Speciation patterns of Aedes mosquitoes in the Scutellaris Group: a mitochondrial perspective.

The Scutellaris Group of Aedes comprises 47 mosquito species, including Aedes albopictus. While Ae. albopictus is widely distributed, the other species are mostly found in the Asia-Pacific region. Evolutionary history researches of Aedes species within the Scutellaris Group have mainly focused on Ae. albopictus, a species that raises significant public health concerns, neglecting the other species. In this study, we aimed to assess genetic diversity and estimate speciation times of several species within the Scutellaris Group. Mosquitoes were therefore collected from various Asia-Pacific countries. Their mitochondrial cytochrome c oxidase subunit 1 (cox1) and subunit 3 (cox3) sequences were analyzed alongside those of other Scutellaris Group species available in the GenBank database. To estimate the divergence time, we analyzed 1849 cox1 gene sequences from 21 species, using three species (Aedes aegypti, Aedes notoscriptus and Aedes vigilax) as outgroups. We found that most of the speciation dates occurred during the Paleogene and the Neogene periods. A separation between the Scutellaris Subgroup and the Albopictus Subgroup occurred approximately 64-61 million years ago (MYA). We also identified a split between species found in Asia/Micronesia and those collected in Melanesia/Polynesia approximately 36-35 MYA. Our findings suggest that the speciation of Aedes species within the Scutellaris Group may be driven by diversity in mammalian hosts, climate and environmental changes, and geological dynamics rather than human migration.

Evaluating vector competence for Yellow fever in the Caribbean.

The mosquito-borne disease, Yellow fever (YF), has been largely controlled via mass delivery of an effective vaccine and mosquito control interventions. However, there are warning signs that YF is re-emerging in both Sub-Saharan Africa and South America. Imported from Africa in slave ships, YF was responsible for devastating outbreaks in the Caribbean. In Martinique, the last YF outbreak was reported in 1908 and the mosquito Aedes aegypti was incriminated as the main vector. We evaluated the vector competence of fifteen Ae. aegypti populations for five YFV genotypes (Bolivia, Ghana, Nigeria, Sudan, and Uganda). Here we show that mosquito populations from the Caribbean and the Americas were able to transmit the five YFV genotypes, with YFV strains for Uganda and Bolivia having higher transmission success. We also observed that Ae. aegypti populations from Martinique were more susceptible to YFV infection than other populations from neighboring Caribbean islands, as well as North and South America. Our vector competence data suggest that the threat of re-emergence of YF in Martinique and the subsequent spread to Caribbean nations and beyond is plausible.

Hospital and urban wastewaters shape the matrix and active resistome of environmental biofilms.

Understanding the dynamics of antibiotic resistance gene (ARG) transfer and dissemination in natural environments remains challenging. Biofilms play a crucial role in bacterial survival and antimicrobial resistance (AMR) dissemination in natural environments, particularly in aquatic systems. This study focused on hospital and urban wastewater (WW) biofilms to investigate the potential for ARG dissemination through mobile genetic elements (MGEs). The analysis included assessing the biofilm extracellular polymeric substances (EPS), microbiota composition as well as metatranscriptomic profiling of the resistome and mobilome. We produced both in vitro and in situ biofilms and performed phenotypic and genomic analyses. In the in vitro setup, untreated urban and hospital WW was used to establish biofilm reactors, with ciprofloxacin added as a selective agent at minimal selective concentration. In the in situ setup, biofilms were developed directly in hospital and urban WW pipes. We first showed that a) the composition of EPS differed depending on the growth environment (in situ and in vitro) and the sampling origin (hospital vs urban WW) and that b) ciprofloxacin impacted the composition of the EPS. The metatranscriptomic approach showed that a) expression of several ARGs and MGEs increased upon adding ciprofloxacin for biofilms from hospital WW only and b) that the abundance and type of plasmids that carried individual or multiple ARGs varied depending on the WW origins of the biofilms. When the same plasmids were present in both, urban and hospital WW biofilms, they carried different ARGs.  We showed that hospital and urban wastewaters shaped the structure and active resistome of environmental biofilms, and we confirmed that hospital WW is an important hot spot for the dissemination and selection of antimicrobial resistance. Our study provides a comprehensive assessment of WW biofilms as crucial hotspots for ARG transfer. Hospital WW biofilms exhibited distinct characteristics, including higher eDNA abundance and expression levels of ARGs and MGEs, highlighting their role in antimicrobial resistance dissemination. These findings emphasize the importance of understanding the structural, ecological, functional, and genetic organization of biofilms in anthropized environments and their contribution to antibiotic resistance dynamics.

