infecting Botrytis cinerea isolates from Argentina

BackgroundBotrytis cinerea is an ascomycete with a high genetic diversity and complex population structure, as reported from several hosts and sites. However, nothing is known about its genetic diversity in Argentina.AimsThe aim of this work is to estimate the genetic diversity of a local population of B. cinerea isolates obtained from grapevine in Argentina.MethodsIn this work, 35 strains that had been isolated from grapevines were genotyped for the presence of transposable elements and PCR-based RFLP molecular markers. The obtained results were compared with those from a large French population of the fungus, and used to perform a population genetics analysis using the Genepop software.ResultsAll the analysed isolates were classified as Group II (according to the most recent proposed classification) and showed a high degree of genetic diversity, with 14 different haplotypes. A significant difference in allele frequency was recorded between the local and French populations.ConclusionsThese comparisons between fungal populations, led to the detection of a high level of diversity and the differentiation between local and French groups of isolates. This was confirmed by an Fst value of 0.3332, which was higher than that reported for other pairwise comparisons of populations. This work constitutes the first report on the genetic diversity of B. cinerea isolates and their population structure in Argentina.     

Genetic Diversity of Amylomyces rouxii from Ragi tapai in Java Island Based on Ribosomal Regions ITS1/ITS2 and D1/D2

Amylomyces rouxii is commonly found as amylolytic fungi in tapai fermentation. However, its diversity is rarely reported despite being often used for food production in Southeast Asia. This research aims to analyze the genetic diversity and the distribution pattern of A. rouxii from Ragi tapai in Java Island, Indonesia. We isolated the fungus from samples obtained from Ragi tapai producing centers in Bandung, Sumedang, Muntilan, Blora, Yogyakarta, and Bondowoso. The obtained isolates were molecularly identified based on the ribosomal regions ITS1/ITS2 and D1/D2, then analyzed for phylogenetic tree reconstruction, genetic distance, genetic variation, and haplotype networking. Six isolates showed specific morphological traits of A. rouxii. However, phylogenetic tree reconstruction on the ribosomal genes showed that the isolates were grouped into two different clades related to two species. Clade A included BDG, SMD, and MTL isolates related to A. rouxii, whereas clade B included YOG, BLR, and BDS isolates related to Mucor indicus. The genetic distances between clades for ITS1/ITS2 and D1/D2 were 0.6145 and 0.1556, respectively. In conclusion, we confirmed the genetic diversity of molds from Ragi tapai in Java Island and showed that the isolates are not only related to A. rouxii as reported before.     

Genetic diversity of Metarhizium anisopliae var. anisopliae in southwestern British Columbia

The abundance and genetic diversity of the entomopathogenic fungus, Metarhizium anisopliae var. anisopliae, in southwestern British Columbia (BC) and southern Alberta was examined. The fungus was found to be widespread in soil throughout southwestern BC, and was recovered from 56% of 85 sample sites. In contrast to southwestern BC, no M. anisopliae isolates were recovered in southern Alberta. An automated fluorescent amplified fragment length polymorphism (AFLP) method was used to examine genetic diversity. In excess of 200 isolates were characterized. The method identified 211 polymorphic amplicons, ranging in size from ≈92 to 400 base pairs, and it was found to be reproducible with a resolution limit of 86.2% similarity. The AFLP method distinguished Metarhizium from other entomopathogenic fungal genera, and demonstrated considerable genetic diversity (25 genotypes) among the reference strains of M. anisopliae isolates examined (i.e. recovered from various substrates and geographical locations). Although 13 genotypes of M. anisopliae var. anisopliae were recovered from southwestern BC soils, the vast majority of isolates (91%) belonged to one of two closely-related genotypes. Furthermore, these two genotypes predominated in urban, agricultural and forest soils. The reasons for the limited diversity of M. anisopliae var. anisopliae in southwestern BC are uncertain. However, findings of this study are consistent with island biogeography theory, and have significant implications for the development of this fungus for microbial control of pest insects.     

