Alisa Wilantho

Metagenomic profiles of free-living archaea, bacteria and small eukaryotes in coastal areas of Sichang island, Thailand

BackgroundTha Wang and Tham Phang coasts, though situated at similar oceanographic positions on Sichang island, Chonburi province, Thailand, are different in bay geography and amount of municipal disturbances. These affect the marine ecosystems. The study used metagenomics combined with 16S and 18S rDNA pyrosequencing to identify types and distributions of archaea, bacteria, fungi and small eukaryotes of sizes ranges 0.45 and ~30 μm.ResultsFollowing the open bay geography and minimal municipal sewages, Tham Phang coast showed the cleaner water properties, described by color, salinity, pH, conductivity and percent dissolved oxygen. The 16S and 18S rDNA metagenomic profiles for Tha Wang and Tham Phang coasts revealed many differences, highlighting by low Lennon and Yue & Clayton theta similarity indices (66.03-73.03% for 16S rDNA profiles, 2.85-25.38% for 18S rDNA profiles). For 16S rDNA, the percent compositions of species belonging to Proteobacteria, Bacteroidetes, Cyanobacteria, Firmicutes, Verrucomicrobia, Gammatimonadetes, Tenericutes, Acidobacteria, Spirochaetes, Chlamydiae, Euryarchaeota, Nitrospirae, Planctomycetes, Thermotogae and Aquificae were higher or distinctly present in Tha Wang. In Tham Phang, except Actinobacteria, the fewer number of prokaryotic species existed. For 18S rDNA, fungi represented 74.745% of the species in Tha Wang, whereas only 6.728% in Tham Phang. Basidiomycota (71.157%) and Ascomycota (3.060%) were the major phyla in Tha Wang. Indeed, Tha Wang-to-Tham Phang percent composition ratios for fungi Basidiomycota and Chytridiomycota were 1264.701 and 25.422, respectively. In Tham Phang, Brachiopoda (lamp shells) and Mollusca (snails) accounted for 80.380% of the 18S rDNA species detected, and their proportions were approximately tenfold greater than those in Tha Wang. Overall, coastal Tham Phang comprised abundant animal species.ConclusionsTha Wang contained numerous archaea, bacteria and fungi, many of which could synthesize useful biotechnology gas and enzymes that could also function in high-saline and high-temperature conditions. Tham Phang contained less abundant archaea, bacteria and fungi, and the majority of the extracted metagenomes belonged to animal kingdom. Many microorganisms in Tham Phang were essential for nutrient-recycling and pharmaceuticals, for instances, Streptomyces, Pennicilium and Saccharomyces. Together, the study provided metagenomic profiles of free-living prokaryotes and eukaryotes in coastal areas of Sichang island.

Microbial ecology of Thailand tsunami and non-tsunami affected terrestrials.

The effects of tsunamis on microbial ecologies have been ill-defined, especially in Phang Nga province, Thailand. This ecosystem was catastrophically impacted by the 2004 Indian Ocean tsunami as well as the 600 year-old tsunami in Phra Thong island, Phang Nga province. No study has been conducted to elucidate their effects on microbial ecology. This study represents the first to elucidate their effects on microbial ecology. We utilized metagenomics with 16S and 18S rDNA-barcoded pyrosequencing to obtain prokaryotic and eukaryotic profiles for this terrestrial site, tsunami affected (S1), as well as a parallel unaffected terrestrial site, non-tsunami affected (S2). S1 demonstrated unique microbial community patterns than S2. The dendrogram constructed using the prokaryotic profiles supported the unique S1 microbial communities. S1 contained more proportions of archaea and bacteria domains, specifically species belonging to Bacteroidetes became more frequent, in replacing of the other typical floras like Proteobacteria, Acidobacteria and Basidiomycota. Pathogenic microbes, including Acinetobacter haemolyticus, Flavobacterium spp. and Photobacterium spp., were also found frequently in S1. Furthermore, different metabolic potentials highlighted this microbial community change could impact the functional ecology of the site. Moreover, the habitat prediction based on percent of species indicators for marine, brackish, freshwater and terrestrial niches pointed the S1 to largely comprise marine habitat indicating-species.