Climate change and vector-borne diseases: a multi-omics approach of temperature-induced changes in the mosquito.

BACKGROUND: Climate change and globalization contribute to the expansion of mosquito vectors and their associated pathogens. Long spared, temperate regions have had to deal with the emergence of arboviruses traditionally confined to tropical regions. Chikungunya virus (CHIKV) was reported for the first time in Europe in 2007, causing a localized outbreak in Italy, which then recurred repeatedly over the years in other European localities. This raises the question of climate effects, particularly temperature, on the dynamics of vector-borne viruses. The objective of this study is to improve the understanding of the molecular mechanisms set up in the vector in response to temperature. METHODS: We combine three complementary approaches by examining Aedes albopictus mosquito gene expression (transcriptomics), bacterial flora (metagenomics) and CHIKV evolutionary dynamics (genomics) induced by viral infection and temperature changes. RESULTS: We show that temperature alters profoundly mosquito gene expression, bacterial microbiome and viral population diversity. We observe that (i) CHIKV infection upregulated most genes (mainly in immune and stress-related pathways) at 20°C but not at 28°C, (ii) CHIKV infection significantly increased the abundance of Enterobacteriaceae Serratia marcescens at 28°C and (iii) CHIKV evolutionary dynamics were different according to temperature. CONCLUSION: The substantial changes detected in the vectorial system (the vector and its bacterial microbiota, and the arbovirus) lead to temperature-specific adjustments to reach the ultimate goal of arbovirus transmission; at 20°C and 28°C, the Asian tiger mosquito Ae. albopictus was able to transmit CHIKV at the same efficiency. Therefore, CHIKV is likely to continue its expansion in the northern regions and could become a public health problem in more countries than those already affected in Europe.

Zika vector competence data reveals risks of outbreaks: the contribution of the European ZIKAlliance project.

First identified in 1947, Zika virus took roughly 70 years to cause a pandemic unusually associated with virus-induced brain damage in newborns. Zika virus is transmitted by mosquitoes, mainly Aedes aegypti, and secondarily, Aedes albopictus, both colonizing a large strip encompassing tropical and temperate regions. As part of the international project ZIKAlliance initiated in 2016, 50 mosquito populations from six species collected in 12 countries were experimentally infected with different Zika viruses. Here, we show that Ae. aegypti is mainly responsible for Zika virus transmission having the highest susceptibility to viral infections. Other species play a secondary role in transmission while Culex mosquitoes are largely non-susceptible. Zika strain is expected to significantly modulate transmission efficiency with African strains being more likely to cause an outbreak. As the distribution of Ae. aegypti will doubtless expand with climate change and without new marketed vaccines, all the ingredients are in place to relive a new pandemic of Zika.

Impact of temperature on dengue and chikungunya transmission by the mosquito Aedes albopictus.

