Genetic and phenotype analysis of Borrelia valaisiana sp.nov. (Borrelia genomic groups VS116 and M19)

To clarify the taxonomic status of two recently described Borrelia genomic groups, groups VS116 and M19, three group VS116 strains and eight group M19 strains isolated from Ixodes ricinus ticks in Switzerland, The Netherlands, and the United Kingdom were characterized. PCR-restriction fragment length polymorphism (RFLP) analysis of the 5S-23S intergenic spacer amplicon, rRNA gene restriction analysis, 16S rRNA gene sequence analysis, randomly amplified polymorphic DNA (RAPD) fingerprinting, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and immunoblotting with monoclonal antibodies were used for genetic and phenotypic analysis. The PCR-RFLP and RAPD patterns of three group VS116 strains and eight group M19 strains were identical but differed from those of Borrelia burgdorferi sensu stricto, Borrelia garinii, Borrelia afzelii, and Borrelia japonica. DNAs from all group VS116 and M19 strains yielded three fragments (6.9, 3.2, and 1.4 kb) and four fragments (2.1, 1.2, 0.8, and 0.6 kb) after digestion with EcoRV and HindIII, respectively, hybridizing with an Escherichia coli 16S + 23S cDNA probe. The SDS-PAGE protein profiles of group VS116 and M19 strains were heterogeneous. Phylogenetic analysis of the partial 16S rRNA gene sequences showed that group VS116 and M19 spirochetes were members of a Borrelia species distinct from previously characterized members of the genus Borrelia. Based on our present study and data from previous DNA-DNA hybridizations, a new Borrelia species, Borrelia valaisiana sp.nov., in the B. burgdorferi complex, is proposed. Strain VS116 is the type strain of this new specie

MLVA-16 typing of 295 marine mammal <i>Brucella</i> isolates from different animal and geographic origins identifies 7 major groups within <i>Brucella ceti</i> and <i>Brucella pinnipedialis</i>

BackgroundSince 1994, Brucella strains have been isolated from a wide range of marine mammals. They are currently recognized as two new Brucella species, B. pinnipedialis for the pinniped isolates and B. ceti for the cetacean isolates in agreement with host preference and specific phenotypic and molecular markers. In order to investigate the genetic relationships within the marine mammal Brucella isolates and with reference to terrestrial mammal Brucella isolates, we applied in this study the Multiple Loci VNTR (Variable Number of Tandem Repeats) Analysis (MLVA) approach. A previously published assay comprising 16 loci (MLVA-16) that has been shown to be highly relevant and efficient for typing and clustering Brucella strains from animal and human origin was used.Results294 marine mammal Brucella strains collected in European waters from 173 animals and a human isolate from New Zealand presumably from marine origin were investigated by MLVA-16. Marine mammal Brucella isolates were shown to be different from the recognized terrestrial mammal Brucella species and biovars and corresponded to 3 major related groups, one specific of the B. ceti strains, one of the B. pinnipedialis strains and the last composed of the human isolate. In the B. ceti group, 3 subclusters were identified, distinguishing a cluster of dolphin, minke whale and porpoise isolates and two clusters mostly composed of dolphin isolates. These results were in accordance with published analyses using other phenotypic or molecular approaches, or different panels of VNTR loci. The B. pinnipedialis group could be similarly subdivided in 3 subclusters, one composed exclusively of isolates from hooded seals (Cystophora cristata) and the two others comprising other seal species isolates.ConclusionThe clustering analysis of a large collection of marine mammal Brucella isolates from European waters significantly strengthens the current view of the population structure of these two species, and their relative position with respect to the rest of the Brucella genus. MLVA-16 is confirmed as being a rapid, highly discriminatory and reproducible method to classify Brucella strains including the marine mammal isolates. The Brucella2009 MLVA-16 genotyping database available at http://mlva.u-psud.fr is providing a detailed coverage of all 9 currently recognized Brucella species

Molecular typing of mycobacterium tuberculosis isolates circulating in Jiangsu Province, China

