Integrating short- and full-length 16S rRNA gene sequencing to elucidate microbiome profiles in Pacific white shrimp (Litopenaeus vannamei) ponds.

Despite their immense economic value as a key aquaculture species, the production of Pacific white shrimp (Litopenaeus vannamei) faces significant challenges from intensive farming practices and disease outbreaks. Routine microbial profiling for disease surveillance could be a promising approach to anticipate and control disease outbreaks. To achieve this, accuracy in microbial profiling in shrimp ponds is crucial for enabling targeted action and prevention. Extensive documentation emphasizes that, beyond biological factors (related to the host, diet, or health status during the rearing period), technical elements, including sequencing techniques significantly influence bacterial community profiling. This study investigated the influence of short- and long-read sequencing of 16S rRNA genes on the microbial profiles in shrimp intestines, water, and sediments. The origin of the samples (intestine or environmental) in shrimp culture ponds primarily drove the observed differences in core microbial species. The ecological niches accounted for 56% of bacterial community variations in culture ponds. Both sequencing approaches showed consistent results in identifying higher-rank taxa and assessing alpha and beta diversity. However, at the species level, full-length 16S rRNA gene sequences provided better resolution than V3-V4 sequences. For routine microbial profiling in shrimp culture ponds, our study suggests that short-read sequences were sufficient for determining overall bacterial community.IMPORTANCEThis interdisciplinary study investigated the influence of sequencing techniques on bacterial communities profiling within Pacific white shrimp (Litopenaeus vannamei) ponds. By integrating aquaculture, microbiology, and environmental science, we revealed the role of ecological niches and factors like salinity and pH on microbiota diversity and composition in shrimp intestines, pond water, and sediment. Additionally, we compared the taxonomic resolution using partial versus full-length 16S rRNA gene sequences, highlighting the value of longer amplicons for precise identification of key taxa. These findings provide novel insights into microbial dynamics underlying environmental effects in shrimp aquaculture. Comprehensive characterization of the pond microbiome could lead to management strategies that promote shrimp health and productivity. Furthermore, the potential of a multi-omics approach for integrating complementary data streams to elucidate environment-microbiome-host interactions was highlighted.

Distinct gut microbiomes in Thai patients with colorectal polyps.

BACKGROUND: Colorectal polyps that develop via the conventional adenoma-carcinoma sequence [e.g., tubular adenoma (TA)] often progress to malignancy and are closely associated with changes in the composition of the gut microbiome. There is limited research concerning the microbial functions and gut microbiomes associated with colorectal polyps that arise through the serrated polyp pathway, such as hyperplastic polyps (HP). Exploration of microbiome alterations associated with HP and TA would improve the understanding of mechanisms by which specific microbes and their metabolic pathways contribute to colorectal carcinogenesis. AIM: To investigate gut microbiome signatures, microbial associations, and microbial functions in HP and TA patients. METHODS: Full-length 16S rRNA sequencing was used to characterize the gut microbiome in stool samples from control participants without polyps [control group (CT), n = 40], patients with HP (n = 52), and patients with TA (n = 60). Significant differences in gut microbiome composition and functional mechanisms were identified between the CT group and patients with HP or TA. Analytical techniques in this study included differential abundance analysis, co-occurrence network analysis, and differential pathway analysis. RESULTS: Colorectal cancer (CRC)-associated bacteria, including Streptococcus gallolyticus (S. gallolyticus), Bacteroides fragilis, and Clostridium symbiosum, were identified as characteristic microbial species in TA patients. Mediterraneibacter gnavus, associated with dysbiosis and gastrointestinal diseases, was significantly differentially abundant in the HP and TA groups. Functional pathway analysis revealed that HP patients exhibited enrichment in the sulfur oxidation pathway exclusively, whereas TA patients showed dominance in pathways related to secondary metabolite biosynthesis (e.g., mevalonate); S. gallolyticus was a major contributor. Co-occurrence network and dynamic network analyses revealed co-occurrence of dysbiosis-associated bacteria in HP patients, whereas TA patients exhibited co-occurrence of CRC-associated bacteria. Furthermore, the co-occurrence of SCFA-producing bacteria was lower in TA patients than HP patients. CONCLUSION: This study revealed distinct gut microbiome signatures associated with pathways of colorectal polyp development, providing insights concerning the roles of microbial species, functional pathways, and microbial interactions in colorectal carcinogenesis.

