Importance of the Two Dissimilatory (Nar) Nitrate Reductases in the Growth and Nitrate Reduction of the Methylotrophic Marine Bacterium Methylophaga nitratireducenticrescens JAM1

Methylophaga nitratireducenticrescens JAM1 is the only reported Methylophaga species capable of growing under anaerobic conditions with nitrate as electron acceptor. Its genome encodes a truncated denitrification pathway, which includes two nitrate reductases, Nar1 and Nar2; two nitric oxide reductases, Nor1 and Nor2; and one nitrous oxide reductase, Nos; but no nitrite reductase (NirK or NirS). The transcriptome of strain JAM1 cultivated with nitrate and methanol under anaerobic conditions showed the genes for these enzymes were all expressed. We investigated the importance of Nar1 and Nar2 by knocking out narG1, narG2 or both genes. Measurement of the specific growth rate and the specific nitrate reduction rate of the knockout mutants JAM1ΔnarG1 (Nar1) and JAM1ΔnarG2 (Nar2) clearly demonstrated that both Nar systems contributed to the growth of strain JAM1 under anaerobic conditions, but at different levels. The JAM1ΔnarG1 mutant exhibited an important decrease in the nitrate reduction rate that consequently impaired its growth under anaerobic conditions. In JAM1ΔnarG2, the mutation induced a 20-h lag period before nitrate reduction occurred at specific rate similar to that of strain JAM1. The disruption of narG1 did not affect the expression of narG2. However, the expression of the Nar1 system was highly downregulated in the presence of oxygen with the JAM1ΔnarG2 mutant. These results indicated Nar1 is the major nitrate reductase in strain JAM1 but Nar2 appears to regulate the expression of Nar1

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency-Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research.Peer reviewe

The pentachlorophenol-dehalogenating Desulfitobacterium hafniense strain PCP-1.

International audienceIn this report, a complete description of Desulfitobacterium hafniense strain PCP-1 is presented. The D. hafniense strain PCP-1 was isolated from a methanogenic consortium for its capacity to dehalogenate pentachlorophenol (PCP) into 3-chlorophenol. This strain is also capable of dehalogenating several other chloroaromatic compounds and tetrachloroethene into trichloroethene. Four gene loci encoding putative chlorophenol-reductive dehalogenases (CprA2 to CprA5) were detected, and the products of two of these loci have been demonstrated to dechlorinate different chlorinated phenols. Strain PCP-1 was used in laboratory-scale bioprocesses to degrade PCP present in contaminated environments. Desulfitobacterium hafniense PCP-1 is an excellent candidate for the development of efficient bioprocesses to degrade organohalide compounds

Co-culturing Hyphomicrobium nitrativorans strain NL23 and Methylophaga nitratireducenticrescens strain JAM1 allows sustainable denitrifying activities under marine conditions

Background Hyphomicrobium nitrativorans strain NL23 and Methylophaga nitratireducenticrescens strain JAM1 are the principal bacteria involved in the denitrifying activities of a methanol-fed, fluidized-bed marine denitrification system. Strain NL23 possesses the complete denitrification pathway, but cannot grow under marine conditions in pure cultures. Strain JAM1 is a marine bacterium that lacks genes encoding a dissimilatory nitrite (NO2−) reductase and therefore cannot reduce NO2−. Here, we report the characterization of some of their physiological traits that could influence their co-habitation. We also perform co-cultures to assess the potential synergy between the two strains under marine and denitrifying conditions. Methodology Anoxic planktonic pure cultures of both strains were grown with different concentrations of nitrate (NO3−). Anoxic planktonic co-cultures could only be cultured on low NaCl concentrations for strain NL23 to grow. Biofilm co-cultures were achieved in a 500-mL bioreactor, and operated under denitrifying conditions with increasing concentrations of NaCl. NO3− and NO2− concentrations and the protein content were measured to derive the denitrification rates. The concentrations of both strains in co-cultures were determined by quantitative PCR (qPCR). Ectoine concentration was measured by mass spectrometry in the biofilm co-culture. The biofilm was visualized by fluorescence in situ hybridization. Reverse-transcription-qPCR and RNA-seq approaches were used to assess changes in the expression profiles of genes involved in the nitrogen pathways in the biofilm cultures. Results Planktonic pure cultures of strain JAM1 had a readiness to reduce NO3− with no lag phase for growth in contrast to pure cultures of strain NL23, which had a 2-3 days lag phase before NO3− starts to be consumed and growth to occur. Compared to strain NL23, strain JAM1 has a higher µmax for growth and higher specific NO3− reduction rates. Denitrification rates were twice higher in the planktonic co-cultures than those measured in strain NL23 pure cultures. The biofilm co-cultures showed sustained denitrifying activities and surface colonization by both strains under marine conditions. Increase in ectoine concentrations was observed in the biofilm co-culture with the increase of NaCl concentrations. Changes in the relative transcript levels were observed in the biofilm culture with genes encoding NapA and NapGH in strain NL23. The type of medium had a great impact on the expression of genes involved in the N-assimilation pathways in both strains. Conclusions These results illustrate the capacity of both strains to act together in performing sustainable denitrifying activities under marine conditions. Although strain JAM1 did not contribute in better specific denitrifying activities in the biofilm co-cultures, its presence helped strain NL23 to acclimate to medium with NaCl concentrations >1.0%