Population structure of four Thai indigenous chicken breeds

BackgroundIn recent years, Thai indigenous chickens have increasingly been bred as an alternative in Thailand poultry market. Due to their popularity, there is a clear need to improve the underlying quality and productivity of these chickens. Studying chicken genetic variation can improve the chicken meat quality as well as conserving rare chicken species. To begin with, a minimal set of molecular markers that can characterize the Thai indigenous chicken breeds is required.ResultsUsing AFLP-PCR, 30 single nucleotide polymorphisms (SNPs) from Thai indigenous chickens were obtained by DNA sequencing. From these SNPs, we genotyped 465 chickens from 7 chicken breeds, comprising four Thai indigenous chicken breeds- Pradhuhangdum (PD), Luenghangkhao (LK), Dang (DA) and Chee (CH), one wild chicken - the red jungle fowls (RJF), and two commercial chicken breeds - the brown egg layer (BL) and commercial broiler (CB). The chicken genotypes reveal unique genetic structures of the four Thai indigenous chicken breeds. The average expected heterozygosities of PD= 0.341, LK= 0.357, DA=0.349 and CH= 0.373, while the references RJF= 0.327, CB=0.324 and BL= 0.285. The FST values among Thai indigenous chicken breeds vary from 0.051 to 0.096. The FST values between the pairs of Thai indigenous chickens and RJF vary from 0.083 to 0.105 and the FST values between the Thai indigenous chickens and the two commercial chicken breeds vary from 0.116 to 0.221. A neighbour-joining tree of all individual chickens showed that the Thai indigenous chickens were clustered into four groups which were closely related to the wild RJF but far from the commercial breeds. Such commercial breeds were split into two closely groups. Using genetic admixture analysis, we observed that the Thai indigenous chicken breeds are likely to share common ancestors with the RJF, while both commercial chicken breeds share the same admixture pattern.ConclusionThese results indicated that the Thai indigenous chicken breeds may descend from the same ancestors. These indigenous chicken breeds were more closely related to red jungle fowls than those of the commercial breeds. These findings showed that the proposed SNP panel can effectively be used to characterize the four Thai indigenous chickens.

Genetic analysis of Thai cattle reveals a Southeast Asian indicine ancestry

Cattle commonly raised in Thailand have characteristics of Bos indicus (zebu). We do not know when or how cattle domestication in Thailand occurred, and so questions remain regarding their origins and relationships to other breeds. We obtained genome-wide SNP genotypic data of 28 bovine individuals sampled from four regions: North (Kho-Khaolampoon), Northeast (Kho-Isaan), Central (Kho-Lan) and South (Kho-Chon) Thailand. These regional varieties have distinctive traits suggestive of breed-like genetic variations. From these data, we confirmed that all four Thai varieties are Bos indicus and that they are distinct from other indicine breeds. Among these Thai cattle, a distinctive ancestry pattern is apparent, which is the purest within Kho-Chon individuals. This ancestral component is only present outside of Thailand among other indicine breeds in Southeast Asia. From this pattern, we conclude that a unique Bos indicus ancestor originated in Southeast Asia, and native Kho-Chon Thai cattle retain the signal of this ancestry with limited admixture of other bovine ancestors.

Pre-docking filter for protein and ligand 3D structures.

Virtual drug screening using protein-ligand docking techniques is a time-consuming process, which requires high computational power for binding affinity calculation. There are millions of chemical compounds available for docking. Eliminating compounds that are unlikely to exhibit high binding affinity from the screening set should speed-up the virtual drug screening procedure. We performed docking of 6353 ligands against twenty-one protein X-ray crystal structures. The docked ligands were ranked according to their calculated binding affinities, from which the top five hundred and the bottom five hundred were selected. We found that the volume and number of rotatable bonds of the top five hundred docked ligands are similar to those found in the crystal structures and corresponded with the volume of the binding sites. In contrast, the bottom five hundred set contains ligands that are either too large to enter the binding site, or too small to bind with high specificity and affinity to the binding site. A pre-docking filter that takes into account shapes and volumes of the binding sites as well as ligand volumes and flexibilities can filter out low binding affinity ligands from the screening sets. Thus, the virtual drug screening procedure speed is increased.