Probiotic Limosilactobacillus reuteri KUB-AC5 decreases urothelial cell invasion and enhances macrophage killing of uropathogenic Escherichia coli in vitro study.

Introduction: Bacterial urinary tract infections (UTI) are among the most common infectious diseases worldwide. The rise of multidrug-resistant (MDR) uropathogenic Escherichia coli (UPEC) UTI cases is a significant threat to healthcare systems. Several probiotic bacteria have been proposed as an alternative to combat MDR UTI. Lactic acid bacteria in the genus Limosilactobacillus are some of the most studied and used probiotics. However, strain-specific effects play a critical role in probiotic properties. L. reuteri KUB-AC5 (AC5), isolated from the chicken gut, confers antimicrobial and immunobiotic effects against some human pathogens. However, the antibacterial and immune modulatory effects of AC5 on UPEC have never been explored. Methods: Here, we investigated both the direct and indirect effects of AC5 against UPEC isolates (UTI89, CFT073, and clinical MDR UPEC AT31) in vitro. Using a spot-on lawn, agar-well diffusion, and competitive growth assays, we found that viable AC5 cells and cell-free components of this probiotic significantly reduced the UPEC growth of all strains tested. The human bladder epithelial cell line UM-UC-3 was used to assess the adhesion and pathogen-attachment inhibition properties of AC5 on UPEC. Results and discussion: Our data showed that AC5 can attach to UM-UC-3 and decrease UPEC attachment in a dose-dependent manner. Pretreatment of UPEC-infected murine macrophage RAW264.7 cells with viable AC5 (multiplicity of infection, MOI = 1) for 24 hours enhanced macrophage-killing activity and increased proinflammatory (Nos2, Il6, and Tnfa) and anti-inflammatory (Il10) gene expression. These findings indicate the gut-derived AC5 probiotic could be a potential urogenital probiotic against MDR UTI.

Distinct Gut Microbial Signature and Host Genetic Variants in Association with Liver Fibrosis Severity in Patients with MASLD.

Gut microbiota might affect the severity and progression of metabolic dysfunction-associated steatotic liver disease (MASLD). We aimed to characterize gut dysbiosis and clinical parameters regarding fibrosis stages assessed by magnetic resonance elastography. This study included 156 patients with MASLD, stratified into no/mild fibrosis (F0-F1) and moderate/severe fibrosis (F2-F4). Fecal specimens were sequenced targeting the V4 region of the 16S rRNA gene and analyzed using bioinformatics. The genotyping of PNPLA3, TM6SF2, and HSD17B13 was assessed by allelic discrimination assays. Our data showed that gut microbial profiles between groups significantly differed in beta-diversity but not in alpha-diversity indices. Enriched Fusobacterium and Escherichia_Shigella, and depleted Lachnospira were found in the F2-F4 group versus the F0-F1 group. Compared to F0-F1, the F2-F4 group had elevated plasma surrogate markers of gut epithelial permeability and bacterial translocation. The bacterial genera, PNPLA3 polymorphisms, old age, and diabetes were independently associated with advanced fibrosis in multivariable analyses. Using the Random Forest classifier, the gut microbial signature of three genera could differentiate the groups with high diagnostic accuracy (AUC of 0.93). These results indicated that the imbalance of enriched pathogenic genera and decreased beneficial bacteria, in association with several clinical and genetic factors, were potential contributors to the pathogenesis and progression of MASLD.

Draft genome sequence of Methylobacterium sp. 37f isolated from forest soil.

