Genomic Insights into Denitrifying Methane-Oxidizing Bacteria Gemmobacter fulva sp. Nov., Isolated from an Anabaena Culture

The genus Gemmobacter grows phototrophically, aerobically, or anaerobically, and utilizes methylated amine. Here, we present two high-quality complete genomes of the strains con4 and con5T isolated from a culture of Anabaena. The strains possess sMMO (soluble methane monooxygenase)-oxidizing alkanes to carbon dioxide. Functional genes for methane-oxidation (prmAC, mimBD, adh, gfa, fdh) were identified. The genome of strain con5T contains nirB, nirK, nirQ, norB, norC, and norG genes involved in dissimilatory nitrate reduction. The presence of nitrite reductase gene (nirK) and the nitric-oxide reductase gene (norB) indicates that it could potentially use nitrite as an electron acceptor in anoxic environments. Taxonomic investigations were also performed on two strains through polyphasic methods, proposing two isolates as a novel species of the genus Gemmobacter. The findings obtained through the whole genome analyses provide genome-based evidence of complete oxidation of methane to carbon dioxide. This study provides a genetic blueprint of Gemmobacter fulva con5T and its biochemical characteristics, which help us to understand the evolutionary biology of the genus Gemmobacter.

Enhancing Denitrification in Constructed Wetland with Algae Addition

Constructed wetlands (CWs) can be used for tertiary treatment of wastewater; however, carbon source shortages limit denitrification. We studied the effect of algae addition as an external carbon source in CWs and found that the nitrogen removal efficiency of CWs is highly dependent on the algae dosage. Optimal nitrogen removal can be achieved by adding 80 mg·L− 1 dry weight algae to the influent when the chemical oxygen demand/nitrogen (COD/N) ratio reaches 5.3. Longitudinal changes in the nitrogen concentrations, organic matter concentrations, and nitrogen functional genes were also analyzed. The algae addition strengthened the anoxic environment, boosted the volatile fatty acid concentrations, and proliferated the nitrite reductase gene (nirS) and the nitrite oxidoreductase alpha subunit gene (nxrA), thereby expanding the active space for denitrification. The integration of algal ponds with CWs could potentially provide enough carbon to enhance denitrification during treatment of wastewater with a low COD/N ratio.

The Pseudomonas aeruginosa phosphodiesterase gene nbdA is transcriptionally regulated by RpoS and AmrZ

Pseudomonas aeruginosa is an opportunistic pathogen causing serious infections in immune compromised persons. These infections are difficult to erase with antibiotics, due to the formation of biofilms. The biofilm lifecycle is regulated by the second messenger molecule c-di-GMP (bis-3,5-cyclic di-guanosine monophosphate). P. aeruginosa encodes 40 genes for enzymes presumably involved in the biosynthesis and degradation of c-di-GMP. A tight regulation of expression, subcellular localized function and protein interactions control the activity of these enzymes. In this work we elucidated the transcriptional regulation of the gene encoding the membrane-bound phosphodiesterase NbdA. We previously reported a transcriptional and posttranslational role of nitric oxide (NO) on nbdA and its involvement in biofilm dispersal. NO is released from macrophages during infections but can also be produced by P. aeruginosa itself during anaerobic denitrification. Recently however, contradictory results about the role of NbdA within NO-induced biofilm dispersal were published. Therefore, the transcriptional regulation of nbdA was reevaluated to obtain insights into this discrepancy. Determination of the transcriptional start site of nbdA by 5'-RACE and subsequent identification of the promoter region revealed a shortened open reading frame (ORF) in contrast to the annotated one. In addition, putative binding sites for RpoS and AmrZ were discovered in the newly defined promoter region. Employing chromosomally integrated transcriptional lacZ reporter gene fusions demonstrated a RpoS-dependent activation and AmrZ repression of nbdA transcription. In order to investigate the impact of NO on nbdA transcription, conditions mimicking exogenous and endogenous NO were applied. While neither exogenous nor endogenous NO had an influence on nbdA promoter activity, deletion of the nitrite reductase gene nirS strongly increased nbdA transcription independently of its enzymatic activity during denitrification. The latter supports a role of NirS in P. aeruginosa apart from its enzymatic function.

Isolation and Characterization of an Aerobic Denitrifier Bacillus sp. SC16 from an Intensive Aquaculture Pond

Overloading of ammonia and nitrite nitrogen in aquaculture can result in toxicity to aquatic animals. In order to eliminate the hazardous substances, a highly efficient denitrifying bacterium, Bacillus sp. SC16, was identified in a fishery pond and isolated subsequently. The strain SC16 could remove nitrate up to 97%, ammonia up to 36.6%, and nitrite up to 99.99% when incubated with nitrate at an initial concentration of 306.9 mg·L−1 for 72 h, ammonia at 165.49 mg·L−1 for 48 h, and nitrite at 200 mg·L−1 for 24 h under aerobic conditions. The nitrite reductase gene was identified as the nirK gene. The transcriptional levels of the nirK gene in strain SC16 incubated with ammonia, nitrate, and nitrite showed similar expression patterns. When the strain SC16 was used to treat the aquaculture water, the concentration of ammonia decreased significantly, from 8.35 mg·L−1 to 4.56 mg·L−1, and there was almost no accumulation of nitrite by the end of experiment. Therefore, the results indicated that Bacillus sp. SC16 could be a promising candidate for aquaculture water treatment.

