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Abstract This study established a novel infield sensing approach based on detection of the volatile compound markers in skin secretions. This was based on analysis of volatile compounds in axillary sweat samples collected from RT-PCR-proven Coronavirus disease 2019 (COVID-19) positive and negative populations using gas chromatography-mass spectrometry (GC–MS). The analysis proposed the possible markers of the monoaromatic compounds and ethyl hexyl acrylate. A portable photo ionization detector (PID) incorporated with the selective material towards the marker compounds was then developed with the pressurized injection approach. This provided the accuracy of 100% in the research phase (n = 125). The developed approach was then applied for screening of 2207 COVID-19 related cases covering the periods of the Alpha, Beta, Delta and Omicron variants of SARS-CoV-2 infection in Bangkok, Thailand. This offered the sensitivity, specificity and accuracy ranges of 92–99, 93–98 and 95–97%, respectively.

Background The appropriate sample handling for human fecal microbiota studies is essential to prevent changes in bacterial composition and quantities that could lead to misinterpretation of the data. Methods This study firstly identified the potential effect of aerobic and anaerobic fecal sample collection and transport materials on microbiota and quantitative microbiota in healthy and fat-metabolic disorder Thai adults aged 23–43 years. We employed metagenomics followed by 16S rRNA gene sequencing and 16S rRNA gene qPCR, to analyze taxonomic composition, alpha diversity, beta diversity, bacterial quantification, Pearson’s correlation with clinical factors for fat-metabolic disorder, and the microbial community and species potential metabolic functions. Results Our study successfully obtained microbiota results in percent and quantitative compositions. Each sample exhibited quality sequences with a >99% Good’s coverage index, and a relatively plateau rarefaction curve. Alpha diversity indices showed no statistical difference in percent and quantitative microbiota OTU richness and evenness, between aerobic and anaerobic sample transport materials. Obligate and facultative anaerobic species were analyzed and no statistical difference was observed. Supportively, the beta diversity analysis by non-metric multidimensional scale (NMDS) constructed using various beta diversity coefficients showed resembling microbiota community structures between aerobic and anaerobic sample transport groups (P = 0.86). On the other hand, the beta diversity could distinguish microbiota community structures between healthy and fat-metabolic disorder groups (P = 0.02), along with Pearson’s correlated clinical parameters (i.e., age, liver stiffness, GGT, BMI, and TC), the significantly associated bacterial species and their microbial metabolic functions. For example, genera such as Ruminococcus and Bifidobacterium in healthy human gut provide functions in metabolisms of cofactors and vitamins, biosynthesis of secondary metabolites against gut pathogens, energy metabolisms, digestive system, and carbohydrate metabolism. These microbial functional characteristics were also predicted as healthy individual biomarkers by LEfSe scores. In conclusion, this study demonstrated that aerobic sample collection and transport (

Ocular IPL therapy has recently been widely used for MGD, especially for patients not showing improvement with traditional therapies (warm compresses and lid scrubs) to clean debris and reduce bacterial overgrowth. Insights on the ocular microbiome and quantitative microbiome in MGD after a course of IPL could provide useful data on bacterial community monitoring and associated mechanisms linked with IPL. Ocular swabs were obtained from a severe MGD patient and age-sex matched healthy for metagenomics, followed by 16S rRNA gene sequencing and qPCR. Of 10 samples, including left and right eyes (el, er) of severe MGD females before (Db) and after 2-4 IPLs (Da2, Da3, and Da4) and the matched non-MGD females (H), both of ~40 years Using 16S rRNA gene sequencing as microbiota and combined 16S rRNA gene qPCR as quantitative microbiota revealed significant disperse in the microbiome structures of Db compared with Da and H (HOMOVA, p