In this study, we report the draft genome sequence data of Methylobacterium sp. 37f, isolated from soil beneath Quercus semiserrata Roxb. in Thailand. The genome consists of 5,305,449 base pairs, with a GC content of 67.5%.

Gut Microbiota in Patients with Non-Alcoholic Fatty Liver Disease without Type 2 Diabetes: Stratified by Body Mass Index.

The relationship between gut dysbiosis and body mass index (BMI) in non-diabetic patients with non-alcoholic fatty liver disease (NAFLD) is not adequately characterized. This study aimed to assess gut microbiota's signature in non-diabetic individuals with NAFLD stratified by BMI. The 16S ribosomal RNA sequencing was performed for gut microbiota composition in 100 patients with NAFLD and 16 healthy individuals. The differential abundance of bacterial composition between groups was analyzed using the DESeq2 method. The alpha diversity (Chao1, Shannon, and observed feature) and beta diversity of gut microbiota significantly differed between patients with NAFLD and healthy controls. However, significant differences in their diversities were not observed among subgroups of NAFLD. At the phylum level, there was no trend of an elevated Firmicutes/Bacteroidetes ratio according to BMI. At the genus level, patients with lean NAFLD displayed a significant enrichment of Escherichia-Shigella and the depletion of Lachnospira and Subdoligranulum compared to the non-lean subgroups. Combining these bacterial genera could discriminate lean from non-lean NAFLD with high diagnostic accuracy (AUC of 0.82). Non-diabetic patients with lean NAFLD had a significant difference in bacterial composition compared to non-lean individuals. Our results might provide evidence of gut microbiota signatures associated with the pathophysiology and potential targeting therapy in patients with lean NAFLD.

Hydrolysis of ionic liquid-treated substrate with an Iocasia fonsfrigidae strain SP3-1 endoglucanase.

Recently, we reported the discovery of a novel endoglucanase of the glycoside hydrolase family 12 (GH12), designated IfCelS12A, from the haloalkaliphilic anaerobic bacterium Iocasia fonsfrigidae strain SP3-1, which was isolated from a hypersaline pond in the Samut Sakhon province of Thailand (ca. 2017). IfCelS12A exhibits high substrate specificity on carboxymethyl cellulose and amorphous cellulose but low substrate specificity on b-1,3;1,4-glucan. Unlike some endoglucanases of the GH12 family, IfCelS12A does not exhibit hydrolytic activity on crystalline cellulose (i.e., Avicel™). High-Pressure Liquid Chromatography (HPLC) and Thin Layer Chromatography (TLC) analyses of products resulting from IfCelS12-mediated hydrolysis indicate mode of action for this enzyme. Notably, IfCelS12A preferentially hydrolyzes cellotetraoses, cellopentaoses, and cellohexaoses with negligible activity on cellobiose or cellotriose. Kinetic analysis with cellopentaose and barely b-D-glucan as cellulosic substrates were conducted. On cellopentaose, IfCelS12A demonstrates a 16-fold increase in activity (KM = 0.27 mM; kcat = 0.36 s-1; kcat/KM = 1.34 mM-1 s-1) compared to the enzymatic hydrolysis of barley b-D-glucan (KM: 0.04 mM, kcat: 0.51 s-1, kcat/KM = 0.08 mM-1 s-1). Moreover, IfCelS12A enzymatic efficacy is stable in hypersaline sodium chlorids (NaCl) solutions (up to 10% NaCl). Specifically, IfCel12A retains notable activity after 24 h at 2M NaCl (10% saline solution). IfCelS12A used as a cocktail component with other cellulolytic enzymes and in conjunction with mobile sequestration platform technology offers additional options for deconstruction of ionic liquid-pretreated cellulosic feedstock. KEY POINTS: • IfCelS12A from an anaerobic alkaliphile Iocasia fronsfrigidae shows salt tolerance • IfCelS12A in cocktails with other enzymes efficiently degrades cellulosic biomass • IfCelS12A used with mobile enzyme sequestration platforms enhances hydrolysis.