The mosquito Aedes albopictus is an invasive species first detected in Europe in Albania in 1979, and now established in 28 European countries. Temperature is a limiting factor in mosquito activities and in the transmission of associated arboviruses namely chikungunya (CHIKV) and dengue (DENV). Since 2007, local transmissions of CHIKV and DENV have been reported in mainland Europe, mainly in South Europe. Thus, the critical question is how far north transmission could occur. In this context, the Albanian infestation by Ae. albopictus is of interest because the species is present up to 1200 m of altitude; this allows using altitude as a proxy for latitude. Here we show that Ae. albopictus can transmit CHIKV at 28 °C as well as 20 °C, however, the transmission of DENV is only observed at 28 °C. We conclude that if temperature is the key environmental factor limiting transmission, then transmission of CHIKV, but not DENV is feasible in much of Europe.

Characteristics of Helicobacter pylori strains isolated from Mauritanian patients.

BACKGROUND: Helicobacter pylori (H pylori) is responsible for various diseases including cancer It co-evolved with humans, and human migrations shaped the expansion and the diversity of strains around the world. The risk of developing a disease depends on virulence factors, mainly the cytotoxin-associated gene A protein (CagA). The aim of this study was to determine the cagA status in H pylori strains from Mauritanian patients and to search for a relationship with endoscopic and histologic findings. MATERIAL AND METHODS: H pylori was searched in gastric biopsies taken during endoscopy in patients with gastro-duodenal symptoms. RT-PCR was used for the diagnosis and resistance to clarithromycin. The cagA status was determined with PCR and the EPIYA-cagA polymorphism with sequencing. RESULTS: At all, 76/78 (97.4%) biopsies were positive. The rate of clarithromycin resistance was 4/76 (5.26%) due to the A2143G mutation, with a mixed population in 2 cases. The cagA gene was present in 23/76 (30.26%) biopsies, and the EPIYA motif was ABC in 21 (91.3%). High bacterial load and inflammation were significantly associated with cagA-positive status (P < .01). Phylogenetic analysis of the glmM and hspA genes highlighted a mixture of African and European genes in strains of H pylori isolated from patients of Moor origin. CONCLUSION: We report a high prevalence of H pylori infection in Mauritanian patients, a low rate of clarithromycin resistance (5.26%) and high bacterial load and inflammation associated with cagA-positive status. The phylogenetic analysis highlights the mix of different populations leading to the Moor ethnicity.

Evolution and biological significance of flaviviral elements in the genome of the arboviral vector Aedes albopictus.

Since its genome details are publically available, the mosquito Aedes albopictus has become the central stage of attention for deciphering multiple biological and evolutionary aspects at the root of its success as an invasive species. Its genome of 1,967 Mb harbours an unusual high number of non-retroviral integrated RNA virus sequences (NIRVS). NIRVS are enriched in piRNA clusters and produce piRNAs, suggesting an antiviral effect. Here, we investigated the evolutionary history of NIRVS in geographically distant Ae. albopictus populations by comparing genetic variation as derived by neutral microsatellite loci and seven selected NIRVS. We found that the evolution of NIRVS was far to be neutral with variations both in their distribution and sequence polymorphism among Ae. albopictus populations. The Flaviviral elements AlbFlavi2 and AlbFlavi36 were more deeply investigated in their association with dissemination rates of dengue virus (DENV) and chikungunya virus (CHIKV) in Ae. albopictus at both population and individual levels. Our results show a complex association between NIRVS and DENV/CHIKV opening a new avenue for investigating the functional role of NIRVS as antiviral elements shaping vector competence of mosquitoes to arboviruses.

Manual and expert annotation of the nearly complete genome sequence of Staphylococcus sciuri strain ATCC 29059: A reference for the oxidase-positive staphylococci that supports the atypical phenotypic features of the species group.