Meibomian gland dysfunction (MGD) is a prevalent worldwide eye disorder that causes eye irritation, inflammation, chronic dryness, and blurred vision. Traditional therapies offer temporary improvement, but their efficacy varies in severe MGD cases. Ocular intense pulsed light (IPL) has emerged as a novel therapy, providing long-term symptom relief and shorter treatment durations compared to traditional approaches. However, the impact of IPL on the bacterial community within the eyes remains limited. To address this, we conducted a preliminary study using metagenomics and next-generation sequencing. We compared the bacterial eyelash communities of Thai females with severe MGD before and after 2-4 IPL treatments, and against a group of healthy females. Our findings revealed higher bacterial diversity in healthy individuals compared to severe MGD cases. IPL treatments increased diversity in the MGD group, making their core bacterial community more similar to that of healthy subjects. Notably, the presence of Koribacteraceae in severe MGD and Bifidobacterium in healthy individuals and post-IPL-treated MGD exemplified this shift. Clustering analysis showed a closer relationship between post-IPL-treated MGH and healthy subjects, while the pre-IPL treatment group formed a separate branch. These results suggest that IPL treatment can reshape the eyelash microbiome in MGD cases, but further research is needed to understand the implications and the microbiome’s role in MGD pathogenesis and treatment response.

Abstract High rates of new cervical cancer cases and deaths occur in low- and middle-income countries yearly, and one reason was found related to limitation of regular cervical cancer screening in local and low-resource settings. HPV has over 150 types, yet certain 14–20 high-risk and 13–14 low-risk types are common, and, thus, most conventional HPV nucleic acid assays, for examples, Cobas 4800 HPV test (Roche Diagnostics, New Jersey, USA) and REBA HPV-ID (Molecules and Diagnostics, Wonju, Republic of Korea) were developed to cover these types. We thereby utilized bioinformatics combined with recent isothermal amplification technique at 35–42 °C to firstly describe multiplex recombinase polymerase amplification assay that is specific to these common 20 high-risk and 14 low-risk types, and also L1 and E6/E7 genes that target different stages of cervical cancer development. Multiplex primer concentrations and reaction incubation conditions were optimized to allow simultaneous two gene detections at limit of detection of 1000 copies (equivalent to 2.01 fg) for L1 and 100 copies (0.0125 fg) for E6/E7, respectively. The assay was validated against urogenital and other pathogens, normal flora, and human control. In 130 real clinical sample tests, the assay demonstrated 100% specificity, 78% diagnostic accuracy, and 75% sensitivity compared with REBA HPV-ID test, and is much more rapid (15–40 min), less expensive (~ 3–4 USD/reaction) and does not require instrumentation (35–42 °C reaction condition so hand holding or tropical temperature is possible). Hence, the developed novel assay provides alternative screening tool for potential local screening. Furthermore, as this assay uses safe chemical reagents, it is safe for users.

Abstract Nucleic acid staining dyes are important tools for the analysis and visualizing of DNA/RNA in vitro and in the cells. Nevertheless, the range of commercially accessible dyes is still rather limited, and they are often very costly. As a result, finding nontoxic, easily accessible dyes, with desirable optical characteristics remains important. Styryl dyes have recently gained popularity as potential biological staining agents with many appealing properties, including a straightforward synthesis procedure, excellent photostability, tunable fluorescence, and high fluorescence quantum yield in the presence of nucleic acid targets with low background fluorescence signals. In addition to fluorescence, styryl dyes are strongly colored and exhibit solvatochromic properties which make them useful as colorimetric stains for low-cost and rapid testing of nucleic acids. In this work, novel dicationic styryl dyes bearing quaternary ammonium groups are designed to improve binding strength and optical response with target nucleic acids which contain a negatively charged phosphate backbone. Optical properties of the newly synthesized styryl dyes have been studied in the presence and absence of nucleic acid targets with the aim to find new dyes that can sensitively and specifically change fluorescence and/or color in the presence of nucleic acid targets. The binding interaction and optical response of the dicationic styryl dyes with nucleic acid were superior to the corresponding monocationic styryl dyes. Applications of the developed dyes for colorimetric detection of DNA in vitro and imaging of cellular nucleic acids are also demonstrated.

