Identification of polyhydroxyalkanoates in Halococcus and other haloarchaeal species

Polyhydroxyalkanoates (PHAs) are accumulated in many prokaryotes. Several members of the Halobacteriaceae produce poly-3-hydroxybutyrate (PHB), but it is not known if this is a general property of the family. We evaluated identification methods for PHAs with 20 haloarchaeal species, three of them isolates from Permian salt. Staining with Sudan Black B, Nile Blue A, or Nile Red was applied to screen for the presence of PHAs. Transmission electron microscopy and 1H-nuclear magnetic resonance spectroscopy were used for visualization of PHB granules and chemical confirmation of PHAs in cell extracts, respectively. We report for the first time the production of PHAs by Halococcus sp. (Halococcus morrhuae DSM 1307T, Halococcus saccharolyticus DSM 5350T, Halococcus salifodinae DSM 8989T, Halococcus dombrowskii DSM 14522T, Halococcus hamelinensis JCM 12892T, Halococcus qingdaonensis JCM 13587T), Halorubrum sp. (Hrr. coriense DSM 10284T, Halorubrum chaoviator DSM 19316T, Hrr. chaoviator strains NaxosII and AUS-1), haloalkaliphiles (Natronobacterium gregoryi NCMB 2189T, Natronococcus occultus DSM 3396T) and Halobacterium noricense DSM 9758T. No PHB was detected in Halobacterium salinarum NRC-1 ATCC 700922, Hbt. salinarum R1 and Haloferax volcanii DSM 3757T. Most species synthesized PHAs when growing in synthetic as well as in complex medium. The polyesters were generally composed of PHB and poly-ß-hydroxybutyrate-co-3-hydroxyvalerate (PHBV). Available genomic data suggest the absence of PHA synthesis in some haloarchaea and in all other Euryarchaeota and Crenarchaeota. Homologies between haloarchaeal and bacterial PHA synthesizing enzymes had indicated to some authors probable horizontal gene transfer, which, considering the data obtained in this study, may have occurred already before Permian times

An Environment-Sensitive Synthetic Microbial Ecosystem

Microbial ecosystems have been widely used in industrial production, but the inter-relationships of organisms within them haven't been completely clarified due to complex composition and structure of natural microbial ecosystems. So it is challenging for ecologists to get deep insights on how ecosystems function and interplay with surrounding environments. But the recent progresses in synthetic biology show that construction of artificial ecosystems where relationships of species are comparatively clear could help us further uncover the meadow of those tiny societies. By using two quorum-sensing signal transduction circuits, this research designed, simulated and constructed a synthetic ecosystem where various population dynamics formed by changing environmental factors. Coherent experimental data and mathematical simulation in our study show that different antibiotics levels and initial cell densities can result in correlated population dynamics such as extinction, obligatory mutualism, facultative mutualism and commensalism. This synthetic ecosystem provides valuable information for addressing questions in ecology and may act as a chassis for construction of more complex microbial ecosystems

Analysis of the unexplored features of rrs (16S rDNA) of the Genus Clostridium

<p>Abstract</p> <p>Background</p> <p>Bacterial taxonomy and phylogeny based on <it>rrs </it>(16S rDNA) sequencing is being vigorously pursued. In fact, it has been stated that novel biological findings are driven by comparison and integration of massive data sets. In spite of a large reservoir of <it>rrs </it>sequencing data of 1,237,963 entries, this analysis invariably needs supplementation with other genes. The need is to divide the genetic variability within a taxa or genus at their <it>rrs </it>phylogenetic boundaries and to discover those fundamental features, which will enable the bacteria to naturally fall within them. Within the large bacterial community, <it>Clostridium </it>represents a large genus of around 110 species of significant biotechnological and medical importance. Certain <it>Clostridium </it>strains produce some of the deadliest toxins, which cause heavy economic losses. We have targeted this genus because of its high genetic diversity, which does not allow accurate typing with the available molecular methods.</p> <p>Results</p> <p>Seven hundred sixty five <it>rrs </it>sequences (> 1200 nucleotides, nts) belonging to 110 <it>Clostridium </it>species were analyzed. On the basis of 404 <it>rrs </it>sequences belonging to 15 <it>Clostridium </it>species, we have developed species specific: (i) phylogenetic framework, (ii) signatures (30 nts) and (iii) <it>in silico </it>restriction enzyme (14 Type II REs) digestion patterns. These tools allowed: (i) species level identification of 95 <it>Clostridium </it>sp. which are presently classified up to genus level, (ii) identification of 84 novel <it>Clostridium </it>spp. and (iii) potential reduction in the number of <it>Clostridium </it>species represented by small populations.</p> <p>Conclusions</p> <p>This integrated approach is quite sensitive and can be easily extended as a molecular tool for diagnostic and taxonomic identification of any microbe of importance to food industries and health services. Since rapid and correct identification allows quicker diagnosis and consequently treatment as well, it is likely to lead to reduction in economic losses and mortality rates.</p

Clostridia in Premature Neonates' Gut: Incidence, Antibiotic Susceptibility, and Perinatal Determinants Influencing Colonization