Staphylococcus sciuri is considered to be one of the most ancestral species in the natural history of the Staphylococcus genus that consists of 48 validly described species. It belongs to the basal group of oxidase-positive and novobiocin-resistant staphylococci that diverged from macrococci approximately 250 million years ago. Contrary to other groups, the S. sciuri species group has not developed host-specific colonization strategies. Genome analysis of S. sciuri ATCC 29059 provides here the first genetic basis for atypical traits that would support the switch between the free-living style and the infective state in animals and humans. From among the most remarkable features, it was noticed in this extensive study that there were a number of phosphoenolpyruvate:carbohydrate phosphotransferase systems (PTS), almost twice as many as any other staphylococci, and the co-occurrence of mevalonate and non-mevalonate pathways for isoprenoid synthesis. The sequenced strain was devoid of the main virulence factors present in Staphylococcus aureus, although it exhibited numerous heme and iron acquisition systems, as well as crt and aldH genes necessary for gold pigment synthesis. The sensing and signaling networks, exemplified by a large and typical repertoire of two-component regulatory systems and a complete panel of master regulators, such as agr, rex, mgrA, rot, sarA and sarR genes, depict the background in which S. aureus virulence genes were later acquired. An additional sigma factor, a distinct set of electron transducer elements and many gene operons similar to those found in Bacillus spp. would constitute the most visible remnant links with Bacillaceae organisms.

Comparative genomics of Clostridium bolteae and Clostridium clostridioforme reveals species-specific genomic properties and numerous putative antibiotic resistance determinants.

BACKGROUND: Clostridium bolteae and Clostridium clostridioforme, previously included in the complex C. clostridioforme in the group Clostridium XIVa, remain difficult to distinguish by phenotypic methods. These bacteria, prevailing in the human intestinal microbiota, are opportunistic pathogens with various drug susceptibility patterns. In order to better characterize the two species and to obtain information on their antibiotic resistance genes, we analyzed the genomes of six strains of C. bolteae and six strains of C. clostridioforme, isolated from human infection. RESULTS: The genome length of C. bolteae varied from 6159 to 6398 kb, and 5719 to 6059 CDSs were detected. The genomes of C. clostridioforme were smaller, between 5467 and 5927 kb, and contained 5231 to 5916 CDSs. The two species display different metabolic pathways. The genomes of C. bolteae contained lactose operons involving PTS system and complex regulation, which contribute to phenotypic differentiation from C. clostridioforme. The Acetyl-CoA pathway, similar to that of Faecalibacterium prausnitzii, a major butyrate producer in the human gut, was only found in C. clostridioforme. The two species have also developed diverse flagella mobility systems contributing to gut colonization. Their genomes harboured many CDSs involved in resistance to beta-lactams, glycopeptides, macrolides, chloramphenicol, lincosamides, rifampin, linezolid, bacitracin, aminoglycosides and tetracyclines. Overall antimicrobial resistance genes were similar within a species, but strain-specific resistance genes were found. We discovered a new group of genes coding for rifampin resistance in C. bolteae. C. bolteae 90B3 was resistant to phenicols and linezolide in producing a 23S rRNA methyltransferase. C. clostridioforme 90A8 contained the VanB-type Tn1549 operon conferring vancomycin resistance. We also detected numerous genes encoding proteins related to efflux pump systems. CONCLUSION: Genomic comparison of C. bolteae and C. clostridiofrome revealed functional differences in butyrate pathways and in flagellar systems, which play a critical role within human microbiota. Most of the resistance genes detected in both species were previously characterized in other bacterial species. A few of them were related to antibiotics inactive against Clostridium spp. Some were part of mobile genetic elements suggesting that these commensals of the human microbiota act as reservoir of antimicrobial resistances.

Importance of mosquito "quasispecies" in selecting an epidemic arthropod-borne virus.