The impacts of metabolomic changes (reduced short-chain-fatty acids; SCFAs) in uremic condition is not fully understood. Once daily Candida gavage with or without probiotics (different times of administration) for 1 week prior to bilateral nephrectomy (Bil Nep) in 8-week-old C57BL6 mice as the possible models more resemble human conditions were performed. Candida-administered Bil Nep mice demonstrated more severe conditions than Bil Nep alone as indicated by mortality (n = 10/group) and other 48 h parameters (n = 6-8/group), including serum cytokines, leaky gut (FITC-dextran assay, endotoxemia, serum beta-glucan, and loss of Zona-occludens-1), and dysbiosis (increased Enterobacteriaceae with decreased diversity in microbiome analysis) (n = 3/group for fecal microbiome) without the difference in uremia (serum creatinine). With nuclear magnetic resonance metabolome analysis (n = 3-5/group), Bil Nep reduced fecal butyric (and propionic) acid and blood 3-hydroxy butyrate compared with sham and Candida-Bil Nep altered metabolomic patterns compared with Bil Nep alone. Then, Lacticaseibacillus rhamnosus dfa1 (SCFA-producing Lacticaseibacilli) (n = 8/group) attenuated the model severity (mortality, leaky gut, serum cytokines, and increased fecal butyrate) of Bil Nep mice (n = 6/group) (regardless of Candida). In enterocytes (Caco-2 cells), butyrate attenuated injury induced by indoxyl sulfate (a gut-derived uremic toxin) as indicated by transepithelial electrical resistance, supernatant IL-8, NFκB expression, and cell energy status (mitochondria and glycolysis activities by extracellular flux analysis). In conclusion, the reduced butyrate by uremia was not enhanced by Candida administration; however, the presence of Candida in the gut induced a leaky gut that was attenuated by SCFA-producing probiotics. Our data support the use of probiotics in uremia.

Abstract A symbiosis of bacterial community (sometimes called microbiota) play essential roles in developmental life cycle and health of coral, starting since a larva. For examples, coral bacterial holobionts function nitrogen fixation, carbon supply, sulfur cycling and antibiotic production. Yet, a study of the dynamic of bacteria associated coral larvae development is complicated owning to a vast diversity and culturable difficulty of bacteria; hence this type of study remains unexplored for Acropora humilis larvae in Thai sea. This study represented the first to utilize 16S rRNA gene sequencing to describe the timely bacterial compositions during successfully cultured and reared A. humilis larval transformation in aquaculture (gametes were collected from Sattahip Bay, Chonburi province, Thailand), from gamete spawning (0 h) and fertilization stage (1 h), to embryonic cleavage (8 h), round cell development (28, 39 and 41 h), and planula formation (48 h). The sequencing results as estimated by Good’s coverage at genus level covered 99.65 ± 0.24% of total bacteria. While core phyla of bacteria were observed (Proteobacteria, Actinobacteria, Firmicutes and Bacteroidetes), changes in bacterial population structures and differential predominant core bacterial orders were denoted for each larval developmental stage, from fertilization to embryonic cleavage and subsequently from the embryonic cleavage to round cell development (P = 0.007). For instances, Pseudoalteromonas and Oceanospirillales were found prevalent at 8 h, and Rhizobiales were at 48 h. The bacterial population structures from the round cell stage, particularly at 41 h, showed gradual drift towards those of the planula formation stage, suggesting microbial selection. Overall, this study provides preliminary insights into the dynamics of bacterial community and their potentially functional association (estimated from the bacterial compositions) during the developmental embryonic A. humilis in a cultivation system in Southeast Asia region.

Abstract Obesity, a major healthcare problem worldwide, induces metabolic endotoxemia through the gut translocation of lipopolysaccharides (LPS), a major cell wall component of Gram-negative bacteria, causing a chronic inflammatory state. A combination of several probiotics including Lactobacillus acidophilus 5 (LA5), a potent lactic acid-producing bacterium, has previously been shown to attenuate obesity. However, data on the correlation between a single administration of LA5 versus microbiota alteration might be helpful for the probiotic adjustment. LA5 was administered daily together with a high-fat diet (HFD) for 8 weeks in mice. Furthermore, the condition media of LA5 was also tested in a hepatocyte cell-line (HepG2 cells). Accordingly, LA5 attenuated obesity in mice as demonstrated by weight reduction, regional fat accumulation, lipidemia, liver injury (liver weight, lipid compositions, and liver enzyme), gut permeability defect, endotoxemia, and serum cytokines. Unsurprisingly, LA5 improved these parameters and acidified fecal pH leads to the attenuation of fecal dysbiosis. The fecal microbiome analysis in obese mice with or without LA5 indicated; (i) decreased Bacteroidetes (Gram-negative anaerobes that predominate in non-healthy conditions), (ii) reduced total fecal Gram-negative bacterial burdens (the sources of gut LPS), (iii) enhanced Firmicutes (Gram-positive bacteria with potential benefits) and (iv) increased Verrucomycobia, especially Akkermansia muciniphila, a bacterium with the anti-obesity property. With LA5 administration, A. muciniphila in the colon were more than 2,000 folds higher than the regular diet mice as determined by 16S rRNA. Besides, LA5 produced anti-inflammatory molecules with a similar molecular weight to LPS that reduced cytokine production in LPS-activated HepG2 cells. In conclusion, LA5 attenuated obesity through (i) gut dysbiosis attenuation, partly through the promotion of A. muciniphila (probiotics with the difficulty in preparation processes), (ii) reduced endotoxemia, and (iii) possibly decreased liver injury by producing the anti-inflammatory molecules.

