Field testing of arsenic in groundwater samples of Bangladesh using a test kit based on lyophilized bioreporter bacteria.

A test kit based on living, lyophilized bacterial bioreporters emitting bioluminescence as a response to arsenite and arsenate was applied during a field campaign in six villages across Bangladesh. Bioreporter field measurements of arsenic in groundwater from tube wells were in satisfying agreement with the results of spectroscopic analyses of the same samples conducted in the lab. The practicability of the bioreporter test in terms of logistics and material requirements, suitability for high sample throughput, and waste disposal was much better than that of two commercial chemical test kits that were included as references. The campaigns furthermore demonstrated large local heterogeneity of arsenic in groundwater, underscoring the use of well switching as an effective remedy to avoid high arsenic exposure.

RP 59500 prophylaxis of experimental endocarditis due to erythromycin-susceptible and -resistant isogenic pairs of viridans group streptococci.

RP 59500 is a new injectable streptogramin composed of two synergistic components (quinupristin and dalfopristin) which are active against a number of erythromycin-susceptible and -resistant gram-positive bacteria. The following experiments investigate the ability of RP 59500 to prevent experimental endocarditis due to either of two erythromycin-susceptible streptococcal isolates or their constitutively erythromycin-resistant Tn916 delta E transconjugants. RP 59500 had low MICs (0.125 to 0.5 mg/liter) for all four test organisms and was substantially bactericidal in vitro. Rats with catheter-induced aortic vegetations were given single-dose antibiotic prophylaxis 30 to 60 min before bacterial inoculation through a computerized pump system which permitted the simulation of drug kinetics for humans produced by either 7 mg of RP 59500 per kg of body weight or 1 g of vancomycin. Single-dose RP 59500 prophylaxis successfully prevented endocarditis due to both the erythromycin-susceptible parent strains and their erythromycin-resistant derivatives in rats challenged with the minimal inoculum infecting 90% of controls. In addition, RP 59500 also prevented infection in animals challenged with fivefold-larger inocula of the erythromycin-susceptible parent strains. Vancomycin successfully prevented endocarditis due to any of the four test organisms. These results underline the in vivo efficacy of RP 59500 against both erythromycin-susceptible and -resistant streptococci. Such good results against the resistant strains would not be expected with erythromycin or clindamycin, which are the standard macrolidelincosamide-streptogramin antibiotics used for endocarditis prophylaxis in humans. An oral form of RP 59500 which might advantageously replace some of the older prophylactic regimens is currently being developed.

Genetics of narcolepsy and other major sleep disorders.

One third of the population is affected by a sleep disorder with a major social, medical, and economic impact. Although very little is known about the genetics of normal sleep, familial and twin studies indicate an important influence of genetic factors. Most sleep disorders run in families and in several of them the contribution of genetic factors is increasingly recognised. With recent advances in the genetics of narcolepsy and the role of the hypocretin/orexin system, the possibility that other gene defects may contribute to the pathophysiology of major sleep disorders is worth indepth investigation.

A new factor stimulating peptidoglycan hydrolysis to separate daughter cells in Caulobacter crescentus.

Cell division in Gram-negative bacteria involves the co-ordinated invagination of the three cell envelope layers to form two new daughter cell poles. This complex process starts with the polymerization of the tubulin-like protein FtsZ into a Z-ring at mid-cell, which drives cytokinesis and recruits numerous other proteins to the division site. These proteins are involved in Z-ring constriction, inner- and outer-membrane invagination, peptidoglycan remodelling and daughter cell separation. Three papers in this issue of Molecular Microbiology, from the teams of Lucy Shapiro, Martin Thanbichler and Christine Jacobs-Wagner, describe a novel protein, called DipM for Division Involved Protein with LysM domains, that is required for cell division in Caulobacter crescentus. DipM localizes to the mid-cell during cell division, where it is necessary for the hydrolysis of the septal peptidoglycan to remodel the cell wall. Loss of DipM results in severe defects in cell envelope constriction, which is deleterious under fast-growth conditions. State-of-the-art microscopy experiments reveal that the peptidoglycan is thicker and that the cell wall is incorrectly organized in DipM-depleted cells compared with wild-type cells, demonstrating that DipM is essential for reorganizing the cell wall at the division site, for envelope invagination and cell separation in Caulobacter.