Discovery of a Novel Cellobiose Dehydrogenase from Cellulomonas palmilytica EW123 and Its Sugar Acids Production.

Cellobiose dehydrogenases (CDHs) are a group of enzymes belonging to the hemoflavoenzyme group, which are mostly found in fungi. They play an important role in the production of acid sugar. In this research, CDH annotated from the actinobacterium Cellulomonas palmilytica EW123 (CpCDH) was cloned and characterized. The CpCDH exhibited a domain architecture resembling class-I CDH found in Basidiomycota. The cytochrome c and flavin-containing dehydrogenase domains in CpCDH showed an extra-long evolutionary distance compared to fungal CDH. The amino acid sequence of CpCDH revealed conservative catalytic amino acids and a distinct flavin adenine dinucleotide region specific to CDH, setting it apart from closely related sequences. The physicochemical properties of CpCDH displayed optimal pH conditions similar to those of CDHs but differed in terms of optimal temperature. The CpCDH displayed excellent enzymatic activity at low temperatures (below 30°C), unlike other CDHs. Moreover, CpCDH showed the highest substrate specificity for disaccharides such as cellobiose and lactose, which contain a glucose molecule at the non-reducing end. The catalytic efficiency of CpCDH for cellobiose and lactose were 2.05 x 105 and 9.06 x 104 (M-1 s-1), respectively. The result from the Fourier-transform infrared spectroscopy (FT-IR) spectra confirmed the presence of cellobionic and lactobionic acids as the oxidative products of CpCDH. This study establishes CpCDH as a novel and attractive bacterial CDH, representing the first report of its kind in the Cellulomonas genus.

Soil microbiome analysis reveals effects of periodic waterlogging stress on sugarcane growth.

Sugarcane is one of the major agricultural crops with high economic importance in Thailand. Periodic waterlogging has a long-term negative effect on sugarcane development, soil properties, and microbial diversity, impacting overall sugarcane production. Yet, the microbial structure in periodically waterlogged sugarcane fields across soil compartments and growth stages in Thailand has not been documented. This study investigated soil and rhizosphere microbial communities in a periodic waterlogged field in comparison with a normal field in a sugarcane plantation in Ratchaburi, Thailand, using 16S rRNA and ITS amplicon sequencing. Alpha diversity analysis revealed comparable values in periodic waterlogged and normal fields across all growth stages, while beta diversity analysis highlighted distinct microbial community profiles in both fields throughout the growth stages. In the periodic waterlogged field, the relative abundance of Chloroflexi, Actinobacteria, and Basidiomycota increased, while Acidobacteria and Ascomycota decreased. Beneficial microbes such as Arthrobacter, Azoarcus, Bacillus, Paenibacillus, Pseudomonas, and Streptomyces thrived in the normal field, potentially serving as biomarkers for favorable soil conditions. Conversely, phytopathogens and growth-inhibiting bacteria were prevalent in the periodic waterlogged field, indicating unfavorable conditions. The co-occurrence network in rhizosphere of the normal field had the highest complexity, implying increased sharing of resources among microorganisms and enhanced soil biological fertility. Altogether, this study demonstrated that the periodic waterlogged field had a long-term negative effect on the soil microbial community which is a key determining factor of sugarcane growth.

Influence of Prolonged Whole Egg Supplementation on Insulin-like Growth Factor 1 and Short-Chain Fatty Acids Product: Implications for Human Health and Gut Microbiota.