<p>Abstract</p> <p>Background</p> <p>Globally, China is the second place with high burden of tuberculosis (TB). To explore the characteristics of the pathogens of <it>Mycobacterium tuberculosis </it>(MTB) circulating in this area is helpful for understanding and controlling the spread of the strains. Recent developments in molecular biology have allowed prompt identification and tracking specific strains of MTB spreading through the population.</p> <p>Methods</p> <p>Spacer-oligonucleotide typing (spoligotyping) and mycobacterial interspersed repetitive units variable number tandem repeat (MIRU-VNTR) were performed in combination to yield specific genetic profiles of 260 MTB strains isolated from 30 counties of Jiangsu province in China between June and July 2010. The spoligotyping results were in comparison to the world Spoligotyping Database of Institute Pasteur de Guadeloupe (SpolDB4). Drug susceptibility test (DST) was performed on all strains by proportion method on Lowenstein-Jensen (LJ) culture media.</p> <p>Results</p> <p>Based on the spoligotyping method, 246 strains displayed known patterns and 14 were absent in the database. Predominant spoligotypes belonged to the Beijing family (80.4%). By using the 24-loci VNTR typing scheme, 224 different patterns were identified, including 20 clusters and 204 unique patterns. The largest clade comprised 195 strains belonging to the Beijing family. The combination of spoligotyping and 24-loci MIRU-VNTR demonstrated maximal discriminatory power. Furthermore, we observed a significant association between Beijing family strains and drug-resistant phenotypes. The Beijing family strains presented increased risks for developing multi-drug resistant TB, with the OR (95% CI) of 11.07(1.45-84.50).</p> <p>Conclusions</p> <p>The present study demonstrated that Beijing family isolates were the most prevalent strains circulating in Jiangsu province of China. The utility of spoligotyping in combination with 24-loci MIRU-VNTR might be a useful tool for epidemiological analysis of MTB transmission.</p

Diversity of Mycobacterium tuberculosis genotypes circulating in Ndola, Zambia

<p>Abstract</p> <p>Background</p> <p>Tuberculosis (TB) is one of the major public health problems in Zambia. However, information about lineages of <it>M. tuberculosis </it>complex (MTBC) isolates useful for epidemiology investigations is unknown. In this study, we investigated the diversity of MTBC isolates from Ndola, a typical Zambian urbanized city with a documented high HIV prevalence.</p> <p>Methods</p> <p>This was part of a prospective cohort study in subjects with sputum smear-positive pulmonary TB. Spoligotyping was used to genotype the MTBC isolates and establish the circulating lineages. The 15-locus Mycobacterial Interspersed Repetitive Units - Variable Number Tandem Repeats (MIRU-VNTR) typing was used to study recent transmission.</p> <p>Results</p> <p>A total of 98 different spoligotypes were identified among 273 MTBC isolates. The majority (64.8%) of the isolates belonged to 9 known families, while 96 (35.2%) of the isolates were orphans. While LAM (41.8%) was the largest spoligotype family observed, most of the isolates (87.7%) belonging to the SAF1 family, with a significant portion coming from the T (13.6%), and X (5.9%) families. A few isolates (3.6%) belonged to the CAS, EAI, H, S, X1-LAM9 or U families. MIRU-VNTR typing was highly discriminatory (h = 0.988) among the 156 isolates tested in our sample, and increased the discrimination among 82 SAF1 isolates from 6 to 46 distinct patterns. In addition, 3.2% (5/156) of cases with available MIRU-VNTR results harbored more than one MTBC strain.</p> <p>Conclusions</p> <p>Our findings show a limited diversity of MTBC in Ndola with a high clustering rate (37.7%), which indicates that recent transmission plays an appreciable role in the dynamics of TB disease in this setting. This conclusion emphasizes the importance of early diagnosis and timely treatment. The results also confirm that MIRU-VNTR typing is suitable for studying the molecular epidemiology of TB in Ndola.</p

Genomes of the Most Dangerous Epidemic Bacteria Have a Virulence Repertoire Characterized by Fewer Genes but More Toxin-Antitoxin Modules

We conducted a comparative genomic study based on a neutral approach to identify genome specificities associated with the virulence capacity of pathogenic bacteria. We also determined whether virulence is dictated by rules, or if it is the result of individual evolutionary histories. We systematically compared the genomes of the 12 most dangerous pandemic bacteria for humans ("bad bugs") to their closest non-epidemic related species ("controls").We found several significantly different features in the "bad bugs", one of which was a smaller genome that likely resulted from a degraded recombination and repair system. The 10 Cluster of Orthologous Group (COG) functional categories revealed a significantly smaller number of genes in the "bad bugs", which lacked mostly transcription, signal transduction mechanisms, cell motility, energy production and conversion, and metabolic and regulatory functions. A few genes were identified as virulence factors, including secretion system proteins. Five "bad bugs" showed a greater number of poly (A) tails compared to the controls, whereas an elevated number of poly (A) tails was found to be strongly correlated to a low GC% content. The "bad bugs" had fewer tandem repeat sequences compared to controls. Moreover, the results obtained from a principal component analysis (PCA) showed that the "bad bugs" had surprisingly more toxin-antitoxin modules than did the controls.We conclude that pathogenic capacity is not the result of "virulence factors" but is the outcome of a virulent gene repertoire resulting from reduced genome repertoires. Toxin-antitoxin systems could participate in the virulence repertoire, but they may have developed independently of selfish evolution