Genetic Polymorphism Characteristics of Brucella canis Isolated in China

In China, brucellosis is an endemic disease typically caused by Brucella melitensis infection (biovars 1 and 3). Brucella canis infection in dogs has not traditionally recognized as a major problem. In recent years however, brucellosis resulting from Brucella canis infection has also been reported, suggesting that infections from this species may be increasing. Data concerning the epidemiology of brucellosis resulting from Brucella canis infection is limited. Therefore, the purpose of this study was to assess the diversity among Chinese Brucella canis strains for epidemiological purposes. First, we employed a 16-marker VNTR assay (Brucella MLVA-16) to assess the diversity and epidemiological relationship of 29 Brucella canis isolates from diverse locations throughout China with 38 isolates from other countries. MLVA-16 analysis separated the 67 Brucella canis isolates into 57 genotypes that grouped into five clusters with genetic similarity coefficients ranging from 67.73 to 100%. Moreover, this analysis revealed a new genotype (2-3-9-11-3-1-5-1:118), which was present in two isolates recovered from Guangxi in 1986 and 1987. Second, multiplex PCR and sequencing analysis were used to determine whether the 29 Chinese Brucella canis isolates had the characteristic BMEI1435 gene deletion. Only two isolates had this deletion. Third, amplification of the omp25 gene revealed that 26 isolates from China had a T545C mutation. Collectively, this study reveals that considerable diversity exists among Brucella canis isolates in China and provides resources for studying the genetic variation and microevolution of Brucella.     

Assessment of ecological diversity of rhizobacterial communities in vermicompost and analysis of their potential to improve plant growth

Present study was designed to determine the microbial diversity from three distinctive sites (amended with vermicompost) of Gujarat, India. A set of 76 strains were screened from total of 438 strains that exhibit plant growthpromoting (PGP) and antagonistic potential isolated from sites PS1 (Mehsana district), BS2 (Dantiwada district) and VS3 (Gandhinagar district). Their diversity indices were studied for determining the species richness and evenness of screened isolates. Results revealed that site BS2 showed the most significant diversity indices in terms of Shannon (H′ 1.525) and Simpson (1/D 5.120) than other two samples. Principal component analysis showed that bacterial diversity (H′) was correlated with the soil characteristics. Chickpea and groundnut plants inoculated with MBCU1 and MBCU3 isolates showed an increase in the vegetative growth parameters that evaluate plant growth when compared to uninoculated controls. Strains MBCU1 and MBCU3 were identified as Pseudomonas stutzeri and Pseudomonas mosselii, respectively, according to sequence analysis of the 16S rRNA gene. These both isolates belong to site BS2 and they showed specific PGP traits suggesting that these isolates can promote plant growth by more than one mechanism with respect to their higher diversity index.     

Exploring the biodiversity of two groups of Oenococcus oeni isolated from grape musts and wines: Are they equally diverse?

One hundred and four Oenococcus oeni isolates were characterised by the carbohydrate fermentation (CH) profile and DNA fingerprinting. Forty-four isolates came from grape must, and 60 from wines sampled at the end of alcoholic fermentation or during malolactic fermentation. The grape must isolates fermented more CH than the wine isolates. In genotypical terms, no clear boundary between grape must and wine isolates was found. Diversities were deduced by considering the isolates of grape must and of wine separately and jointly. By considering only CH fermentation abilities, the group of grape must isolates gave higher diversity index (DI_CH) values than those isolated from wine; i.e., these isolates were metabolically more diverse. The contrary occurred when the DNA fingerprints were used to calculate DI_RAPD-VNTR: wine isolates were genotypically more diverse than grape must ones. With a polyphasic approach, which considered metabolic and genotypic data, the diversity index of both isolate groups (from grape must and wine) was the same, 0.993, which was slightly lower than that calculated from all the isolates (0.997).     