The gut microbiota exert a profound influence on human health and metabolism, with microbial metabolites playing a pivotal role in shaping host physiology. This study investigated the impact of prolonged egg supplementation on insulin-like growth factor 1 (IGF-1) and circulating short-chain fatty acids (SCFAs). In a subset of a cluster-randomized trial, participants aged 8-14 years were randomly assigned into three groups: (1) Whole Egg (WE)-consuming 10 additional eggs per week [n = 24], (2) Protein Substitute (PS)-consuming yolk-free egg substitute equivalent to 10 eggs per week [n = 25], and (3) Control Group (C) [n = 26]. At week 35, IGF-1 levels in WE significantly increased (66.6 ± 27.7 ng/mL, p < 0.05) compared to C, with positive SCFA correlations, except acetate. Acetate was stable in WE, increasing in PS and C. Significant propionate differences occurred between WE and PS (14.8 ± 5.6 µmol/L, p = 0.010). WE exhibited notable changes in the relative abundance of the Bifidobacterium and Prevotella genera. Strong positive SCFA correlations were observed with MAT-CR-H4-C10 and Libanicoccus, while Roseburia, Terrisporobacter, Clostridia_UCG-014, and Coprococcus showed negative correlations. In conclusion, whole egg supplementation improves growth factors that may be related to bone formation and growth; it may also promote benefits to gut microbiota but may not affect SCFAs.

Microbiome analysis of thai traditional fermented soybeans reveals short-chain fatty acid-associated bacterial taxa.

Thua Nao is a Thai traditional fermented soybean food and low-cost protein supplement. This study aimed to evaluate the bacterial community in Thua Nao from northern Thailand and assess potentially active short-chain fatty acids (SCFAs)-related bacteria. Sixty-five Thua Nao consisting of 30 wet and 35 dried samples were collected from six provinces: Chiang Rai, Chiang Mai, Mae Hong Son, Lampang, Lamphun, and Phayao. Bacterial diversity was significantly higher in the wet samples than in the dried samples. The dominant phyla were Firmicutes (92.7%), Proteobacteria (6.7%), Actinobacteriota (0.42%), and Bacteroidota (0.26%). The genus Bacillus (67%) was the most represented in all samples. Lactobacillus, Enterococcus, and Globicatella were enriched in the wet samples. Assessment of the SCFA-microbiota relationships revealed that high butyrate and propionate concentrations were associated with an increased Clostridiales abundance, and high acetate concentrations were associated with an increased Weissella abundance. Wet products contained more SCFAs, including acetate (P = 2.8e-08), propionate (P = 0.0044), butyrate (P = 0.0021), and isovalerate (P = 0.017), than the dried products. These results provide insight into SCFA-microbiota associations in Thua Nao, which may enable the development of starter cultures for SCFA-enriched Thua Nao production.

Prolonged Egg Supplement Advances Growing Child's Growth and Gut Microbiota.

Protein-energy malnutrition still impacts children's growth and development. We investigated the prolonged effects of egg supplementation on growth and microbiota in primary school children. For this study, 8-14-year-old students (51.5% F) in six rural schools in Thailand were randomly assigned into three groups: (1) whole egg (WE), consuming 10 additional eggs/week (n = 238) (n = 238); (2) protein substitute (PS), consuming yolk-free egg substitutes equivalent to 10 eggs/week (n = 200); and (3) control group (C, (n = 197)). The outcomes were measured at week 0, 14, and 35. At the baseline, 17% of the students were underweight, 18% were stunted, and 13% were wasted. At week 35, compared to the C group the weight and height difference increased significantly in the WE group (3.6 ± 23.5 kg, p < 0.001; 5.1 ± 23.2 cm, p < 0.001). No significant differences in weight or height were observed between the PS and C groups. Significant decreases in atherogenic lipoproteins were observed in the WE, but not in PS group. HDL-cholesterol tended to increase in the WE group (0.02 ± 0.59 mmol/L, ns). The bacterial diversity was similar among the groups. The relative abundance of Bifidobacterium increased by 1.28-fold in the WE group compared to the baseline and differential abundance analysis which indicated that Lachnospira increased and Varibaculum decreased significantly. In conclusion, prolonged whole egg supplementation is an effective intervention to improve growth, nutritional biomarkers, and gut microbiota with unaltered adverse effects on blood lipoproteins.

Insights into the genome of Methylobacterium sp. NMS14P, a novel bacterium for growth promotion of maize, chili, and sugarcane.

