Chun Shiong Chong

Recent discoveries and applications of Anoxybacillus

The Bacillaceae family members are a good source of bacteria for bioprocessing and biotransformation involving whole cells or enzymes. In contrast to Bacillus and Geobacillus, Anoxybacillus is a relatively new genus that was proposed in the year 2000. Because these bacteria are alkali-tolerant thermophiles, they are suitable for many industrial applications. More than a decade after the first report of Anoxybacillus, knowledge accumulated from fundamental and applied studies suggests that this genus can serve as a good alternative in many applications related to starch and lignocellulosic biomasses, environmental waste treatment, enzyme technology, and possibly bioenergy production. This current review provides the first summary of past and recent discoveries regarding the isolation of Anoxybacillus, its medium requirements, its proteins that have been characterized and cloned, bioremediation applications, metabolic studies, and genomic analysis. Comparisons to some other members of Bacillaceae and possible future applications of Anoxybacillus are also discussed.

Analysis of Anoxybacillus Genomes from the Aspects of Lifestyle Adaptations, Prophage Diversity, and Carbohydrate Metabolism

Species of Anoxybacillus are widespread in geothermal springs, manure, and milk-processing plants. The genus is composed of 22 species and two subspecies, but the relationship between its lifestyle and genome is little understood. In this study, two high-quality draft genomes were generated from Anoxybacillus spp. SK3-4 and DT3-1, isolated from Malaysian hot springs. De novo assembly and annotation were performed, followed by comparative genome analysis with the complete genome of Anoxybacillus flavithermus WK1 and two additional draft genomes, of A. flavithermus TNO-09.006 and A. kamchatkensis G10. The genomes of Anoxybacillus spp. are among the smaller of the family Bacillaceae. Despite having smaller genomes, their essential genes related to lifestyle adaptations at elevated temperature, extreme pH, and protection against ultraviolet are complete. Due to the presence of various competence proteins, Anoxybacillus spp. SK3-4 and DT3-1 are able to take up foreign DNA fragments, and some of these transferred genes are important for the survival of the cells. The analysis of intact putative prophage genomes shows that they are highly diversified. Based on the genome analysis using SEED, many of the annotated sequences are involved in carbohydrate metabolism. The presence of glycosyl hydrolases among the Anoxybacillus spp. was compared, and the potential applications of these unexplored enzymes are suggested here. This is the first study that compares Anoxybacillus genomes from the aspect of lifestyle adaptations, the capacity for horizontal gene transfer, and carbohydrate metabolism.

Characterization of aluminum resistant Anoxybacillus sp. SK 3-4 isolated from a hot spring

The Anoxybacillus sp. SK 3–4, previously isolated from a hot spring, was screened for its heavy metals resistance (Al3+, Mn2+, Cu2+, Co2+, Zn2+, and Ni2+) and the strain was found to be most resistant to aluminum. Significant growth of the strain was observed when it was grown in medium containing aluminum (200 mg L−1–800 mg L−1) with relative growth rates ranging between 77% and 100%. A gene encoding the aluminum resistance protein (accession number: WP_021095658.1) was found in genome of strain SK 3–4, which revealed high sequence identity (>95%) to its homologues from Anoxybacillus species. Sequence comparisons with two functionally characterized aluminum resistance proteins, namely G2alt and ALU1‐P, showed 97% and 81% of sequence identity, respectively. Four putative metal binding sites were detected in SK 3–4 aluminum resistance protein and G2alt at same amino acid residue positions of 186, 195, 198, and 201. Strain SK 3–4 was found to be able to remove aluminum from aqueous solution. This study demonstrated that Anoxybacillus sp. SK 3–4 could be applied in the treatment of aluminum contaminated wastewater.

Global transcriptomic response of Anoxybacillus sp. SK 3-4 to aluminum exposure.

Anoxybacillus sp. SK 3‐4 is a Gram‐positive, rod‐shaped bacterium and a member of family Bacillaceae. We had previously reported that the strain is an aluminum resistant thermophilic bacterium. This is the first report to provide a detailed analysis of the global transcriptional response of Anoxybacillus when the cells were exposed to 600 mg L−1 of aluminum. The transcriptome was sequenced using Illumina MiSeq sequencer. Total of 708 genes were differentially expressed (fold change >2.00) with 316 genes were up‐regulated while 347 genes were down‐regulated, in comparing to control with no aluminum added in the culture. Based on Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, the majority of genes encoding for cell metabolism such as glycolysis, sulfur metabolism, cysteine and methionine metabolism were up‐regulated; while most of the gene associated with tricarboxylic acid cycle (TCA cycle) and valine, leucine and isoleucine metabolism were down‐regulated. In addition, a significant number of the genes encoding ABC transporters, metal ions transporters, and some stress response proteins were also differentially expressed following aluminum exposure. The findings provide further insight and help us to understand on the resistance of Anoxybacillus sp. SK 3‐4 toward aluminium.