Most arthropod-borne viruses (arboviruses), perpetuated by alternation between a vertebrate host and an insect vector, are likely to emerge through minor genetic changes enabling the virus to adapt to new hosts. In the past decade, chikungunya virus (CHIKV; Alphavirus, Togaviridae) has emerged on La Réunion Island following the selection of a unique substitution in the CHIKV E1 envelope glycoprotein (E1-A226V) of an East-Central-South African (ECSA) genotype conferring a higher transmission rate by the mosquito Aedes albopictus. Assumed to have occurred independently on at least four separate occasions, this evolutionary convergence was suspected to be responsible for CHIKV worldwide expansion. However, assumptions on CHIKV emergence were mainly based on viral genetic changes and the role of the mosquito population quasispecies remained unexplored. Here we show that the nature of the vector population is pivotal in selecting the epidemic CHIKV. We demonstrate using microsatellites mosquito genotyping that Ae. albopictus populations are genetically differentiated, contributing to explain their differential ability to select the E1-226V mutation. Aedes albopictus, newly introduced in Congo coinciding with the first CHIKV outbreak, was not able to select the substitution E1-A226V nor to preferentially transmit a CHIKV clone harboring the E1-226V as did Ae. albopictus from La Réunion.

Diagnosis of Helicobacter pylori recurrence: relapse or reinfection? Usefulness of molecular tools.

BACKGROUND AND AIMS: Infection due to Helicobacter pylori causes many gastrointestinal diseases including peptic ulcers and gastric carcinoma. Their treatment and prevention depends on the successful eradication of H. pylori. However, even after a well-conducted treatment, H. pylori persists in about 10-30% of patients. Recurrent infections can correspond to relapse or to re-infection and require appropriate medical care. In this study, we explore retrospectively three clinical cases using molecular methods, and propose new guidelines for the diagnosis of recurrence. MATERIAL AND METHODS: Ten colonies of H. pylori were selected from the primary culture of biopsy samples taken from the antrum and fundus for each patient. The genotype of each isolated colony was determined by analyzing the polymorphism of two housekeeping genes, hspA and glmM. The genome-wide composition of H. pylori strains was studied using in house macro-arrays designed. RESULTS: Relapses were demonstrated by the stability of genotypes and the slight genetic variability of strains on macro-arrays. Two patients suffered from relapses, one and three years after H. pylori treatment. For the third patient, both the polymorphism of glmM and hspA genotypes and the diversity of CDSs identified on macro-arrays suggested that several episodes of re-infection occurred, 1-8 years after eradication. CONCLUSION: For the three clinical cases, molecular methods allowed identifying the causes of recurrent infections. We suggest to study genotype to distinguish between relapse and re-infection in order to adapt the treatment and the follow-up of patients to the nature of recurrence.

Impact of human migrations on diversity of Helicobacter pylori in Cambodia and New Caledonia.

BACKGROUND: Helicobacter pylori is a major gastric bacterial pathogen, presumed to have established itself in the human stomach approximately 100,000 years ago. Helicobacter pylori co-evolved with its host, and human migrations shaped the expansion and the diversity of strains around the world. Here, we investigated the population structure and the genomic diversity of H. pylori in New Caledonia and Cambodia, where humans of different origins are living. METHODS: Both multilocus sequence typing (MLST) and macro-array experiments were performed to assess polymorphism of housekeeping genes and to compare differences in gene contents among strains of H. pylori. RESULTS: The macro-array analysis based on variations of the flexible gene pools was consistent with the contribution of ancestral H. pylori populations to modern strains. Most of the CDS variably present encode proteins of unknown function, selfish DNA, and transposases. In New Caledonia-where humans are of several ethnic origins-strains belonged to four different genetic populations, reflecting the diversity of human populations. Melanesians and Polynesians were infected mainly by strains assigned to hspMaori, whereas Caucasians were infected by hspWAfrica, hpEurope, and hpNEAfrica strains. In contrast, strains from Khmer patients belonged to only two subpopulations: hspEAsia and hpEurope. In the two countries, both ancient and recent human migrations may have influenced the diversity of H. pylori. CONCLUSION: Our present results are consistent with the possibility of admixture of strains in multiethnic communities. This increases the global polymorphism of H. pylori without evidence of functional change or impact on fitness and virulence.

A crypt-specific core microbiota resides in the mouse colon.

