High Genotypic Similarity Research Articles

Antimicrobial Resistance and Molecular Characterization of Staphylococcus aureus Recovered from Cows with Clinical Mastitis in Dairy Herds from Southeastern Brazil.

Staphylococcus aureus is a contagious pathogen frequently associated with bovine mastitis in Brazil. Molecular characterization of Staph. aureus isolated from affected mammary quarters of cows with clinical mastitis (CM) can provide data on epidemiological behavior of this pathogen and antimicrobial susceptibility (AMS) assessment at the genotypic level. This study genotypically characterized Staph. aureus isolates recovered from cows with CM and determined the association of genotypes and AMS. A total of 84 Staph. aureus strains identified from affected mammary quarters of cows with CM in 13 dairy herds from Southeastern Brazil were submitted for susceptibility testing to 10 antimicrobials using the technique of minimal inhibitory concentration. The same isolates were also genotyped using the spa-typing methodology. Results showed a high genotypic similarity between the Staph. aureus isolates within and between herds, which were categorized as resistant to most antimicrobials, especially to β-lactam antibiotics. In addition, differences in AMS were observed among genotypic clusters, which may affect the efficacy of antimicrobials used to treat CM in different dairy herds.

Investigating extradomiciliary transmission of tuberculosis: An exploratory approach using social network patterns of TB cases and controls and the genotyping of Mycobacterium tuberculosis

Extradomiciliary contacts have been overlooked in the study of TB transmission due to difficulties in identifying actual contacts in large populations. Complex network analysis provides a framework to model the structure of contacts, specially extradomiciliary ones. We conducted a study of incident sputum-positive TB cases and healthy controls occurring in a moderate TB burden city. Cases and controls were interviewed to obtain data regarding the usual locations of residence, work, study, and leisure. Mycobacterium tuberculosis isolated from sputum was genotyped. The collected data were used to build networks based on a framework of putative social interactions indicating possible TB transmission. A user-friendly open source environment (GraphTube) was setup to extract information from the collected data. Networks based on the likelihood of patient-patient, patient-healthy, and healthy-healthy contacts were setup, depending on a constraint of geographical distance of places attended by the volunteers. Using a threshold for the geographical distance of 300 m, the differences between TB cases and controls are revealed. Several clusters formed by social network nodes with high genotypic similarity were characterized. The developed framework provided consistent results and can be used to support the targeted search of potentially infected individuals and to help to understand the TB transmission.

Diverse cellular colonizing endophytic bacteria in field shoots and in vitro cultured papaya with physiological and functional implications.

The study was envisaged to assess the extent of normally uncultivable endophytic bacteria in field papaya plants and in vitro established cultures adopting cultivation vs molecular analysis and microscopy. Surface-sterilized axillary shoot-buds of papaya 'Arka Surya' revealed high bacterial diversity as per 16S rRNA metagene amplicon sequencing (6 phyla, 10 classes, 21 families) with an abundance of Pseudomonas (Gammaproteobacteria), which also formed a common contaminant for in vitro cultured field explants. Molecular analysis of seedling shoot-tip-derived healthy proliferating cultures of three genotypes ('Arka Surya', 'Arka Prabhath', 'Red Lady') with regular monthly subculturing also displayed high bacterial diversity (11-16 phyla, >25 classes, >50 families, >200 genera) about 12-18 months after initial establishment. 'Arka Surya' and 'Red Lady' cultures bore predominantly Actinobacteria (75-78%) while 'Arka Prabhath' showed largely Alphaproteobacteria corroborating the slowly activated Methylobacterium sp. Bright-field direct microscopy on tissue sections and tissue homogenate and epi-fluorescence microscopy employing bacterial DNA probe SYTO-9 revealed abundant intracellular bacteria embracing the next-generation sequencing elucidated high taxonomic diversity. Phylogenetic investigation of communities by reconstruction of unobserved states- PICRUSt- functional annotation suggested significant operational roles for the bacterial-biome. Metabolism, environmental information processing, and genetic information processing constituted major Kyoto Encyclopedia of Genes and Genomes KEGG attributes. Papaya stocks occasionally displayed bacterial growth on culture medium arising from the activation of originally uncultivable organisms to cultivation. The organisms included Bacillus (35%), Methylobacterium (15%), Pseudomonas (10%) and seven other genera (40%). This study reveals a hidden world of diverse and abundant conventionally uncultivable cellular-colonizing endophytic bacteria in field shoots and micropropagating papaya stocks with high genotypic similarity and silent participation in various plant processes/pathways.