A novel methylotrophic bacterium designated as NMS14P was isolated from the root of an organic coffee plant (Coffea arabica) in Thailand. The 16S rRNA sequence analysis revealed that this new isolate belongs to the genus Methylobacterium, and its novelty was clarified by genomic and comparative genomic analyses, in which NMS14P exhibited low levels of relatedness with other Methylobacterium-type strains. NMS14P genome consists of a 6,268,579 bp chromosome, accompanied by a 542,519 bp megaplasmid and a 66,590 bp plasmid, namely pNMS14P1 and pNMS14P2, respectively. Several genes conferring plant growth promotion are aggregated on both chromosome and plasmids, including phosphate solubilization, indole-3-acetic acid (IAA) biosynthesis, cytokinins (CKs) production, 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity, sulfur-oxidizing activity, trehalose synthesis, and urea metabolism. Furthermore, pangenome analysis showed that NMS14P possessed the highest number of strain-specific genes accounting for 1408 genes, particularly those that are essential for colonization and survival in a wide array of host environments, such as ABC transporter, chemotaxis, quorum sensing, biofilm formation, and biosynthesis of secondary metabolites. In vivo tests have supported that NMS14P significantly promoted the growth and development of maize, chili, and sugarcane. Collectively, NMS14P is proposed as a novel plant growth-promoting Methylobacterium that could potentially be applied to a broad range of host plants as Methylobacterium-based biofertilizers to reduce and ultimately substitute the use of synthetic agrochemicals for sustainable agriculture.

Characterization of the Gut Microbiota in Urban Thai Individuals Reveals Enterotype-Specific Signature.

Gut microbiota play vital roles in human health, utilizing indigestible nutrients, producing essential substances, regulating the immune system, and inhibiting pathogen growth. Gut microbial profiles are dependent on populations, geographical locations, and long-term dietary patterns resulting in individual uniqueness. Gut microbiota can be classified into enterotypes based on their patterns. Understanding gut enterotype enables us to interpret the capability in macronutrient digestion, essential substance production, and microbial co-occurrence. However, there is still no detailed characterization of gut microbiota enterotype in urban Thai people. In this study, we characterized the gut microbiota of urban Thai individuals by amplicon sequencing and classified their profiles into enterotypes, including Prevotella (EnP) and Bacteroides (EnB) enterotypes. Enterotypes were associated with lifestyle, dietary habits, bacterial diversity, differential taxa, and microbial pathways. Microbe-microbe interactions have been studied via co-occurrence networks. EnP had lower α-diversities than those in EnB. A correlation analysis revealed that the Prevotella genus, the predominant taxa of EnP, has a negative correlation with α-diversities. Microbial function enrichment analysis revealed that the biosynthesis pathways of B vitamins and fatty acids were significantly enriched in EnP and EnB, respectively. Interestingly, Ruminococcaceae, resistant starch degraders, were the hubs of both enterotypes, and strongly correlated with microbial diversity, suggesting that traditional Thai food, consisting of rice and vegetables, might be the important drivers contributing to the gut microbiota uniqueness in urban Thai individuals. Overall findings revealed the biological uniqueness of gut enterotype in urban Thai people, which will be advantageous for developing gut microbiome-based diagnostic tools.

Genomics and cellulolytic, hemicellulolytic, and amylolytic potential of Iocasia fonsfrigidae strain SP3-1 for polysaccharide degradation.