UNLABELLED: In an attempt to explore the microbial content of functionally critical niches of the mouse gastrointestinal tract, we targeted molecular microbial diagnostics of the crypts that contain the intestinal stem cells, which account for epithelial regeneration. As current evidence indicates, the gut microbiota affects epithelial regeneration; bacteria that are likely to primarily participate in this essential step of the gut, microbiota cross talk, have been identified. We show in this article that only the cecal and colonic crypts harbor resident microbiota in the mouse and that regardless of the line and breeding origin of these mice, this bacterial population is unexpectedly dominated by aerobic genera. Interestingly, this microbiota resembles the restricted microbiota found in the midgut of invertebrates; thus, the presence of our so-called "crypt-specific core microbiota" (CSCM) in the mouse colon potentially reflects a coevolutionary process under selective conditions that can now be addressed. We suggest that CSCM could play both a protective and a homeostatic role within the colon. This article is setting the bases for such studies, particularly by providing a bona fide--and essentially cultivable--crypt microbiota of reference. IMPORTANCE: Metagenomic typing of the whole-gut luminal microbiome was recently provided, revealing great opportunities for physiological and physiopathological analysis of the host-microbiota interface. On this basis, it appears increasingly important to analyze which niches of the gut exposed to a particular microbiota are of major functional importance, specifically focusing on the crypt, which accounts for permanent epithelial renewal, and to analyze how this microbiota compares to its luminal counterpart in composition and quantity. Crypt-specific core microbiotas may show themselves as important elements regarding crypt protection and homeostasis of its functions.

Molecular evidence of host influences on the evolution and spread of human tapeworms.

The taeniasis/cysticercosis complex is included in the list of neglected zoonotic diseases by the World Health Organization due to its significant impact on public health in tropical areas. Cysticercosis is still endemic in many regions of Asia, Africa and Latin America. Long absent in Europe and in other developed countries, cysticercosis has recently re-emerged in the United States and Canada, due to immigration, travel and local transmission. This has encouraged the use of molecular data to understand better the influence of animal and human hosts on the emergence and spread of Taenia species. The increasing number of mitochondrial sequences now available from human tapeworms and recent advances in computational tools has enabled reconstruction of the biogeography and evolutionary history of these organisms. New molecular data have provided insights into the biogeography of T. solium, T. asiatica and T. saginata. A Bayesian statistical framework using variable evolutionary rates from lineage to lineage has allowed an improved timescale analysis of human tapeworms. The dates of divergence obtained were compared to the timing of evolutionary events in the history of their hosts, based on the hypothesis that Taenia spp. and their hosts share a common history. Herein, we review changes in the definitive and secondary hosts and human interactions that underlie the differentiation and evolution of tapeworms. Species diversification of Taenia seems to be closely linked with the evolution of intermediate hosts in response to climatic events during the Pleistocene. Different genotypes of T. solium emerged when European and Asian wild boar Sus spp. populations diverged. Taenia saginata emerged when wild cattle Bos primigenius evolved and when zebu Bos indicus and taurine Bos taurus ancestors separated. Humans through migrations and later with the development of farming and animal husbandry may have had a significant impact on the spread and diversification of tapeworms. Migrations of Homo erectus from Africa to Asia and later of Homo sapiens facilitated the diversification and dispersal of T. solium and T. saginata populations. The development of animal husbandry, making Sus scrofa and Bos taurus preferential intermediate hosts, led to the worldwide distribution of parasites. New molecular data combined with an innovative dating method allow us to explain the ways in which ancient human migrations promoted the emergence and spread of taeniasis and cysticercosis around the world. Another intriguing phenomenon explained better by our approach is the influence of human settlement on the spread of these parasites in recently inhabited areas. The diverse nature of T. solium currently observed in Madagascar may correspond to multiple imports of the parasite during Austronesian migrations, while in Mexico a recent influence of humans during the colonial period is more likely. Human activities, especially food preparation and husbandry methods, remain responsible for the transmission and persistence of these parasites.