Yeast-like fungi and yeasts in withered grape carposphere: Characterization of Aureobasidium pullulans population and species diversity

Yeast-like fungi and yeasts residing on carposphere of withered grapes for Italian passito wine production have been scarcely investigated. In the present study, isolates from single berries, both sound and damaged, of Nosiola, Corvina and Garganega varieties were analyzed at the end of the withering process. Great variation of cell concentration among single berries was observed. In sound berries, yeast-like fungi were significantly more frequent than yeasts. Species identification of isolates was carried out by BLAST comparative analysis on gene databases and phylogenetic approach. All yeast-like fungi isolates belonged to Aureobasidium pullulans. They displayed different culture and physiological characteristics and inhibitory capacity against phytopathogenic fungi. Moreover, PCR profile analysis revealed high genotypic similarity among these strains. A total of 35 species were recognized among yeast isolates. Ascomycetes prevailed over basidiomycetes. To the best of our knowledge, Naganishia onofrii and Rhodosporidiobolus odoratus were identified for the first time among yeasts isolated from grapes, must or wine. Hanseniaspora uvarum and Starmerella bacillaris were the most frequent species. Most species were found only in one grape variety (nine in Nosiola, 10 in Corvina and five in Garganega). The sanitary state of withered grapes could have an important impact on the structure of these epiphytic populations.

Pulse-Field Gel Electrophoresis (PFGE) of Salmonella Serovar Infantis Isolates From Poultry

Background: Salmonella is one of the most important zoonotic agents known to infect humans and a wide range of animals, including poultry. Salmonella Infantis has been one of the 15 most frequently isolated serovars throughout the world. Despite its clinical importance, little is known about the molecular characteristics of S. Infantis strains from Iran. OBJECTIVES: The purpose of this study was mainly to type a number of S. Infantis isolates obtained from Iranian poultry flocks in the last decade by pulse-field gel electrophoresis (PFGE). METHODS: Forty five Salmonella Infantis isolates, mostly from poultry origin, were subjected to PFGE according to protocol of the CDC PulseNet. RESULTS: PFGE revealed 27 pulsotypes and eight clusters among 45 isolates based on the number of observed bands among the pulsotypes. The distribution of 45 isolates among the 27 pulsotypes was variable and included from one to nine isolates. One pulsotype included nine (20%) isolates. The genotypic similarity among 45 isolates was more than 90%. CONCLUSIONS: This study showed the value of PFGE in determining the genotypic similarity among S. Infantis isolates. The high genotypic similarity shown by PFGE among the S. Infantis isolates of this study suggested that the majority of S. Infantis isolates studied may have descended from a common ancestor that has differed little and is responsible for the contamination of poultry flocks and possibly humans as well

Pathogenic potential and genotypic diversity of Campylobacter jejuni: a neglected food-borne pathogen in Brazil.

Using PFGE and flaA-SVR sequencing, the 111 C. jejuni strains studied were grouped into three and two clusters, respectively, and some strains of different origin presented a similarity of ≥80 %. In total, 35 flaA-SVR alleles were detected. Alleles gt45, gt49 and gt57 were the most prevalent, in contrast with those frequently described in the PubMLST database. All 111 C. jejuni strains contained the genes flaA, flhA, cadF, docA, cdtA, cdtB, cdtC, iamA, ciaB, sodB, dnaJ, pldA, racR and csrA. The wlaN gene was detected in 11 strains (9.9 %), and the virB11 in just one strain (0.9 %). In conclusion, the pathogenic potential of the C. jejuni strains studied was highlighted by the high frequency of the majority of the virulence genes searched. The flaA-SVR sequencing and PFGE results showed that some of the strains studied presented a high genotypic similarity, suggesting potential for transmission between animal sources and humans in this country. Altogether, the results characterize further C. jejuni isolates from Brazil, an important producer and exporter of chicken meat.