The microbiological signature of human cutaneous leishmaniasis lesions exhibits restricted bacterial diversity compared to healthy skin

Localised cutaneous leishmaniasis (LCL) is the most common form of cutaneous leishmaniasis characterised by single or multiple painless chronic ulcers, which commonly presents with secondary bacterial infection. Previous culture-based studies have found staphylococci, streptococci, and opportunistic pathogenic bacteria in LCL lesions, but there have been no comparisons to normal skin. In addition, this approach has strong bias for determining bacterial composition. The present study tested the hypothesis that bacterial communities in LCL lesions differ from those found on healthy skin (HS). Using a high throughput amplicon sequencing approach, which allows for better populational evaluation due to greater depth coverage and the Quantitative Insights Into Microbial Ecology pipeline, we compared the microbiological signature of LCL lesions with that of contralateral HS from the same individuals.Streptococcus, Staphylococcus,Fusobacterium and other strict or facultative anaerobic bacteria composed the LCL microbiome. Aerobic and facultative anaerobic bacteria found in HS, including environmental bacteria, were significantly decreased in LCL lesions (p < 0.01). This paper presents the first comprehensive microbiome identification from LCL lesions with next generation sequence methodology and shows a marked reduction of bacterial diversity in the lesions.

Complex genetics in idiopathic hypogonadotropic hypogonadism.

Idiopathic hypogonadotropic hypogonadism (IHH) is an important human disease model. Investigations of the genetics of IHH have facilitated insights into critical pathways regulating sexual maturation and fertility. IHH has been traditionally considered a monogenic disorder. This model holds that a single gene defect is responsible for the disease in each patient. In the case of IHH, 30% of cases are explained by mutations in one of eleven genes. In recent years, several lines of evidence have challenged the monogenic paradigm in IHH. First, disease-associated mutations display striking incomplete penetrance and variable expressivity within and across IHH families. Second, each locus is responsible for only a small percentage of cases. Third, more than one disease-associated mutation seems to be segregating in some families with IHH, and their combined or separate presence in individuals accounts for the variability in disease severity. Finally, IHH is not strictly a congenital and life-long disorder; occasionally it manifests itself during adulthood (adult-onset IHH); in other cases, the disease is not permanent, as evidenced by normal activity of the hypothalamic-pituitary-gonadal axis after discontinuation of treatment in adulthood (IHH reversal). Together, these observations suggest that IHH is not strictly a monogenic mendelian disease, as previously thought. Rather, it is emerging as a digenic, and potentially oligogenic disease, in which hormonal and/or environmental factors may critically influence genetic predisposition and clinical course. Future investigations of IHH should characterize the extent of the involvement of multiple genes in disease pathogenesis, and elucidate the contributions of epigenetic factors.

Assessment of airborne microorganisms by real-time PCR: optimistic findings and research challenges

Most airborne microorganisms are natural components of our ecosystem. Soil, vegetation and animals, including humans, are sources for aerial release of these living or dead cells. In the past, assessment of airborne microorganisms was mainly restricted to occupational health concerns. Indeed, in several occupations, exposure to very high concentrations of non-infectious airborne bacteria and fungi, result in allergenic, toxic or irritant reactions. Recently, the threat of bioterrorism and pandemics have highlighted the urgent need to increase knowledge of bioaerosol ecology. More fundamentally, airborne bacterial and fungal communities begin to draw much more consideration from environmental microbiologists, who have neglected this area for a long time. This increased interest of scientists is to a great part due to the development and use of real-time PCR techniques to identify and quantify airborne microorganisms. Even if the advantages of the PCR technology are obvious, researchers are confronted with new problems. This review describes the methodological state of the art in bioaerosols field and emphasizes the future challenges and perspectives of the real-time PCR-based methods for airborne microorganism studies.

Catabolite repression control of pyocyanin biosynthesis at an intersection of primary and secondary metabolism in Pseudomonas aeruginosa.

In Pseudomonas aeruginosa, the catabolite repression control (Crc) protein repressed the formation of the blue pigment pyocyanin in response to a preferred carbon source (succinate) by interacting with phzM mRNA, which encodes a key enzyme in pyocyanin biosynthesis. Crc bound to an extended imperfect recognition sequence that was interrupted by the AUG translation initiation codon.

Four main virotypes among extended-spectrum-β-lactamase-producing isolates of Escherichia coli O25b:H4-B2-ST131: bacterial, epidemiological, and clinical characteristics.