Fine-Scale Phylogeographic Structure of Borrelia lusitaniae Revealed by Multilocus Sequence Typing

Borrelia lusitaniae is an Old World species of the Lyme borreliosis (LB) group of tick-borne spirochetes and prevails mainly in countries around the Mediterranean Basin. Lizards of the family Lacertidae have been identified as reservoir hosts of B. lusitaniae. These reptiles are highly structured geographically, indicating limited migration. In order to examine whether host geographic structure shapes the evolution and epidemiology of B. lusitaniae, we analyzed the phylogeographic population structure of this tick-borne bacterium using a recently developed multilocus sequence typing (MLST) scheme based on chromosomal housekeeping genes. A total of 2,099 questing nymphal and adult Ixodes ricinus ticks were collected in two climatically different regions of Portugal, being ∼130 km apart. All ticks were screened for spirochetes by direct PCR. Attempts to isolate strains yielded 16 cultures of B. lusitaniae in total. Uncontaminated cultures as well as infected ticks were included in this study. The results using MLST show that the regional B. lusitaniae populations constitute genetically distinct populations. In contrast, no clear phylogeographic signals were detected in sequences of the commonly used molecular markers ospA and ospC. The pronounced population structure of B. lusitaniae over a short geographic distance as captured by MLST of the housekeeping genes suggests that the migration rates of B. lusitaniae are rather low, most likely because the distribution of mediterranean lizard populations is highly parapatric. The study underlines the importance of vertebrate hosts in the geographic spread of tick-borne microparasites

The Secret Life of the Anthrax Agent Bacillus anthracis: Bacteriophage-Mediated Ecological Adaptations

Ecological and genetic factors that govern the occurrence and persistence of anthrax reservoirs in the environment are obscure. A central tenet, based on limited and often conflicting studies, has long held that growing or vegetative forms of Bacillus anthracis survive poorly outside the mammalian host and must sporulate to survive in the environment. Here, we present evidence of a more dynamic lifecycle, whereby interactions with bacterial viruses, or bacteriophages, elicit phenotypic alterations in B. anthracis and the emergence of infected derivatives, or lysogens, with dramatically altered survival capabilities. Using both laboratory and environmental B. anthracis strains, we show that lysogeny can block or promote sporulation depending on the phage, induce exopolysaccharide expression and biofilm formation, and enable the long-term colonization of both an artificial soil environment and the intestinal tract of the invertebrate redworm, Eisenia fetida. All of the B. anthracis lysogens existed in a pseudolysogenic-like state in both the soil and worm gut, shedding phages that could in turn infect non-lysogenic B. anthracis recipients and confer survival phenotypes in those environments. Finally, the mechanism behind several phenotypic changes was found to require phage-encoded bacterial sigma factors and the expression of at least one host-encoded protein predicted to be involved in the colonization of invertebrate intestines. The results here demonstrate that during its environmental phase, bacteriophages provide B. anthracis with alternatives to sporulation that involve the activation of soil-survival and endosymbiotic capabilities

Brucellosis as an Emerging Threat in Developing Economies:Lessons from Nigeria

Nigeria is the most populous country in Africa, has a large proportion of the world's poor livestock keepers, and is a hotspot for neglected zoonoses. A review of the 127 accessible publications on brucellosis in Nigeria reveals only scant and fragmented evidence on its spatial and temporal distribution in different epidemiological contexts. The few bacteriological studies conducted demonstrate the existence of Brucella abortus in cattle and sheep, but evidence for B. melitensis in small ruminants is dated and unclear. The bulk of the evidence consists of seroprevalence studies, but test standardization and validation are not always adequately described, and misinterpretations exist with regard to sensitivity and/or specificity and ability to identify the infecting Brucella species. Despite this, early studies suggest that although brucellosis was endemic in extensive nomadic systems, seroprevalence was low, and brucellosis was not perceived as a real burden; recent studies, however, may reflect a changing trend. Concerning human brucellosis, no studies have identified the Brucella species and most reports provide only serological evidence of contact with Brucella in the classical risk groups; some suggest brucellosis misdiagnoses as malaria or other febrile conditions. The investigation of a severe outbreak that occurred in the late 1970s describes the emergence of animal and human disease caused by the settling of previously nomadic populations during the Sahelian drought. There appears to be an increasing risk of re-emergence of brucellosis in sub-Saharan Africa, as a result of the co-existence of pastoralist movements and the increase of intensive management resulting from growing urbanization and food demand. Highly contagious zoonoses like brucellosis pose a threat with far-reaching social and political consequences