Genotypic diversity and pathogenic potential of Yersinia enterocolitica biotype 2 strains isolated in Brazil.

To investigate the pathogenic potential and genotypic diversity of Yersinia enterocolitica biotype 2 strains isolated in Brazil and to compare these strains with other Y. enterocolitica biotypes using ERIC-PCR and PFGE. Forty strains of Y. enterocolitica biotype 2 (B2) isolated from humans (5), the environment (34) and animal (1), in Brazil over 19 years were studied. In addition to these isolates, we also analysed 26 Y. enterocolitica strains belonging to the biotypes 1A, 1B, and 3-5. All of the B2 strains contained the genes inv, ail, ystA, hreP, tccC and myfA. The genes fepD and fes were detected in 39 (97·5%) strains, virF was found in three (7·5%) strains, and ystB and fepA were not detected in any strains. The B2 strains showed genotypic similarities of more than 84·8% by ERIC-PCR and of more than 69·0% by PFGE. The pathogenic potential of the B2 strains examined in this study was highlighted by the occurrence of the majority of the virulence markers searched. The results of the ERIC-PCR and PFGE showed that the B2 strains evaluated in this study had a high genotypic similarity, suggesting that these strains differed little over the 19 year study period and that the environment was a possible source of contamination of humans and animals in Brazil. Furthermore, the ERIC-PCR technique grouped the strains belonging to Y. enterocolitica biotypes 1A, 1B, 2, 3, 4 and 5 according to their pathogenicity. This study provided new information about the pathogenic potential and genotypic similarity of Y. enterocolitica B2 isolated from diverse sources in Brazil. Furthermore, ERIC-PCR showed to be a valuable tool for grouping Y. enterocolitica of different biotypes according their pathogenicity.

High Genotypic and Phenotypic Similarity Among Shiga Toxin–ProducingEscherichia coliO111 Environmental and Outbreak Strains

Escherichia coli serogroup O111 is among the six most commonly reported non-O157:H7 Shiga toxin-producing E. coli (STEC), which are emerging as important foodborne pathogens. We have assembled a collection of environmental and clinical strains of E. coli O111 from diverse sources and investigated various genotypic and phenotypic characteristics of these strains to gain a better understanding of the epidemiology and biology of this serogroup. Sixty-three percent of the strains (24/38) were of H-type 8, which dominated the environmental- and outbreak-strains group, whereas the sporadic-case strains were more heterogeneous in H-type. All of the environmental and outbreak strains harbored the Shiga toxin 1 gene (stx1), eae, and ehx, and a subset of these also carried the Shiga toxin 2 gene (stx2). Only 9 of 16 sporadic-case strains produced stx1 and/or stx2, and these were mostly of H-type 8 and 10. Pulsed-field gel electrophoresis analysis revealed a cluster of environmental, outbreak, and sporadic illness strains with high phylogenetic similarity. Strains in this pulsogroup were all of the H8 type and STEC pathotype, and carried eae and ehx. Smaller clusters of highly similar STEC O111 strains included outbreak and sporadic illness strains isolated during different time periods or from different geographical locations. A distinct aggregative behavior was observed in the cultures of all environmental and outbreak STEC O111 strains, but not in those of sporadic-case strains. Among environmental and outbreaks strains, aggregation was positively correlated with production of curli fimbriae and RpoS function, and negatively with cellulose synthesis, while the nonaggregative behavior of sporadic-case strains correlated (positively) only with cellulose production. Our results indicate that STEC O111 strains sharing high genotypic similarity and important phenotypic traits with STEC O111 outbreak strains are present in the agricultural environment and may contribute to the burden of foodborne disease.