Background: Cellulolytic, hemicellulolytic, and amylolytic (CHA) enzyme-producing halophiles are understudied. The recently defined taxon Iocasia fonsfrigidae consists of one well-described anaerobic bacterial strain: NS-1T. Prior to characterization of strain NS-1T, an isolate designated Halocella sp. SP3-1 was isolated and its genome was published. Based on physiological and genetic comparisons, it was suggested that Halocella sp. SP3-1 may be another isolate of I. fronsfrigidae. Despite being geographic variants of the same species, data indicate that strain SP3-1 exhibits genetic, genomic, and physiological characteristics that distinguish it from strain NS-1T. In this study, we examine the halophilic and alkaliphilic nature of strain SP3-1 and the genetic substrates underlying phenotypic differences between strains SP3-1 and NS-1T with focus on sugar metabolism and CHA enzyme expression. Methods: Standard methods in anaerobic cell culture were used to grow strains SP3-1 as well as other comparator species. Morphological characterization was done via electron microscopy and Schaeffer-Fulton staining. Data for sequence comparisons (e.g., 16S rRNA) were retrieved via BLAST and EzBioCloud. Alignments and phylogenetic trees were generated via CLUTAL_X and neighbor joining functions in MEGA (version 11). Genomes were assembled/annotated via the Prokka annotation pipeline. Clusters of Orthologous Groups (COGs) were defined by eegNOG 4.5. DNA-DNA hybridization calculations were performed by the ANI Calculator web service. Results: Cells of strain SP3-1 are rods. SP3-1 cells grow at NaCl concentrations of 5-30% (w/v). Optimal growth occurs at 37 °C, pH 8.0, and 20% NaCl (w/v). Although phylogenetic analysis based on 16S rRNA gene indicates that strain SP3-1 belongs to the genus Iocasia with 99.58% average nucleotide sequence identity to Iocasia fonsfrigida NS-1T, strain SP3-1 is uniquely an extreme haloalkaliphile. Moreover, strain SP3-1 ferments D-glucose to acetate, butyrate, carbon dioxide, hydrogen, ethanol, and butanol and will grow on L-arabinose, D-fructose, D-galactose, D-glucose, D-mannose, D-raffinose, D-xylose, cellobiose, lactose, maltose, sucrose, starch, xylan and phosphoric acid swollen cellulose (PASC). D-rhamnose, alginate, and lignin do not serve as suitable culture substrates for strain SP3-1. Thus, the carbon utilization profile of strain SP3-1 differs from that of I. fronsfrigidae strain NS-1T. Differences between these two strains are also noted in their lipid composition. Genomic data reveal key differences between the genetic profiles of strain SP3-1 and NS-1T that likely account for differences in morphology, sugar metabolism, and CHA-enzyme potential. Important to this study, I. fonsfrigidae SP3-1 produces and extracellularly secretes CHA enzymes at different levels and composition than type strain NS-1T. The high salt tolerance and pH range of SP3-1 makes it an ideal candidate for salt and pH tolerant enzyme discovery.

Cellulomonas palmilyticum sp. nov., from earthworm soil biofertilizer with the potential to degrade oil palm empty fruit bunch.

Oil palm empty fruit bunch (OPEFB) is lignocellulosic waste from the palm oil industry in Southeast Asia. It is difficult to degrade because of its complex matrix and recalcitrant structure. To decompose OPEFB, highly efficient micro-organisms and robust enzymatic systems are required. A bacterium with high degradation ability against untreated OPEFB was isolated from earthworm soil biofertilizer and designated as strain EW123T. Cells were Gram-stain-positive, rod-shaped and catalase-positive. In tests, the strain was negative for mycelium formation, motility, nitrate reductase and urease. The 16S rRNA gene analysis of the isolate showed 98.21 % similarity to Cellulomonas uda NBRC 3747T, whereas similarity to other species was below 98 %. The genome of strain EW123T was 3 834 009 bp long, with 73.97 mol% G+C content. Polar lipid analysis of strain EW123T indicated phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol and aminophospholipid as the lipid components of the cell wall. The major cellular fatty acid was anteiso-C15 : 0 (41.26 %) and the isomer of 2,6-diaminopimelic acid (DAP) was meso-DAP. The average nucleotide identity value between the genome sequences of EW123T and C. uda NBRC 3747T was 88.6 %. In addition, the digital DNA-DNA hybridization and genome average amino acid between those strains were 36.1 and 89.68 %, respectively. The ORF number (186) of carbohydrate-active enzymes, including cellulases, xylanases, mannanase, lipase and lignin-degrading enzymes, was higher than those of related strains. These results indicate that the polyphasic characteristics of EW123T differ from those of other related species in the genus Cellulomonas. We therefore propose a novel species of the genus Cellulomonas, namely Cellulomonas palmilyticum sp. nov. (type strain TBRC 11805T=NBRC 114552T), with the ability to effectively degrade untreated OPEFB.