Bacterial communities on facial skin of teenage and elderly Thai females

The Human Microbiome Project was first established to understand the roles of human-associated microbes to human health and disease. This study presents preliminary findings of Thai female facial skin microbiome using three pooled samples from groups of skin microbiome profiles, namely (1) healthy and (2) acne-prone young adults (teenage.hea and teenage.acn) and (3) healthy elderly adults (elderly.hea) based on standard dermatological criteria. These samples were sequenced using 454-pyrosequencing targeting 16S rRNA (V3–V4 regions). Good’s coverage index of greater than 92% shows sufficient sampling of our data for each group. Three unique OTUs for each microbiome profile (43, 258 and 59 for teenage.hea, teenage.acn and ederly.hea, respectively) were obtained with 134 shared OTUs among the three datasets. Based on Morisita–Horn similarity coefficient, age is the major factor that brings the community relationship factor closer. The comparison among the three datasets reveal majority of Gemmatimonadetes, Planctomycetes and Nitrospirae in the teenage.hea, whereas Firmicutes are more prevalent in teenage.acn and elderly.hea skin types. In addition, when comparing Thai facial microbial diversity with the 16S data from U.S. forehead female database, significant differences were found among orders of bacteria, pointing to possible differences in human ecto-flora.

Diversity of bacterial communities on the facial skin of different age-group Thai males

Background Skin microbiome varies from person to person due to a combination of various factors, including age, biogeography, sex, cosmetics and genetics. Many skin disorders appear to be related to the resident microflora, yet databases of facial skin microbiome of many biogeographies, including Thai, are limited. Methods Metagenomics derived B-RISA and 16S rRNA gene sequencing was utilized to identify the culture-independent bacterial diversity on Thai male faces (cheek and forehead areas). Skin samples were categorized (grouped) into (i) normal (teenage.hea) and (ii) acne-prone (teenage.acn) young adults, and normal (iii) middle-aged (middle.hea) and (iv) elderly (elderly.hea) adults. Results The 16S rRNA gene sequencing was successful as the sequencing depth had an estimated >98% genus coverage of the true community. The major diversity was found between the young and elderly adults in both cheek and forehead areas, followed by that between normal and acne young adults. Detection of representative characteristics indicated that bacteria from the order Rhizobiales, genera Sphingomonas and Pseudoalteromonas, distinguished the elderly.hea microbiota, along the clinical features of wrinkles and pores. Prediction of the metabolic potential revealed reduced metabolic pathways involved in replication and repair, nucleotide metabolism and genetic translation in the elderly.hea compared with that in the teenage.hea. For young adults, some unique compositions such as abundance of Propionibacterium acnes and Staphylococcus epidermidis, with a minor diversity between normal and acne skins, were detected. The metabolic potentials of the acne vs. normal young adults showed that teenage.acn was low in many cellular processes (e.g., cell motility and environmental adaptation), but high in carbohydrate metabolism, which could support acne growth. Moreover, comparison with the age-matched males from the US (Boulder, Colorado) to gain insight into the diversity across national biogeography, revealed differences in the distribution pattern of species, although common bacteria were present in both biogeographical samples. Furthermore, B-RISA served as a crosscheck result to the 16S rRNA gene sequencing (i.e., differences between teenage and elderly microbiota). Conclusions This study revealed and compared the microbial diversity on different aged Thai male faces, and included analyses for representing the bacterial flora, the clinical skin characteristics, and comparison with the US age-matched. The results represent the first skin microbiota of Thai males, and helps the design of a large-scale skin microbiome study of Thais. The findings of the diversity among ages, skin type and national biogeography supported the importance of these traits in the skin microbiome and in developing a safe and sustainable treatment for acne and aging skin diseases.

Microbial communities in the reef water at Kham Island, lower Gulf of Thailand

Coral reefs are among the most biodiverse habitats on Earth, but knowledge of their associated marinemicrobiome remains limited. To increase the understanding of the coral reef ecosystem in the lower Gulf of Thailand, this study utilized 16S and 18S rRNA gene-based pyrosequencing to identify the prokaryotic and eukaryotic microbiota present in the reef water at Kham Island, Trat province, Thailand (N6.97 E100.86). The obtained result was then compared with the published microbiota from different coral reef water and marine sites. The coral reefs at Kham Island are of the fringe type. The reefs remain preserved and abundant. The community similarity indices (i.e., Lennon similarity index, Yue & Clayton similarity index) indicated that the prokaryotic composition of Kham was closely related to that of Kra, another fringing reef site in the lower Gulf of Thailand, followed by coral reef water microbiota at GS048b (Cooks Bay, Fr. Polynesia), Palmyra (Northern Line Islands, United States) and GS108b (Coccos Keeling, Australia), respectively. Additionally, the microbial eukaryotic populations at Kham was analyzed and compared with the available database at Kra. Both eukaryotic microbiota, in summer and winter seasons, were correlated. An abundance of Dinophysis acuminata was noted in the summer season, in accordance with its reported cause of diarrhoeatic shellfish outbreak in the summer season elsewhere. The slightly lower biodiversity in Kham than at Kra might reflect the partly habitat difference due to coastal anthropogenic activities and minor water circulation, as Kham locates close to the mainland and is surrounded by islands (e.g., Chang and Kut islands). The global marine microbiota comparison suggested relatively similar microbial structures among coral sites irrespective of geographical location, supporting the importance of coral-associated marine microbiomes, and Spearman’s correlation analysis between community membership and factors of shore distance and seawater temperature indicated potential correlation of these factors (p-values < 0.05) with Kham, Kra, and some other coral and coastal sites. Together, this study provided the second marine microbial database for the coral reef of the lower Gulf of Thailand, and a comparison of the coral-associated marine microbial diversity among global ocean sites.

