The description of comammox Nitrospira spp., performing complete ammonia-to-nitrate oxidation, and their co-occurrence with canonical β-proteobacterial ammonia oxidizing bacteria (β-AOB) in the environment, calls into question the metabolic potential of comammox Nitrospira and the evolutionary history of their ammonia oxidation pathway. We report four new comammox Nitrospira genomes, constituting two novel species, and the first comparative genomic analysis on comammox Nitrospira. Unlike canonical Nitrospira, comammox Nitrospira genomes lack genes for assimilatory nitrite reduction, suggesting that they have lost the potential to use external nitrite nitrogen sources. By contrast, compared to canonical Nitrospira, comammox Nitrospira harbor a higher diversity of urea transporters and copper homeostasis genes and lack cyanate hydratase genes. Additionally, the two comammox clades differ in their ammonium uptake systems. Contrary to β-AOB, comammox Nitrospira genomes have single copies of the two central ammonia oxidation pathway operons. Similar to ammonia oxidizing archaea and some oligotrophic AOB strains, they lack genes involved in nitric oxide reduction. Furthermore, comammox Nitrospira genomes encode genes that might allow efficient growth at low oxygen concentrations. Regarding the evolutionary history of comammox Nitrospira, our analyses indicate that several genes belonging to the ammonia oxidation pathway could have been laterally transferred from β-AOB to comammox Nitrospira. We postulate that the absence of comammox genes in other sublineage II Nitrospira genomes is the result of subsequent loss.
Previous studies have shown that enhanced biological phosphorus removal (EBPR) performance under continuous aerobic conditions always eventually deteriorates; however, the speed at which this happens depends on the carbon source supplied. The published data suggest that propionate is a better carbon source than acetate is for maintaining operational stability, although it is not clear why. A lab-scale sequencing batch reactor was run initially under conventional anaerobic/aerobic conditions with either acetate or propionate as the carbon source. Chemical and microbiological analyses revealed that both sources performed as expected for such systems. When continuous aerobic conditions were imposed on both these established communities, marked shifts of the "Candidatus Accumulibacter" clades were recorded for both carbon sources. Here, we discuss whether this shift could explain the prolonged EBPR stability observed with propionate.
Polyhydroxyalkanoates (PHAs) are biopolyesters synthesized by microorganisms as intracellular energy reservoirs under stressful environmental conditions. PHA synthase (PhaC) is the key enzyme responsible for PHA biosynthesis, but the importance of its N- and C-terminal ends still remains elusive. Six plasmid constructs expressing truncation variants of Aquitalea sp. USM4 PhaC (PhaC1As) were generated and heterologously expressed in Cupriavidus necator PHB-4. Removal of the first six residues at the N-terminus enabled the modulation of PHA composition without altering the PHA content in cells. Meanwhile, deletion of 13 amino acids from the C-terminus greatly affected the catalytic activity of PhaC1As, retaining only 1.1-7.4% of the total activity. Truncation(s) at the N- and/or C-terminus of PhaC1As gradually diminished the incorporation of comonomer units, and revealed that the N-terminal region is essential for PhaC1As dimerization whereas the C-terminal region is required for stabilization. Notably, transmission electron microscopy analysis showed that PhaC modification affected the morphology of intracellular PHA granules, which until now is only known to be regulated by phasins. This study provided substantial evidence and highlighted the significance of both the N- and C-termini of PhaC1As in regulating intracellular granule morphology, activity, substrate specificity, dimerization and stability of the synthase.