A total of 1,021 extended-spectrum-β-lactamase-producing Escherichia coli (ESBLEC) isolates obtained in 2006 during a Spanish national survey conducted in 44 hospitals were analyzed for the presence of the O25b:H4-B2-ST131 (sequence type 131) clonal group. Overall, 195 (19%) O25b-ST131 isolates were detected, with prevalence rates ranging from 0% to 52% per hospital. Molecular characterization of 130 representative O25b-ST131 isolates showed that 96 (74%) were positive for CTX-M-15, 15 (12%) for CTX-M-14, 9 (7%) for SHV-12, 6 (5%) for CTX-M-9, 5 (4%) for CTX-M-32, and 1 (0.7%) each for CTX-M-3 and the new ESBL enzyme CTX-M-103. The 130 O25b-ST131 isolates exhibited relatively high virulence scores (mean, 14.4 virulence genes). Although the virulence profiles of the O25b-ST131 isolates were fairly homogeneous, they could be classified into four main virotypes based on the presence or absence of four distinctive virulence genes: virotypes A (22%) (afa FM955459 positive, iroN negative, ibeA negative, sat positive or negative), B (31%) (afa FM955459 negative, iroN positive, ibeA negative, sat positive or negative), C (32%) (afa FM955459 negative, iroN negative, ibeA negative, sat positive), and D (13%) (afa FM955459 negative, iroN positive or negative, ibeA positive, sat positive or negative). The four virotypes were also identified in other countries, with virotype C being overrepresented internationally. Correspondingly, an analysis of XbaI macrorestriction profiles revealed four major clusters, which were largely virotype specific. Certain epidemiological and clinical features corresponded with the virotype. Statistically significant virotype-specific associations included, for virotype B, older age and a lower frequency of infection (versus colonization), for virotype C, a higher frequency of infection, and for virotype D, younger age and community-acquired infections. In isolates of the O25b:H4-B2-ST131 clonal group, these findings uniquely define four main virotypes, which are internationally distributed, correspond with pulsed-field gel electrophoresis (PFGE) profiles, and exhibit distinctive clinical-epidemiological associations.

Colistin resistance in a clinical Acinetobacter baumannii strain appearing after colistin treatment: effect on virulence and bacterial fitness.

The fitness and virulence costs associated with the clinical acquisition of colistin resistance by Acinetobacter baumannii were evaluated. The growth of strain CR17 (colistin resistant) was less than that of strain CS01 (colistin susceptible) when the strains were grown in competition (72-h competition index, 0.008). In a murine sepsis model, CS01 and CR17 reached spleen concentrations when coinfecting of 9.31 and 6.97 log10 CFU/g, respectively, with an in vivo competition index of 0.016. Moreover, CS01 was more virulent than CR17 with respect to mortality and time to death.

Mycobacterium tuberculosis transmission in a country with low tuberculosis incidence: role of immigration and HIV infection.

Immigrants from high-burden countries and HIV-coinfected individuals are risk groups for tuberculosis (TB) in countries with low TB incidence. Therefore, we studied their role in transmission of Mycobacterium tuberculosis in Switzerland. We included all TB patients from the Swiss HIV Cohort and a sample of patients from the national TB registry. We identified molecular clusters by spoligotyping and mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) analysis and used weighted logistic regression adjusted for age and sex to identify risk factors for clustering, taking sampling proportions into account. In total, we analyzed 520 TB cases diagnosed between 2000 and 2008; 401 were foreign born, and 113 were HIV coinfected. The Euro-American M. tuberculosis lineage dominated throughout the study period (378 strains; 72.7%), with no evidence for another lineage, such as the Beijing genotype, emerging. We identified 35 molecular clusters with 90 patients, indicating recent transmission; 31 clusters involved foreign-born patients, and 15 involved HIV-infected patients. Birth origin was not associated with clustering (adjusted odds ratio [aOR], 1.58; 95% confidence interval [CI], 0.73 to 3.43; P = 0.25, comparing Swiss-born with foreign-born patients), but clustering was reduced in HIV-infected patients (aOR, 0.49; 95% CI, 0.26 to 0.93; P = 0.030). Cavitary disease, male sex, and younger age were all associated with molecular clustering. In conclusion, most TB patients in Switzerland were foreign born, but transmission of M. tuberculosis was not more common among immigrants and was reduced in HIV-infected patients followed up in the national HIV cohort study. Continued access to health services and clinical follow-up will be essential to control TB in this population.