Analysis of snake venom metalloproteinases from Myanmar Russell's viper transcriptome

&NA; Snake venom metalloproteinases (SVMPs) are the key enzymes in Russells viper (RV) venom which target all important components of haemostasis, such as clotting factors, platelets, endothelial cells and basement membrane. The structural diversity of SVMPs contributes to the broad spectrum of biological activities. The aim of the study was to investigate the SVMP transcript profile to gain better insights into the characteristic clinical manifestations of the Myanmar Russells viper (MRV) bites that distinguish it from the RVs of other habitats. Next generation sequencing (RNA‐Seq) of mRNA from MRV venom glands (2 males and 1 female) was performed on an Illumina HiSeq2000 platform and then de novo assembled using Trinity software. A total of 59 SVMP contigs were annotated through a Blastn search against the serpent nucleotide database from NCBI. Among them, disintegrins were the most abundant transcripts (75%) followed by the P‐III class SVMPs (25%). The P‐II SVMPs were scarce (0.002%), while no P‐I SVMPs were detectable in the transcriptome. For detailed structural analysis, contigs were conceptually translated and compared with amino acid sequences from other RVs and other vipers using Clustal Omega. The RTS‐disintegrin (jerdostatin homolog) was the most abundant among transcripts corresponding to 5 disintegrin isoforms. From 10 isoforms of SVMPs, RVV‐X, and Vipera lebetina apoptosis‐inducing protease (VLAIP) homolog, hereby termed Daboia siamensis AIP (DSAIP), were found to be highly expressed. Venom protein analysis using SDS‐PAGE followed by mass spectrometry revealed that the disintegrin was scarce, while the latter two SVMPs were abundant. These two proteins can contribute to severe clinical manifestations caused by MRV envenomation. HighlightsThe most abundant toxin transcripts in the Myanmar Russells viper transcriptome were annotated as metalloproteinases.Disintegrin transcripts were the most abundant transcripts (75%) followed by P‐III SVMPs (25%).Five isoforms of disintegrins and 10 isoforms of SVMPs were identified in the current transcriptomes.Protein analysis revealed that the disintegrin was scarce, while the P‐III SVMPs were abundant.The highly‐expressed P‐III SVMPs were RVV‐X and the novel Vipera lebetina apoptosis‐inducing protease (VLAIP) homolog.

The Plasmodium berghei RC strain is highly diverged and harbors putatively novel drug resistance variants

Background The current first line drugs for treating uncomplicated malaria are artemisinin (ART) combination therapies. However, Plasmodium falciparum parasites resistant to ART and partner drugs are spreading, which threatens malaria control efforts. Rodent malaria species are useful models for understanding antimalarial resistance, in particular genetic variants responsible for cross resistance to different compounds. Methods The Plasmodium berghei RC strain (PbRC) is described as resistant to different antimalarials, including chloroquine (CQ) and ART. In an attempt to identify the genetic basis for the antimalarial resistance trait in PbRC, its genome was sequenced and compared with five other previously sequenced P. berghei strains. Results We found that PbRC is eight-fold less sensitive to the ART derivative artesunate than the reference strain PbANKA. The genome of PbRC is markedly different from other strains, and 6,974 single nucleotide variants private to PbRC were identified. Among these PbRC private variants, non-synonymous changes were identified in genes known to modulate antimalarial sensitivity in rodent malaria species, including notably the ubiquitin carboxyl-terminal hydrolase 1 gene. However, no variants were found in some genes with strong evidence of association with ART resistance in P. falciparum such as K13 propeller protein. Discussion The variants identified in PbRC provide insight into P. berghei genome diversity and genetic factors that could modulate CQ and ART resistance in Plasmodium spp.