Diversity of Methicillin-Resistant Staphylococcus aureus (MRSA) Strains Isolated from Inpatients of 30 Hospitals in Orange County,California

There is a need for a regional assessment of the frequency and diversity of MRSA to determine major circulating clones and the extent to which community and healthcare MRSA reservoirs have mixed. We conducted a prospective cohort study of inpatients in Orange County, California, systematically collecting clinical MRSA isolates from 30 hospitals, to assess MRSA diversity and distribution. All isolates were characterized by spa typing, with selective PFGE and MLST to relate spa types with major MRSA clones. We collected 2,246 MRSA isolates from hospital inpatients. This translated to 91/10,000 inpatients with MRSA and an Orange County population estimate of MRSA inpatient clinical cultures of 86/100,000 people. spa type genetic diversity was heterogeneous between hospitals, and relatively high overall (72%). USA300 (t008/ST8), USA100 (t002/ST5) and a previously reported USA100 variant (t242/ST5) were the dominant clones across all Orange County hospitals, representing 83% of isolates. Fifteen hospitals isolated more t008 (USA300) isolates than t002/242 (USA100) isolates, and 12 hospitals isolated more t242 isolates than t002 isolates. The majority of isolates were imported into hospitals. Community-based infection control strategies may still be helpful in stemming the influx of traditionally community-associated strains, particularly USA300, into the healthcare setting.     

Evaluation of molecular identification of Aspergillus species causing fungal keratitis

Fungal keratitis caused by the species of Aspergillus is a common and leading problem in developing countries like India. In this study, a total of 135 isolates from Aspergillus keratitis were studied by sequence analyses of the internal transcribed spacer (ITS) region performed by nucleotide-nucleotide BLAST analysis followed by the initial identification of the isolates based on conidial and colony morphology. The sequence analysis revealed several unusual species which were never reported in eye infections such as A. tamrii, A. tubingensis, A. braslliensis, A. nomius, A. pseudonomius, A. sydowii, Eurotium amstelodami. The sequence analysis of the ITS region; the β-tubulin and calmodulin genes brought out the genetic diversity among the isolates as the study intended to locate a more sensitive target sequence to study genetic diversity among a set of test fungal isolates. The PCR amplified sequences of the test isolates of the study as well as sequences belonging to section Flavi obtained from Genbank database were compared and analyzed along with three standard isolates by phylogenetic tree (Neighbor-joining) as to find out a target region/gene that could produce a better resolution to differentiate the isolates. Accordingly, the calmodulin gene had provided better resolution compared to ITS and β-tubulin to study the diversity among the test Aspergillus species isolated from fungal corneal ulcer.     

Genetic Diversity of Mycobacterium tuberculosis from Guadalajara,Mexico and Identification of a Rare Multidrug Resistant Beijing Genotype

Determining the genetic diversity of M. tuberculosis strains allows identification of the distinct Mycobacterium tuberculosis genotypes responsible for tuberculosis in different regions. Several studies have reported the genetic diversity of M. tuberculosis strains in Mexico, but little information is available from the state of Jalisco. Therefore, the aim of this study was to determine the genetic diversity of Mycobacterium tuberculosis clinical isolates from Western Mexico. Sixty-eight M. tuberculosis isolates were tested for susceptibility to first-line drugs using manual Mycobacteria Growth Indicator Tube method and genotyped using spoligotyping and IS6110-restriction fragment length polymorphism (RFLP) pattern analyses. Forty-seven (69.1%) isolates were grouped into 10 clusters and 21 isolates displayed single patterns by spoligotyping. Three of the 21 single patterns corresponded to orphan patterns in the SITVITWEB database, and 1 new type that contained 2 isolates was created. The most prevalent lineages were T (38.2%), Haarlem (17.7%), LAM (17.7%), X (7.4%), S (5.9%), EAI (1.5%) and Beijing (1.5%). Six (12.8%) of the clustered isolates were MDR, and type 406 of the Beijing family was among the MDR isolates. Seventeen (26.2%) isolates were grouped into 8 clusters and 48 isolates displayed single patterns by IS6110-RFLP. Combination of IS6110-RFLP and spoligotyping reduced the clustering rate to 20.0%. The results show that T, Haarlem, and LAM are predominant lineages among clinical isolates of M. tuberculosis in Guadalajara, Mexico. Clustering rates indicated low transmission of MDR strains. We detected a rare Beijing genotype, SIT406, which was a highly resistant strain. This is the first report of this Beijing genotype in Latin America.     

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.     