Polyhydroxyalkanoate (PHA) synthase, PhaC, is a key enzyme in the biosynthesis of PHA, a type of bioplastics with huge potential to replace petroleum-based plastics. While two structures have been determined, the exact mechanism remains unclear partly due to the absence of a tunnel for product passage. A model of the class I PhaC from Aquitalea sp. USM4, characterised with Km of 394 μM and kcat of 476 s-1 on 3-(R)-hydroxybutyryl-CoA, revealed a three-branched channel at the dimeric interface. Two of them are opened to the solvent and are expected to serve as the putative routes for substrate entrance and product exit, while the third is elongated in the class II PhaC1 model from Pseudomonas aeruginosa, indicating a role in accommodating the hydroxyalkanoate (HA) moiety of a HA-CoA substrate. Docking of the two tetrahedral intermediates, formed during the transfer of the growing PHA chain from the catalytic Cys to a new molecule of substrate and back to Cys, suggests a common elongation mechanism requiring the HA moiety of the ligand to rotate ~180°. Substrate specificity is determined in part by a bulky Phe/Tyr/Trp residue in the third branch in class I, which is conserved as Ala in class II to create room for longer substrates.
A novel polyhydroxyalkanoate (PHA)-producing bacterium, Jeongeupia sp. USM3 (JCM 19920) was isolated from the limestone soil at Gua Tempurung, Perak, Malaysia. This is the first report on the complete genome sequence for the genus Jeongeupia. This genome consists of a circular chromosome with a size of 3,788,814 bp and contains 3557 genes. Two PHA synthase (phaC) genes encoding for the key enzyme in the polymerization of PHA monomers and other PHA-associated genes were identified from the genome. Phylogenetic analysis of the PhaC protein sequences has revealed that both PhaC1 and PhaC2 of Jeongeupia sp. USM3 are categorized as Class I PHA synthases with 56% similarity to each other. Both of the PHA synthase genes of this isolate were cloned and heterologously expressed in a PHA mutant strain Cupriavidus necator PHB-4. The ability of the transformants to accumulate PHA showed that both PhaC1 and PhaC2 were functional.
All obligate bacterial endosymbionts of free-living amoebae currently described are affiliated with the alpha-Proteobacteria, the Chlamydiales or the phylum Cytophaga-Flavobacterium-Bacteroides. Here, six rod-shaped gram-negative obligate bacterial endosymbionts of clinical and environmental isolates of Acanthamoeba spp. from the USA and Malaysia are reported. Comparative 16S rDNA sequence analysis demonstrated that these endosymbionts form a novel, monophyletic lineage within the beta-Proteobacteria, showing less than 90% sequence similarity to all other recognized members of this subclass. 23S rDNA sequence analysis of two symbionts confirmed this affiliation and revealed the presence of uncommon putative intervening sequences of 146 bp within helix-25 that shared no sequence homology to any other bacterial rDNA. In addition, the 23S rRNA of these endosymbionts displayed one polymorphism at the target site of oligonucleotide probe BET42a that is conserved in all other sequenced beta-Proteobacteria. Intra-cytoplasmatic localization of the endosymbionts within the amoebal host cells was confirmed by electron microscopy and fluorescence in situ hybridization with a specific 16S rRNA-targeted oligonucleotide probe. Based on these findings, the provisional name 'Candidatus Procabacter acanthamoebae' is proposed for classification of a representative of the six endosymbionts of Acanthamoeba spp. studied in this report. Comparative 18S rDNA sequence analysis of the Acanthamoeba host cells revealed their membership with either Acanthamoeba 18S rDNA sequence type T5 (Acanthamoeba lenticulata) or sequence type T4, which comprises the majority of all Acanthamoeba isolates.