Genome Characterisation of an Isoprene-Degrading Alcaligenes sp. Isolated from a Tropical Restored Forest.

Isoprene is a climate-active biogenic volatile organic compound (BVOC), emitted into the atmosphere in abundance, mainly from terrestrial plants. Soil is an important sink for isoprene due to its consumption by microbes. In this study, we report the ability of a soil bacterium to degrade isoprene. Strain 13f was isolated from soil beneath wild Himalayan cherry trees in a tropical restored forest. Based on phylogenomic analysis and an Average Nucleotide Identity score of >95%, it most probably belongs to the species Alcaligenes faecalis. Isoprene degradation by Alcaligenes sp. strain 13f was measured by using gas chromatography. When isoprene was supplied as the sole carbon and energy source at the concentration of 7.2 × 105 ppbv and 7.2 × 106 ppbv, 32.6% and 19.6% of isoprene was consumed after 18 days, respectively. Genome analysis of Alcaligenes sp. strain 13f revealed that the genes that are typically found as part of the isoprene monooxygenase gene cluster in other isoprene-degrading bacteria were absent. This discovery suggests that there may be alternative pathways for isoprene metabolism.

Complete Genome Sequence of Neisseria gonorrhoeae Multilocus Sequence Type ST7363 Isolated from Thailand.

A Neisseria gonorrhoeae multilocus sequence type (MLST) ST7363 strain was isolated from a patient at the Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand, in 2010 and completely sequenced. This strain is susceptible to ceftriaxone and cefixime. A complete circular chromosome and circular plasmids were assembled from combined Oxford Nanopore Technologies (ONT) and Illumina sequencing.

A novel amylolytic/xylanolytic/cellulolytic multienzyme complex from Clostridium manihotivorum that hydrolyzes polysaccharides in cassava pulp.

Some anaerobic bacteria, particularly Clostridium species, produce extracellular cellulolytic and xylanolytic enzymes as multienzyme complexes (MECs). However, an amylolytic/xylanolytic/cellulolytic multienzyme complex (AXC-MEC) from anaerobic bacteria is rarely found. In this work, the glycoprotein AXC-MEC, composed of subunits of amylolytic, xylanolytic, and cellulolytic enzymes, was isolated from crude extracellular enzyme of the mesophilic anaerobic bacterium Clostridium manihotivorum CT4, grown on cassava pulp, using a milled cassava pulp column and Sephacryl S-500 gel filtration chromatography. The isolated AXC-MEC showed a single band upon native-polyacrylamide gel electrophoresis (native-PAGE). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed at least eight protein bands of the multienzyme complex which predominantly exhibited amylolytic enzyme activity, followed by xylanolytic and cellulolytic enzyme activities. The AXC-MEC is highly capable of degrading starch and non-starch polysaccharides present in cassava pulp into glucose and oligosaccharides, without conventional pretreatment. Base on the genomic analysis of C. manihotivorum CT4, we found no evidence of the known structural components of the well-known multienzyme complexes from Clostridium species, cellulosomes such as scaffoldin, cohesin, and dockerin, indicating that AXC-MEC from strain CT4 exhibit a different manner of assembly from the cellulosomes. These results suggest that AXC-MEC from C. manihotivorum CT4 is a new MEC capable of hydrolyzing cassava pulp into value-added products, which will benefit the starch industry. KEY POINTS: • Glycoprotein AXC-MEC was first reported in Clostridium manihotivorum. • Unlike cellulosomes, AXC-MEC consists of amylase, xylanase, and cellulase. • Glucose and oligosaccharides were hydrolysis products from cassava pulp by AXC-MEC.