Genetic diversity of root nodule bacteria nodulating Lotus corniculatus and Anthyllis vulneraria in Sweden

Very little is known about the genetic diversity and phylogeny of rhizobia nodulating Lotus species in northern temperate regions. We have therefore studied the genetic diversity among a total of 61 root nodule bacteria isolated from Lotus corniculatus and Anthyllis vulneraria from different geographic sites and habitats in Sweden by restriction fragment length polymorphism (RFLP) of the internal transcribed spacer between their 16S rRNA and 23S rRNA (IGS) region. A high diversity consisting of 26 IGS types from 54 L. corniculatus isolates and five IGS types from seven A. vulneraria isolates was found. The 16S rRNA sequences and phylogeny of representatives of the different IGS types showed four interesting exceptions from the majority of the isolates belonging to the genus Mesorhizobium: Two isolates were both found to be closely related to Rhodococcus spp., and two other isolates showed close relationship with Geobacillus spp. and Paenibacillus spp., respectively. The nodA sequences and phylogeny showed that all the isolates, including those not belonging to the traditional rhizobia genera, harbored nodA sequences which were typical of Mesorhizobium loti. Generally, the 16S rRNA and nodA phylogenetic trees were not congruent in that isolates with similar 16S rRNA sequences were associated with isolates harboring different nodA sequences. All the isolates were confirmed to nodulate L. corniculatus in an inoculation test. This is the first report of members of these non-rhizobia genera being able to nodulate legumes, and we suggest that they may have acquired their nodulating properties through lateral gene transfer.     

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.     

Epidemiology of Rhodococcus equi Strains on Thoroughbred Horse Farms

Pulsed-field gel electrophoresis of restriction endonuclease-digested genomic DNA from a large collection of clinical isolates of Rhodococcus equi, an important pathogen of foals, was used to compare strain distribution between farms and over time. Forty-four strains were found among 209 isolates, with 5 of these accounting for over half the isolates and the 22 strains isolated more than once accounting for 90% of the isolates. The average genotypic diversity on each farm and in each year was found to be less than the genotypic diversity of the isolates taken as a whole, with 5.2% of the total diversity being due to differences between farms and 5.5% to differences between years. A small number of strains on each farm were found to have caused at least half the clinical cases of disease, and these varied between farms and, to a lesser extent, years. Most strains were found on more than one farm, and some very similar restriction patterns were found among isolates from different continents, indicating that strains can be very widespread. Multiple strains were isolated in five of the six cases in which more than one isolate from a single foal was examined, indicating that disease may commonly be caused by simultaneous infection with multiple strains. It was concluded that there are a number of different strains of R. equi which carry the vapA gene, and these strains tend to be widespread, but individual farms tend to have particular strains associated with them.     

Evaluation of Genetic Diversity among Pseudomonas citronellolis Strains Isolated from Oily Sludge-Contaminated Sites

The diversity among a set of bacterial strains that have the capacity to degrade total petroleum hydrocarbons (TPH) in soil contaminated with oily sludge (hazardous hydrocarbon waste from oil refineries) was determined. TPH is composed of alkane, aromatics, nitrogen-, sulfur-, and oxygen-containing compound, and asphaltene fractions of crude oil. The 150 bacterial isolates which could degrade TPH were isolated from soil samples obtained from diverse geoclimatic regions of India. All the isolates were biochemically characterized and identified with a Biolog microbial identification system and by 16S rDNA sequencing. Pseudomonas citronellolis predominated among the 150 isolates obtained from six different geographically diverse samplings. Of the isolates, 29 strains of P. citronellolis were selected for evaluating their genetic diversity. This was performed by molecular typing with repetitive sequence (Rep)-based PCR with primer sets ERIC (enterobacterial repetitive intergenic consensus), REP (repetitive extragenic palindromes), and BOXAIR and PCR-based ribotyping. Strain-specific and unique genotypic fingerprints were distinguished by these molecular typing strategies. The 29 strains of P. citronellolis were separated into 12 distinguishable genotypic groups by Rep-PCR and into seven genomic patterns by PCR-based ribotyping. The genetic diversity of the strains was related to the different geoclimatic isolation sites, type of oily sludge, and age of contamination of the sites. These results indicate that a combination of Rep-PCR fingerprinting and PCR-based ribotyping can be used as a high-resolution genomic fingerprinting method for elucidating intraspecies diversity among strains of P. citronellolis.