Although premature neonates (PN) gut microbiota has been studied, data about gut clostridial colonization in PN are scarce. Few studies have reported clostridia colonization in PN whereas Bacteroides and bifidobacteria have been seldom isolated. Such aberrant gut microbiota has been suggested to be a risk factor for the development of intestinal infections. Besides, PN are often treated by broad spectrum antibiotics, but little is known about how antibiotics can influence clostridial colonization based on their susceptibility patterns. The aim of this study was to report the distribution of Clostridium species isolated in feces from PN and to determine their antimicrobial susceptibility patterns. Additionally, clostridial colonization perinatal determinants were analyzed.Of the 76 PN followed until hospital discharge in three French neonatal intensive care units (NICUs), 79% were colonized by clostridia. Clostridium sp. colonization, with a high diversity of species, increased throughout the hospitalization. Antibiotic courses had no effect on the clostridial colonization incidence although strains were found susceptible (except C. difficile) to anti-anaerobe molecules tested. However, levels of colonization were decreased by either antenatal or neonatal (during more than 10 days) antibiotic courses (p = 0.006 and p = 0.001, respectively). Besides, incidence of colonization was depending on the NICU (p = 0.048).This study shows that clostridia are part of the PN gut microbiota. It provides for the first time information on the status of clostridia antimicrobial susceptibility in PN showing that strains were susceptible to most antibiotic molecules. Thus, the high prevalence of this genus is not linked to a high degree of resistance to antimicrobial agents or to the use of antibiotics in NICUs. The main perinatal determinant influencing PN clostridia colonization appears to be the NICU environment

Phylogeny in Aid of the Present and Novel Microbial Lineages: Diversity in Bacillus

Bacillus represents microbes of high economic, medical and biodefense importance. Bacillus strain identification based on 16S rRNA sequence analyses is invariably limited to species level. Secondly, certain discrepancies exist in the segregation of Bacillus subtilis strains. In the RDP/NCBI databases, out of a total of 2611 individual 16S rDNA sequences belonging to the 175 different species of the genus Bacillus, only 1586 have been identified up to species level. 16S rRNA sequences of Bacillus anthracis (153 strains), B. cereus (211 strains), B. thuringiensis (108 strains), B. subtilis (271 strains), B. licheniformis (131 strains), B. pumilus (83 strains), B. megaterium (47 strains), B. sphaericus (42 strains), B. clausii (39 strains) and B. halodurans (36 strains) were considered for generating species-specific framework and probes as tools for their rapid identification. Phylogenetic segregation of 1121, 16S rDNA sequences of 10 different Bacillus species in to 89 clusters enabled us to develop a phylogenetic frame work of 34 representative sequences. Using this phylogenetic framework, 305 out of 1025, 16S rDNA sequences presently classified as Bacillus sp. could be identified up to species level. This identification was supported by 20 to 30 nucleotides long signature sequences and in silico restriction enzyme analysis specific to the 10 Bacillus species. This integrated approach resulted in identifying around 30% of Bacillus sp. up to species level and revealed that B. subtilis strains can be segregated into two phylogenetically distinct groups, such that one of them may be renamed

3,4-Methylenedioxymethamphetamine (MDMA) neurotoxicity in rats: a reappraisal of past and present findings

RATIONALE: 3,4-Methylenedioxymethamphetamine (MDMA) is a widely abused illicit drug. In animals, high-dose administration of MDMA produces deficits in serotonin (5-HT) neurons (e.g., depletion of forebrain 5-HT) that have been interpreted as neurotoxicity. Whether such 5-HT deficits reflect neuronal damage is a matter of ongoing debate. OBJECTIVE: The present paper reviews four specific issues related to the hypothesis of MDMA neurotoxicity in rats: (1) the effects of MDMA on monoamine neurons, (2) the use of “interspecies scaling” to adjust MDMA doses across species, (3) the effects of MDMA on established markers of neuronal damage, and (4) functional impairments associated with MDMA-induced 5-HT depletions. RESULTS: MDMA is a substrate for monoamine transporters, and stimulated release of 5-HT, NE, and DA mediates effects of the drug. MDMA produces neurochemical, endocrine, and behavioral actions in rats and humans at equivalent doses (e.g., 1–2 mg/kg), suggesting that there is no reason to adjust doses between these species. Typical doses of MDMA causing long-term 5-HT depletions in rats (e.g., 10–20 mg/kg) do not reliably increase markers of neurotoxic damage such as cell death, silver staining, or reactive gliosis. MDMA-induced 5-HT depletions are accompanied by a number of functional consequences including reductions in evoked 5-HT release and changes in hormone secretion. Perhaps more importantly, administration of MDMA to rats induces persistent anxiety-like behaviors in the absence of measurable 5-HT deficits. CONCLUSIONS: MDMA-induced 5-HT depletions are not necessarily synonymous with neurotoxic damage. However, doses of MDMA which do not cause long-term 5-HT depletions can have protracted effects on behavior, suggesting even moderate doses of the drug may pose risks

Why Functional Pre-Erythrocytic and Bloodstage Malaria Vaccines Fail: A Meta-Analysis of Fully Protective Immunizations and Novel Immunological Model

Background: Clinically protective malaria vaccines consistently fail to protect adults and children in endemic settings, and at best only partially protect infants. Methodology/Principal Findings: We identify and evaluate 1916 immunization studies between 1965-February 2010, and exclude partially or nonprotective results to find 177 completely protective immunization experiments. Detailed reexamination reveals an unexpectedly mundane basis for selective vaccine failure: live malaria parasites in the skin inhibit vaccine function. We next show published molecular and cellular data support a testable, novel model where parasite-host interactions in the skin induce malaria-specific regulatory T cells, and subvert early antigen-specific immunity to parasite-specific immunotolerance. This ensures infection and tolerance to reinfection. Exposure to Plasmodium-infected mosquito bites therefore systematically triggers immunosuppression of endemic vaccine-elicited responses. The extensive vaccine trial data solidly substantiate this model experimentally. Conclusions/Significance: We conclude skinstage-initiated immunosuppression, unassociated with bloodstage parasites, systematically blocks vaccine function in the field. Our model exposes novel molecular and procedural strategies to significantly and quickly increase protective efficacy in both pipeline and currently ineffective malaria vaccines, and forces fundamental reassessment of central precepts determining vaccine development. This has major implications fo