Denitrifying metabolism of the methylotrophic marine bacterium Methylophaga nitratireducenticrescens strain JAM1

Background Methylophaga nitratireducenticrescens strain JAM1 is a methylotrophic, marine bacterium that was isolated from a denitrification reactor treating a closed-circuit seawater aquarium. It can sustain growth under anoxic conditions by reducing nitrate ( ${\mathrm{NO}}_{3}^{-}$ NO 3 − ) to nitrite ( ${\mathrm{NO}}_{2}^{-}$ NO 2 − ). These physiological traits are attributed to gene clusters that encode two dissimilatory nitrate reductases (Nar). Strain JAM1 also contains gene clusters encoding two nitric oxide (NO) reductases and one nitrous oxide (N2O) reductase, suggesting that NO and N2O can be reduced by strain JAM1. Here we characterized further the denitrifying activities of M. nitratireducenticrescens JAM1. Methods Series of oxic and anoxic cultures of strain JAM1 were performed with N2O, ${\mathrm{NO}}_{3}^{-}$ NO 3 − or sodium nitroprusside, and growth and N2O, ${\mathrm{NO}}_{3}^{-}$ NO 3 − , ${\mathrm{NO}}_{2}^{-}$ NO 2 − and N2 concentrations were measured. Ammonium ( ${\mathrm{NH}}_{4}^{+}$ NH 4 + )-free cultures were also tested to assess the dynamics of N2O, ${\mathrm{NO}}_{3}^{-}$ NO 3 − and ${\mathrm{NO}}_{2}^{-}$ NO 2 − . Isotopic labeling of N2O was performed in 15NH4+-amended cultures. Cultures with the JAM1ΔnarG1narG2 double mutant were performed to assess the involvement of the Nar systems on N2O production. Finally, RT-qPCR was used to measure the gene expression levels of the denitrification genes cytochrome bc-type nitric oxide reductase (cnorB1 and cnorB2) and nitrous oxide reductase (nosZ), and also nnrS and norR that encode NO-sensitive regulators. Results Strain JAM1 can reduce NO to N2O and N2O to N2 and can sustain growth under anoxic conditions by reducing N2O as the sole electron acceptor. Although strain JAM1 lacks a gene encoding a dissimilatory ${\mathrm{NO}}_{2}^{-}$ NO 2 − reductase, ${\mathrm{NO}}_{3}^{-}$ NO 3 − -amended cultures produce N2O, representing up to 6% of the N-input. ${\mathrm{NO}}_{2}^{-}$ NO 2 − was shown to be the key intermediate of this production process. Upregulation in the expression of cnorB1, cnorB2, nnrS and norR during the growth and the N2O accumulation phases suggests NO production in strain JAM1 cultures. Discussion By showing that all the three denitrification reductases are active, this demonstrates that M. nitratireducenticrescens JAM1 is one of many bacteria species that maintain genes associated primarily with denitrification, but not necessarily related to the maintenance of the entire pathway. The reason to maintain such an incomplete pathway could be related to the specific role of strain JAM1 in the denitrifying biofilm of the denitrification reactor from which it originates. The production of N2O in strain JAM1 did not involve Nar, contrary to what was demonstrated in Escherichia coli. M. nitratireducenticrescens JAM1 is the only reported Methylophaga species that has the capacity to grow under anoxic conditions by using ${\mathrm{NO}}_{3}^{-}$ NO 3 − and N2O as sole electron acceptors for its growth. It is also one of a few marine methylotrophs that is studied at the physiological and genetic levels in relation to its capacity to perform denitrifying activities

Methylophaga nitratireducenticrescens sp. nov. and Methylophaga frappieri sp. nov., isolated from the biofilm of the methanol-fed denitrification system treating the seawater at the Montreal Biodome.