Describing ancient horizontal gene transfers at the nucleotide and gene levels by comparative pathogenicity island genometrics.

MOTIVATION: Lateral gene transfer is a major mechanism contributing to bacterial genome dynamics and pathovar emergence via pathogenicity island (PAI) spreading. However, since few of these genomic exchanges are experimentally reproducible, it is difficult to establish evolutionary scenarios for the successive PAI transmissions between bacterial genera. Methods initially developed at the gene and/or nucleotide level for genomics, i.e. comparisons of concatenated sequences, ortholog frequency, gene order or dinucleotide usage, were combined and applied here to homologous PAIs: we call this approach comparative PAI genometrics. RESULTS: YAPI, a Yersinia PAI, and related islands were compared with measure evolutionary relationships between related modules. Through use of our genometric approach designed for tracking codon usage adaptation and gene phylogeny, an ancient inter-genus PAI transfer was oriented for the first time by characterizing the genomic environment in which the ancestral island emerged and its subsequent transfers to other bacterial genera.

Genetics of colouration in birds.

Establishing the links between phenotype and genotype is of great importance for resolving key questions about the evolution, maintenance and adaptive function of phenotypic variation. Bird colouration is one of the most studied systems to investigate the role of natural and sexual selection in the evolution of phenotypic diversity. Given the recent advances in molecular tools that allow discovering genetic polymorphisms and measuring gene and protein expression levels, it is timely to review the literature on the genetics of bird colouration. The present study shows that melanin-based colour phenotypes are often associated with mutations at melanogenic genes. Differences in melanin-based colouration are caused by switches of eumelanin to pheomelanin production or by changes in feather keratin structure, melanoblast migration and differentiation, as well as melanosome structure. Similar associations with other types of colourations are difficult to establish, because our knowledge about the molecular genetics of carotenoid-based and structural colouration is quasi inexistent. This discrepancy stems from the fact that only melanin-based colouration shows pronounced heritability estimates, i.e. the resemblance between related individuals is usually mainly explained by genetic factors. In contrast, the expression of carotenoid-based colouration is phenotypically plastic with a high sensitivity to variation in environmental conditions. It therefore appears that melanin-based colour traits are prime systems to understand the genetic basis of phenotypic variation. In this context, birds have a great potential to bring us to new frontiers where many exciting discoveries will be made on the genetics of phenotypic traits, such as colouration. In this context, a major goal of our review is to suggest a number of exciting future avenues.

Development of a multistrain bacterial bioreporter platform for the monitoring of hydrocarbon contaminants in marine environments.

Petroleum hydrocarbons are common contaminants in marine and freshwater aquatic habitats, often occurring as a result of oil spillage. Rapid and reliable on-site tools for measuring the bioavailable hydrocarbon fractions, i.e., those that are most likely to cause toxic effects or are available for biodegradation, would assist in assessing potential ecological damage and following the progress of cleanup operations. Here we examined the suitability of a set of different rapid bioassays (2-3 h) using bacteria expressing the LuxAB luciferase to measure the presence of short-chain linear alkanes, monoaromatic and polyaromatic compounds, biphenyls, and DNA-damaging agents in seawater after a laboratory-scale oil spill. Five independent spills of 20 mL of NSO-1 crude oil with 2 L of seawater (North Sea or Mediterranean Sea) were carried out in 5 L glass flasks for periods of up to 10 days. Bioassays readily detected ephemeral concentrations of short-chain alkanes and BTEX (i.e., benzene, toluene, ethylbenzene, and xylenes) in the seawater within minutes to hours after the spill, increasing to a maximum of up to 80 muM within 6-24 h, after which they decreased to low or undetectable levels. The strong decrease in short-chain alkanes and BTEX may have been due to their volatilization or biodegradation, which was supported by changes in the microbial community composition. Two- and three-ring PAHs appeared in the seawater phase after 24 h with a concentration up to 1 muM naphthalene equivalents and remained above 0.5 muM for the duration of the experiment. DNA-damage-sensitive bioreporters did not produce any signal with the oil-spilled aqueous-phase samples, whereas bioassays for (hydroxy)biphenyls showed occasional responses. Chemical analysis for alkanes and PAHs in contaminated seawater samples supported the bioassay data, but did not show the typical ephemeral peaks observed with the bioassays. We conclude that bacterium-based bioassays can be a suitable alternative for rapid on-site quantitative measurement of hydrocarbons in seawater.