A new intertidal Brachionus and intrageneric phylogenetic relationships among Brachionus as revealed by allometry and CO1-ITS1 gene analysis

Abstract Background The rotifer genus Brachionus is distributed worldwide along a gradient from freshwater to seawater habitats. This genus is recognized as a suitable organism for testing environmental and evolutionary hypotheses. Here we present the description of a new species and the phylogenetic relationship within the genus Brachionus as revealed by morphometric allometry of two representatives and CO1-ITS1 gene analysis of five representatives belonging to Brachionus distributed in the fresh and seawaters of Korea, Japan, and China. Results Similarities of populations were studied using nuclear rDNA ITS1 sequences from Brachionus spp. collected from different geographical areas of Far East Asia. The phylogeographic analysis of nuclear DNA ITS1 and mitochondrial CO1 sequences showed that Brachionus from South Korea formed five distinct clades according to their geographic origin. Interspecific differences suggest that Brachionus species established on conventional morphological characters also forms five separate clades. When ITS1-CO1 of the Asian specimens was compared with representatives of Brachionus worldwide, a high genotypic similarity was found. However, they slightly differed between localities. Conclusions This study sets a first step for an integrative morphological and molecular characterization of the diversity contained within the ecologically and economically important rotifer genus Brachionus.

Evaluation of Two Approaches for Assessing the Genetic Similarity of Virioplankton Populations as Defined by Genome Size

Viral production estimates show that virioplankton communities turn over rapidly in aquatic ecosystems. Thus, it is likely that the genetic identity of viral populations comprising the virioplankton also change over temporal and spatial scales, reflecting shifts in viral-host interactions. However, there are few approaches that can provide data on the genotypic identity of viral populations at low cost and with the sample throughput necessary to assess dynamic changes in the virioplankton. This study examined two of these approaches-T4-like major capsid protein (g23) gene polymorphism and randomly amplified polymorphic DNA-PCR (RAPD-PCR) fingerprinting-to ask how well each technique could track differences in virioplankton populations over time and geographic location. Seasonal changes in overall virioplankton composition were apparent from pulsed-field gel electrophoresis (PFGE) analysis. T4-like phages containing similar g23 proteins were found within both small- and large-genome populations, including populations from different geographic locations and times. The surprising occurrence of T4-like g23 within small genomic groups (23 to 64 kb) indicated that the genome size range of T4-like phages may be broader than previously believed. In contrast, RAPD-PCR fingerprinting detected high genotypic similarity within PFGE bands from the same location, time, and genome size class without the requirement for DNA sequencing. Unlike g23 polymorphism, RAPD-PCR fingerprints showed a greater temporal than geographic variation. Thus, while polymorphism in a viral signature gene, such as g23, can be a powerful tool for inferring evolutionary relationships, the degree to which this approach can capture fine-scale variability within virioplankton populations is less clear.

Emerging multi-pathogen disease caused by Didymella bryoniae and pathogenic bacteria on Styrian oil pumpkin

The Styrian oil pumpkin, Cucurbita pepo L. subsp. pepo var. styriaca Greb. is a crop of cultural, commercial, and medical importance. In the last decade, yield losses of pumpkins increased dramatically. The ascomycetous fungus Didymella bryoniae (Fuckel) Rehm was identified as main causal agent provoking gummy stem blight as well as black rot of pumpkins. We observed a remarkable phenotypic diversity of the fungal pathogen, which contrasted with a high genotypic similarity. Evidence of pathogenictiy of D. bryoniae on Styrian oil pumpkin was demonstrated in a newly developed greenhouse assay. Isolates representing the five observed phenotypic groups fulfilled the Koch’s postulates. In the field, the fungal disease was often associated with bacterial colonization by Pectobacterium carotovorum, Pseudomonas viridiflava, Pseudomonas syringae and Xanthomonas cucurbitae. The pathogenic behaviour of bacterial isolates on pumpkin was confirmed in the greenhouse assay. The high co-incidence of fungal and bacterial disease suggests mutualistic effects in pathogenesis. With a new assay, we found that bacteria can use the mycelium of D. bryoniae for translocation. We argue that the rapid rise of the multi-pathogen disease of pumpkins results from combined action of versatile pathogenic bacteria and the rapid translocation on a structurally versatile mycelium of the fungal pathogen.