International audienceTwo bacterial strains, designated JAM1(T) and JAM7(T), were isolated from a methanol-fed denitrification system treating seawater at the Montreal Biodome, Canada. They were affiliated within the genus Methylophaga of the Gammaproteobacteria by analysis of the 16S rRNA gene sequences. Strain JAM1(T) had the capacity to grow under denitrifying conditions by reducing nitrate into nitrite which is unique among the species of the genus Methylophaga. Major fatty acids were C16:1ω7c or ω6c, C16:0 and C18:1ω7c or ω6c. The major ubiquinone was Q8. Both strains required vitamin B12 and Na(+) ions for growth. The genomes of strains JAM1(T) and JAM7(T) have been completely sequenced and showed a DNA G+C content of 44.7 mol% and 47.8 mol%, respectively. Growth occurred at pH 6-11 and at 0.5-8% NaCl. Both genomes contained predicted ORFs encoding the key enzymes of the ribulose monophosphate pathway. Also, operons encoding two nitrate reductases (Nar), two nitric oxide reductases (Nor), one nitrous oxide reductase (Nos) and one truncated nitrite reductase (NirK) were clustered in a 67 kb chromosomal region in strain JAM1(T). No such operons were found in strain JAM7(T). These results supported the affiliation of the two strains as novel species within the genus Methylophaga. The names Methylophaga nitratireducenticrescens sp. nov. for type strain JAM1(T) (=DSM 25689(T)=ATCC BAA-2433(T)) and Methylophaga frappieri sp. nov. for type strain JAM7(T) (=DSM 25690(T)=ATCC BAA-2434(T)) are proposed

Complete Genome Sequence of Hyphomicrobium nitrativorans Strain NL23, a Denitrifying Bacterium Isolated from Biofilm of a Methanol-Fed Denitrification System Treating Seawater at the Montreal Biodome.

International audienceHyphomicrobium nitrativorans strain NL23 has been isolated from the biofilm of a denitrification system treating seawater. This strain has the capacity to denitrify using methanol as a carbon source. Here, we report the complete genome sequence of this strain in an effort to increase understanding of the function of this bacterium within the biofilm

An environmental survey of surface waters using mitochondrial DNA from human, bovine and porcine origin as fecal source tracking markers.

International audienceFecal contamination of surface waters is one the major sources of waterborne pathogens and consequently, is an important concern for public health. For reliable fecal source tracking (FST) monitoring, there is a need for a multi-marker toolbox as no single all-encompassing method currently exists. Mitochondrial DNA (mtDNA) as a source tracking marker has emerged as a promising animal-specific marker. However, very few comprehensive field studies were done on the occurrence of this marker in surface waters. In this report, water samples were obtained from 82 sites in different watersheds over a six year period. The samples were analyzed for the presence of human, bovine and porcine mtDNA by endpoint nested PCR, along with the human-specific Bacteroidales HF183 marker. These sites represented a mix of areas with different anthropogenic activities, natural, urban and agricultural. The occurrences of mitoHu (human), mitoBo (bovine), mitoPo (porcine) and HF183 specific PCR amplifications from the samples were 46%, 23%, 6% and 50%, respectively. The occurrence of mitoHu and HF183 was high in all environment types with higher occurrence in the natural and urban areas, whereas the occurrence of mitoBo was higher in agricultural areas. FST marker concentrations were measured by real-time PCR for samples positive for these markers. The concentration of the mitoHu markers was one order of magnitude lower than HF183. There was co-linearity between the concentrations of the mitoHu and HF183 markers. Co-linearity was also observed between HF183 concentration and fecal coliform levels. Such a relationship was not observed between the mitoHu concentration and fecal coliform levels. In summary, our results showed a high incidence of human fecal pollution throughout the environment while demonstrating the potential of mtDNA as suitable FST markers