Effects of Oenanthe javanica on Nitrogen Removal in Free-Water Surface Constructed Wetlands under Low-Temperature Conditions

To investigate the role and microorganism-related mechanisms of macrophytes and assess the feasibility of Oenanthe javanica (Blume) DC. in promoting nitrogen removal in free-water surface constructed wetlands (FWS-CWS) under low temperatures (<10 °C), pilot-scale FWS-CWS, planted with O. javanica, were set up and run for batch wastewater treatment in eastern China during winter. The presence of macrophytes observably improved the removal rates of ammonia nitrogen (65%–71%) and total nitrogen (41%–48%) (p < 0.05), with a sharp increase in chemical oxygen demand concentrations (about 3–4 times). Compared to the unplanted systems, the planted systems not only exhibited higher richness and diversity of microorganisms, but also significantly higher abundances of bacteria, ammonia monooxygenase gene (amoA), nitrous oxide reductase gene (nosZ), dissimilatory cd1-containing nitrite reductase gene (nirS), and dissimilatory copper-containing nitrite reductase gene (nirK) in the substrate. Meanwhile, the analysis of the microbial community composition further revealed significant differences. The results indicate that enhanced abundances of microorganisms, and the key functional genes involved with nitrogen metabolism in the planted systems played critical roles in nitrogen removal from wastewater in FWS-CWS. Furthermore, abundant carbon release from the wetland macrophytes could potentially aid nitrogen removal in FWS-CWS during winter.

Potentially Mobile Denitrification Genes Identified inAzospirillumsp. Strain TSH58

ABSTRACTDenitrification ability is sporadically distributed among diverse bacteria, archaea, and fungi. In addition, disagreement has been found between denitrification gene phylogenies and the 16S rRNA gene phylogeny. These facts have suggested potential occurrences of horizontal gene transfer (HGT) for the denitrification genes. However, evidence of HGT has not been clearly presented thus far. In this study, we identified the sequences and the localization of the nitrite reductase genes in the genomes of 41 denitrifyingAzospirillumsp. strains and searched for mobile genetic elements that contain denitrification genes. AllAzospirillumsp. strains examined in this study possessed multiple replicons (4 to 11 replicons), with their sizes ranging from 7 to 1,031 kbp. Among those, the nitrite reductase genenirKwas located on large replicons (549 to 941 kbp). Genome sequencing showed thatAzospirillumstrains that had similarnirKsequences also shared similarnir-norgene arrangements, especially between the TSH58, Sp7T, and Sp245 strains. In addition to the high similarity betweennir-norgene clusters among the threeAzospirillumstrains, a composite transposon structure was identified in the genome of strain TSH58, which contains thenir-norgene cluster and the novel IS6family insertion sequences (ISAz581and ISAz582). ThenirKgene within the composite transposon system was actively transcribed under denitrification-inducing conditions. Although not experimentally verified in this study, the composite transposon system containing thenir-norgene cluster could be transferred to other cells if it is moved to a prophage region and the phage becomes activated and released outside the cells. Taken together, strain TSH58 most likely acquired its denitrification ability by HGT from closely relatedAzospirillumsp. denitrifiers.IMPORTANCEThe evolutionary history of denitrification is complex. While the occurrence of horizontal gene transfer has been suggested for denitrification genes, most studies report circumstantial evidences, such as disagreement between denitrification gene phylogenies and the 16S rRNA gene phylogeny. Based on the comparative genome analyses ofAzospirillumsp. denitrifiers, we identified denitrification genes, includingnirKandnorCBQD, located on a mobile genetic element in the genome ofAzospirillumsp. strain TSH58. ThenirKwas actively transcribed under denitrification-inducing conditions. Since this gene was the sole nitrite reductase gene in strain TSH58, this strain most likely benefitted by acquiring denitrification genes via horizontal gene transfer. This finding will significantly advance our scientific knowledge regarding the ecology and evolution of denitrification.

Genomic Characterization of Urethritis-Associated Neisseria meningitidis Shows that a Wide Range of N. meningitidis Strains Can Cause Urethritis

ABSTRACTNeisseria meningitidis, typically a resident of the oro- or nasopharynx and the causative agent of meningococcal meningitis and meningococcemia, is capable of invading and colonizing the urogenital tract. This can result in urethritis, akin to the syndrome caused by its sister species,N. gonorrhoeae, the etiologic agent of gonorrhea. Recently, meningococcal strains associated with outbreaks of urethritis were reported to share genetic characteristics with the gonococcus, raising the question of the extent to which these strains contain features that promote adaptation to the genitourinary niche, making them gonococcus-like and distinguishing them from otherN. meningitidisstrains. Here, we analyzed the genomes of 39 diverseN. meningitidisisolates associated with urethritis, collected independently over a decade and across three continents. In particular, we characterized the diversity of the nitrite reductase gene (aniA), the factor H-binding protein gene (fHbp), and the capsule biosynthetic locus, all of which are loci previously suggested to be associated with urogenital colonization. We observed notable diversity, including frameshift variants, inaniAandfHbpand the presence of intact, disrupted, and absent capsule biosynthetic genes, indicating that urogenital colonization and urethritis caused byN. meningitidisare possible across a range of meningococcal genotypes. Previously identified allelic patterns in urethritis-associatedN. meningitidisstrains may reflect genetic diversity in the underlying meningococcal population rather than novel adaptation to the urogenital tract.