Phylogenetic analysis of Rhodococcus erythropolis based on the variation of ribosomal proteins as observed by matrix-assisted laser desorption ionization-mass spectrometry without using genome information

Rhodococcus erythropolis strains characterized as antibiotic producers can be classified into three groups according to their antibiotic spectrum and growth compatibility. Due to their high genotypic similarity, the taxonomic relationship of these strains has not been elucidated. In this study, ribosomal protein profiling using matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS) was employed to classify twenty-one strains of R. erythropolis (15 antibiotic producers and 6 non-antibiotic producers). In the first step in this method, a total of 30 intense peaks observed for purified ribosomal subunit proteins of the type strain (R. erythropolis JCM 3201(T)) were selected as the reference peaks. The mass spectra observed for the cell lysates of each sample strain were then checked as to whether peaks were observed at the same masses of the reference peaks. The results of peak matching were processed by cluster analysis, generating a dendrogram. Four major clusters of the R. erythropolis strains corresponded to three antibiotic groups and the non-antibiotic group. Furthermore, the topology of the dendrogram was highly comparable with the phylogenetic tree based on DNA gyrase subunit B gene (gyrB) sequencing. These results indicate that our proposed ribosomal protein profiling method using MALDI-MS is a potentially reliable and sufficiently high-throughput technique for the taxonomic analysis of closely related bacterial strains without using DNA sequence information.

Equine protease inhibitor system as a marker for the diagnosis of chronic obstructive pulmonary disease (COPD)

The protease inhibitor system (PI) was investigated to ascertain if it can be used as a marker of chronic obstructive pulmonary disease (COPD) in thoroughbred horses. Serum samples were taken from healthy thoroughbreds (n = 13) and those diagnosed as having COPD (n = 24) or inflammatory airway disease (IAD, n = 38) as well as from 3,600 undiagnosed thoroughbred horses. PI allelic and genotypic frequencies were estimated using protein electrophoresis on starch and polyacrylamide gels. The four groups of horses showed high genotypic similarity and none of the observed alleles or genotypes of the equine PI system were found to be associated with COPD.

Isolates of Pneumocystis jirovecii from Harare show high genotypic similarity to isolates from London at the superoxide dismutase locus

Pneumocystis jirovecii is the cause of Pneumocystis pneumonia (PCP) in humans. Isolates of P. jirovecii obtained from patients in Harare, Zimbabwe were genotyped at the superoxide dismutase locus. High genotypic similarity to isolates of P. jirovecii obtained from patients in London, UK was observed. These data provide additional support for the hypothesis that P. jirovecii is genetically indistinguishable in isolates from geographically diverse locations.

Genetic structure of Cenococcum geophilum populations in primary successional volcanic deserts on Mount Fuji as revealed by microsatellite markers

Polymorphic simple sequence repeat (SSR) markers were used to investigate the genetic structure in a Cenococcum geophilum population associated with Salix reinii in an early successional volcanic desert at Gotenba, on the south-eastern slope of Mount Fuji in Japan, and in three other populations associated with the same host at more developed sites on the mountain, one at Fujinomiya and two at Subashiri. The genotype richness of C. geophilum tended to be higher in more developed vegetation patches as well as in more developed sites, suggesting that genotype richness increased with advanced succession because new genotypes might have been introduced into these sites over time. High genotypic similarity was observed between the Gotenba and Fujinomiya populations but not between the Gotenba and Subashiri populations, suggesting that C. geophilum genotypes in Gotenba were introduced from the direction of Fujinomiya. Genotypes in the Gotenba population were clearly distinguishable into two groups. The absence of any intermediate genotype suggests the absence of frequent recombination in this C. geophilum population associated with early successional vegetation.