Vitellibacter aquimaris sp. nov., a marine bacterium isolated from seawater

A Gram-staining-negative, aerobic, yellow-orange-pigmented, rod-shaped bacterium designated D-24T was isolated from seawater from sandy shoreline in Johor, Malaysia. The 16S rRNA gene sequence analysis revealed that strain D-24T is affiliated with the genus Vitellibacter. It shared more than 96 % sequence similarity with the types of some of the validly published species of the genus: Vitellibactervladivostokensis KMM 3516T (99.5 %), Vitellibactersoesokkakensis RSSK-12T (97.3 %), VitellibacterechinoideorumCC-CZW007T (96.9 %), VitellibacternionensisVBW088T (96.7 %) and Vitellibacteraestuarii JCM 15496T (96.3 %). DNA-DNA hybridization and genome-based analysis of average nucleotide identity (ANI) of strain D-24T versus V.vladivostokensisKMM 3516T exhibited values of 35.9±0.14 % and 89.26 %, respectively. Strain D-24T showed an even lower ANI value of 80.88 % with V. soesokkakensis RSSK-12T. The major menaquinone of strain D-24T was MK-6, and the predominant fatty acids were iso-C15 : 0 and iso-C17 : 0 3-OH. Strain D-24T contained major amounts of phosphatidylethanolamine, two lipids and two aminolipids, and a phosphoglycolipid that was different to that of other species of the genus Vitellibacter. The genomic DNA G+C content was 40.6 mol%. On the basis of phenotypic properties, DNA-DNA relatedness, ANI value and chemotaxonomic analyses, strain D-24T represents a novel species of the genus Vitellibacter, for which the name Vitellibacter aquimaris sp. nov. is proposed. The type strain is D-24T (=KCTC 42708T=DSM 101732T).

Biofilm-coated macrocomposites for the treatment of high strength agricultural wastewater

AbstractThe processing of palm oil generates highly polluting effluent called palm oil mill effluent (POME). Due to the inefficiency of conventional treatment methods to meet the environmental standard discharge limit, alternative treatment methods could play a positive role in treating POME. In this study, biofilm-coated macrocomposites were used to treat the POME obtained from palm oil industry. Color and chemical oxygen demand (COD) removal were the main parameters analyzed in this study. The rate of color removal was examined using the Elovich equation, Pseudo-first-order, and Pseudo-second-order kinetics models. Pseudo-first-order gave the best model equation for color adsorption of POME. The biofilm that was coated on the macrocomposites consisted of a mixed bacterial culture of Brevibacillus panacihumi, Enterococcus faecalis, Lysinibacillus fusiformis, and the newly identified Klebsiella pneumonia (MABZ). All of these bacteria were previously confirmed as bacteria capable of removing color and COD....

Draft genome sequence of Vitellibacter vladivostokensis KMM 3516T: A protease-producing bacterium

Type strain Vitellibacter vladivostokensis KMM 3516(T) (=NBRC 16718(T)) belongs to the phylum Cytophaga-Flavobacterium-Bacteroides. To date, no genomes of the Vitellibacter spp. have been reported, and their metabolic pathways are unknown. This study reports the draft genome sequence of V. vladivostokensis. Moreover, mining of genes associated with proteolytic enzymes was performed to provide insights for further enzyme characterization.

Microbial diversity of thermophiles with biomass deconstruction potential in a foliage-rich hot spring

The ability of thermophilic microorganisms and their enzymes to decompose biomass have attracted attention due to their quick reaction time, thermostability, and decreased risk of contamination. Exploitation of efficient thermostable glycoside hydrolases (GHs) could accelerate the industrialization of biofuels and biochemicals. However, the full spectrum of thermophiles and their enzymes that are important for biomass degradation at high temperatures have not yet been thoroughly studied. We examined a Malaysian Y‐shaped Sungai Klah hot spring located within a wooded area. The fallen foliage that formed a thick layer of biomass bed under the heated water of the Y‐shaped Sungai Klah hot spring was an ideal environment for the discovery and analysis of microbial biomass decay communities. We sequenced the hypervariable regions of bacterial and archaeal 16S rRNA genes using total community DNA extracted from the hot spring. Data suggested that 25 phyla, 58 classes, 110 orders, 171 families, and 328 genera inhabited this hot spring. Among the detected genera, members of Acidimicrobium, Aeropyrum, Caldilinea, Caldisphaera, Chloracidobacterium, Chloroflexus, Desulfurobacterium, Fervidobacterium, Geobacillus, Meiothermus, Melioribacter, Methanothermococcus, Methanotorris, Roseiflexus, Thermoanaerobacter, Thermoanaerobacterium, Thermoanaerobaculum, and Thermosipho were the main thermophiles containing various GHs that play an important role in cellulose and hemicellulose breakdown. Collectively, the results suggest that the microbial community in this hot spring represents a good source for isolating efficient biomass degrading thermophiles and thermozymes.

Draft genome sequence of Vitellibacter aquimaris D-24T isolated from seawater

Vitellibacter aquimaris D-24T (=KCTC 42708T = DSM 101732T), a halophilic marine bacterium, was isolated from seawater collected from Desaru beach, Malaysia. Here, we present the draft genome sequence of D-24T with a genome size of approximately 3.1 Mbp and G + C content of 39.93%. The genome of D-24T contains genes involved in reducing a potent greenhouse gas (N2O) in the environment and the degradation of proteinaceous compounds. Genome availability will provide insights into potential biotechnological and environmental applications of this bacterium.

Genome sequence of Roseivirga sp. strain D-25 and its potential applications from the genomic aspect

Roseivirga sp. strain D-25 is an aerobic marine bacterium isolated from seawater collected from Desaru beach, Malaysia. To date, the genus Roseivirga consists of only four species with no genome sequence reported. Here, we present the genome sequence of Roseivirga sp. strain D-25 (=KCTC 42709=DSM 101709), with a genome size of approximately 4.08Mbp and G+C content of 39.18%. Genome sequence analysis of strain D-25 revealed the presence of genes related to petroleum hydrocarbon degradation, 2,4,6-trinitrotoluene detoxification, heavy metals bioremediation and production of carotenoids, which shed light on the potential application of this strain.