It is known that the microbial community of the rhizosphere is not only influenced by factors such as root exudates, phenology, and nutrient uptake but also by the plant species. However, studies of bacterial communities associated with tropical rainforest tree root surfaces, or rhizoplane, are lacking. Here, we analyzed the bacterial community of root surfaces of four species of native trees, Agathis borneensis, Dipterocarpus kerrii, Dyera costulata, and Gnetum gnemon, and nearby bulk soils, in a rainforest arboretum in Malaysia, using 454 pyrosequencing of the 16S rRNA gene. The rhizoplane bacterial communities for each of the four tree species sampled clustered separately from one another on an ordination, suggesting that these assemblages are linked to chemical and biological characteristics of the host or possibly to the mycorrhizal fungi present. Bacterial communities of the rhizoplane had various similarities to surrounding bulk soils. Acidobacteria, Alphaproteobacteria, and Betaproteobacteria were dominant in rhizoplane communities and in bulk soils from the same depth (0-10 cm). In contrast, the relative abundance of certain bacterial lineages on the rhizoplane was different from that in bulk soils: Bacteroidetes and Betaproteobacteria, which are known as copiotrophs, were much more abundant in the rhizoplane in comparison to bulk soil. At the genus level, Burkholderia, Acidobacterium, Dyella, and Edaphobacter were more abundant in the rhizoplane. Burkholderia, which are known as both pathogens and mutualists of plants, were especially abundant on the rhizoplane of all tree species sampled. The Burkholderia species present included known mutualists of tropical crops and also known N fixers. The host-specific character of tropical tree rhizoplane bacterial communities may have implications for understanding nutrient cycling, recruitment, and structuring of tree species diversity in tropical forests. Such understanding may prove to be useful in both tropical forestry and conservation.
Knowledge about the biogeography of marine bacterioplankton on the global scale in general and in Southeast Asia in particular has been scarce. This study investigated the biogeography of bacterioplankton community in Singapore seawaters. Twelve stations around Singapore island were sampled on different schedules over 1 year. Using PCR-DNA fingerprinting, DNA cloning and sequencing, and microarray hybridization of the 16S rRNA genes, we observed clear spatial variations of bacterioplankton diversity within the small area of the Singapore seas. Water samples collected from the Singapore Strait (south) throughout the year were dominated by DNA sequences affiliated with Cyanobacteria and Alphaproteobacteria that were believed to be associated with the influx of water from the open seas in Southeast Asia. On the contrary, water in the relatively polluted Johor Strait (north) were dominated by Betaproteobacteria, Gammaproteobacteria, and Bacteroidetes and that were presumably associated with river discharge and the relatively eutrophic conditions of the waterway. Bacterioplankton diversity was temporally stable, except for the episodic surge of Pseudoalteromonas, associated with algal blooms. Overall, these results provide valuable insights into the diversity of bacterioplankton communities in Singapore seas and the possible influences of hydrological conditions and anthropogenic activities on the dynamics of the communities.
Tropical peat swamp forest is a global store of carbon in a water-saturated, anoxic and acidic environment. This ecosystem holds diverse prokaryotic communities that play a major role in nutrient cycling. A study was conducted in which a total of 24 peat soil samples were collected in three forest types in a tropical peat dome in Sarawak, Malaysia namely, Mixed Peat Swamp (MPS), Alan Batu (ABt), and Alan Bunga (ABg) forests to profile the soil prokaryotic communities through meta 16S amplicon analysis using Illumina Miseq. Results showed these ecosystems were dominated by anaerobes and fermenters such as Acidobacteria, Proteobacteria, Actinobacteria and Firmicutes that cover 80-90% of the total prokaryotic abundance. Overall, the microbial community composition was different amongst forest types and depths. Additionally, this study highlighted the prokaryotic communities' composition in MPS was driven by higher humification level and lower pH whereas in ABt and ABg, the less acidic condition and higher organic matter content were the main factors. It was also observed that prokaryotic diversity and abundance were higher in the more oligotrophic ABt and ABg forest despite the constantly waterlogged condition. In MPS, the methanotroph Methylovirgula ligni was found to be the major species in this forest type that utilize methane (CH4), which could potentially be the contributing factor to the low CH4 gas emissions. Aquitalea magnusonii and Paraburkholderia oxyphila, which can degrade aromatic compounds, were the major species in ABt and ABg forests respectively. This information can be advantageous for future study in understanding the underlying mechanisms of environmental-driven alterations in soil microbial communities and its potential implications on biogeochemical processes in relation to peatland management.