Multi-genome identification and characterization of chlamydiae-specific type III secretion substrates: the Inc proteins.

BACKGROUND: Chlamydiae are obligate intracellular bacteria that multiply in a vacuolar compartment, the inclusion. Several chlamydial proteins containing a bilobal hydrophobic domain are translocated by a type III secretion (TTS) mechanism into the inclusion membrane. They form the family of Inc proteins, which is specific to this phylum. Based on their localization, Inc proteins likely play important roles in the interactions between the microbe and the host. In this paper we sought to identify and analyze, using bioinformatics tools, all putative Inc proteins in published chlamydial genomes, including an environmental species. RESULTS: Inc proteins contain at least one bilobal hydrophobic domain made of two transmembrane helices separated by a loop of less than 30 amino acids. Using bioinformatics tools we identified 537 putative Inc proteins across seven chlamydial proteomes. The amino-terminal segment of the putative Inc proteins was recognized as a functional TTS signal in 90% of the C. trachomatis and C. pneumoniae sequences tested, validating the data obtained in silico. We identified a macro domain in several putative Inc proteins, and observed that Inc proteins are enriched in segments predicted to form coiled coils. A surprisingly large proportion of the putative Inc proteins are not constitutively translocated to the inclusion membrane in culture conditions. CONCLUSIONS: The Inc proteins represent 7 to 10% of each proteome and show a great degree of sequence diversity between species. The abundance of segments with a high probability for coiled coil conformation in Inc proteins support the hypothesis that they interact with host proteins. While the large majority of Inc proteins possess a functional TTS signal, less than half may be constitutively translocated to the inclusion surface in some species. This suggests the novel finding that translocation of Inc proteins may be regulated by as-yet undetermined mechanisms.

Molecular analyses of the microbial community composition of an anoxic basin of a municipal wastewater treatment plant reveal a novel lineage of proteobacteria.

A culture-independent molecular phylogenetic approach was used to study prokaryotic diversity in an anoxic activated sludge from a municipal wastewater treatment plant. Two 16S rRNA gene libraries were constructed using total genomic DNA and amplified by polymerase chain reaction using primers specific for archaeal or bacterial domains. Phylogenetic analysis of 132 and 249 almost full-length 16S rRNA genes for Archaea and Bacteria, respectively, was done using the ARB software package. Phylogenetic groups affiliated with the Archaea belong to Euryarchaeota (93.8% of the operational taxonomic units [OTUs]) and Crenarchaeota (6.2% of the OTUs). Within the bacterial library, 84.8% of the OTUs represent novel putative phylotypes never described before and affiliated with ten divisions. The Proteobacteria phylum is the most abundant and diversified phylogenetic group representing 60.4% of the OTUs, followed by Bacteroidetes (22.1%) and gram-positives (6.1%). Interestingly, we detected a novel Proteobacteria monophyletic group distinct from the five known subclasses, which we named New Lineage of Proteobacteria (NLP) lineage, and it is composed of eight clones representing 4.6% of the Proteobacteria. A new 16S rRNA-targeted hybridization probe was designed and fluorescent in situ hybridization analyses shows representatives of NLP as cocci-shaped microorganisms. The Chloroflexi, Acidobacterium, and Nitrospira phyla and TM7 candidate division are each represented by ≤3% of clone sequences. A comprehensive set of eight 16S and 23S rRNA-targeted oligonucleotide probes was used to quantify these major groups by dot blot hybridization within 12 samples. The Proteobacteria accounted for 82.5 ± 4.9%, representing the most abundant phyla. The Bacteroidetes and Planctomycetales groups accounted for 4.9 ± 1.3% and 4 ± 1.7%, respectively. Firmicutes and Actinobacteria together accounted for only 1.9 ± 0.5%. The set of probes covers 93.4 ± 14% of the total bacterial population rRNA within the anoxic basin.

The pig tapeworm Taenia solium, the cause of cysticercosis: Biogeographic (temporal and spacial) origins in Madagascar.

Cysticercosis is a serious public health problem in Madagascar. The prevalence rate of active cysticercosis reached 21% in regions with a high level of livestock farming. Taenia solium of African-American and Asian genotypes are both present on the island. The times of divergence of the 13 specimens studied suggests a very ancient diversification of T. solium. These events are widely thought to be prior to the domestication of pigs, and seem to follow the expansion of Homo in Asia. Multiple human migrations and the diversity of potential intermediate hosts may have led to a complex epidemiological situation on the island.

Molecular analysis of a large subtelomeric nucleotide-binding-site-leucine-rich-repeat family in two representative genotypes of the major gene pools of Phaseolus vulgaris.

In common bean, the B4 disease resistance gene cluster is a complex cluster localized at the end of linkage group (LG) B4, containing at least three R specificities to the fungus Colletotrichum lindemuthianum. To investigate the evolution of this R cluster since the divergence of Andean and Mesoamerican gene pools, DNA sequences were characterized from two representative genotypes of the two major gene pools of common bean (BAT93: Mesoamerican; JaloEEP558: Andean). Sequences encoding 29 B4-CC nucleotide-binding-site-leucine-rich-repeat (B4-CNL) genes were determined-12 from JaloEEP558 and 17 from BAT93. Although sequence exchange events were identified, phylogenetic analyses revealed that they were not frequent enough to lead to homogenization of B4-CNL sequences within a haplotype. Genetic mapping based on pulsed-field gel electrophoresis separation confirmed that the B4-CNL family is a large family specific to one end of LG B4 and is present at two distinct blocks separated by 26 cM. Fluorescent in situ hybridization on meiotic pachytene chromosomes revealed that two B4-CNL blocks are located in the subtelomeric region of the short arm of chromosome 4 on both sides of a heterochromatic block (knob), suggesting that this peculiar genomic environment may favor the proliferation of a large R gene cluster.

Cloacibacillus evryensis gen. nov., sp. nov., a novel asaccharolytic, mesophilic, amino-acid-degrading bacterium within the phylum 'Synergistetes', isolated from an anaerobic sludge digester.

A novel anaerobic, mesophilic, amino-acid-utilizing bacterium, strain 158T, was isolated from an anaerobic digester of a wastewater treatment plant. Cells of strain 158T were non-motile, rod-shaped (2.0-3.0 x 0.8-1.0 microm) and stained Gram-negative. Optimal growth occurred at 37 degrees C and pH 7.0 in an anaerobic basal medium containing 1 % Casamino acids. Strain 158T fermented arginine, histidine, lysine and serine and showed growth on yeast extract, brain-heart infusion (BHI) medium and tryptone, but not on carbohydrates, organic acids or alcohols. The end products of degradation were: acetate, butyrate, H2 and CO2 from arginine; acetate, propionate, butyrate, H2 and CO2 from lysine; and acetate, propionate, butyrate, valerate, H2 and CO2 from histidine, serine, BHI medium, Casamino acids and tryptone. The DNA G+C content was 55.8 mol%. The 16S rRNA gene sequence of strain 158T showed only 92.6 % sequence similarity with that of Synergistes jonesii, the only described species of the 'Synergistes' group. The major cellular fatty acids were iso-C(15:0) (16.63 %), iso-C(15:0) 3-OH (12.41 %) and C(17:1)omega6c (9.46 %) and the polar fatty acids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and phosphatidylmonomethylamine; these fatty acid profiles did not resemble those of any recognized bacterial species. Due to the considerable differences in genotypic, phenotypic and phylogenetic characteristics between strain 158T and those of its nearest relative, it is proposed that strain 158T represents a novel species in a new genus, Cloacibacillus evryensis gen. nov., sp. nov., in the phylum 'Synergistetes'. The type strain is 158T (=DSM 19522T=JCM 14828T).