Abstract Northeastern Thailand relies on agriculture as a major economic activity, and has used high levels of agrochemicals due to low facility, and salty sandy soil. To support soil recovery and sustainable agriculture, local farmers have used organic fertilizers from farmed animal feces. However, knowledge about these animal fecal manures remains minimal restricting their optimal use. Specifically, while bacteria are important for soil and plant growth, an abundance and a diversity of bacterial composition in these animal fecal manures have not been reported to allow selection and adjustment for a more effective organic fertilizer. This study thereby utilized metagenomics combined with 16S rRNA gene quantitative PCR (qPCR) and sequencing to analyze quantitative microbiota profiles in association with nutrients (N, P, K), organic matters, and the other physiochemical properties, of the commonly used earthworm manure and other manures from livestock animals (including breed and feeding diet variations) in the region. Unlike the other manures, the earthworm manure demonstrated more favorable nutrient profiles and physiochemical properties for forming fertile soil. Despite low total microbial biomass, the microbiota were enriched with maximal OTUs and Chao richness, and no plant pathogenic bacteria were found based on the VFDB database. The microbial metabolic potentials supported functions to promote crop growth, such as C, N and P cyclings, xenobiotic degradation, and synthesis of bioactive compounds. Pearson’s correlation analyses indicated that the quantitative microbiota of the earthworm manure were clustered in the same direction as N, and conductivity, salinity, and water content were essential to control the microbiota of animal manures.

As seawater temperature rises, repeated thermal bleaching events have negatively affected the reefs of the Andaman Sea for over decades. Studies on the coral-associated microbial diversity of prokaryotes and microbial eukaryotes (microbiome) in healthy and bleached corals are important to better understand the coral holobionts that involved augmented resistance to stresses, and this information remains limited in the Andaman Sea of Thailand. The present study thereby described the microbiomes of healthy (unbleached) and bleached colonies of four prevalent corals, Acropora humilis, Platygyra sp., Pocillopora damicornis, and Porites lutea, along with the surrounding seawater and sediments, that were collected during a 2016 thermal bleaching event, using 16S and 18S rRNA genes next-generation sequencing (NGS). Both prokaryotic and eukaryotic microbes showed isolated community profiles among sample types (corals, sediment, and seawater) [analysis of similarities (ANOSIM): p = 0.038 for prokaryotes, p < 0.001 for microbial eukaryotes] and among coral genera (ANOSIM: p < 0.001 for prokaryotes and microbial eukaryotes). In bleached state corals, we found differences in microbial compositions from the healthy state corals. Prevalent differences shared among bleached coral genera (shared in at least three coral genera) included a loss of reported coral-beneficial microbes, such as Pseudomonadales, Alteromonadales, and Symbiodinium; meanwhile an increase of putative coral-pathogenic Malassezia and Aspergillus. This difference could affect carbon and nitrogen availability for coral growth, reflective of a healthy or bleached state. Our findings in part supported previously microbial dysbiosis knowledge of thermal bleaching coral microbiomes around South East Asia marine geography, and together ongoing efforts are to support the understanding and management of microbial diversity to reduce the negative impacts to corals in massive thermal bleaching events.

Candida albicans is abundant in the human gut mycobiota but this species does not colonize the mouse gastrointestinal tract. C. albicans administration in dextran-sulfate solution (DSS) induced-colitis mouse model (DSS+Candida) might resemble more to human condition, therefore, a DSS colitis model with Candida administration was studied; first, to test the influence of fungi in DSS model and second, to test the efficacy of Lactobacillus rhamnosus L34. We demonstrated serum (1→3)-β-D-glucan (BG) elevation in patients with IBD and endoscopic moderate colitis in clinical remission, supporting the possible influence of gut fungi toward IBD in human. Then, in mouse model, Candida gavage was found to worsen the DSS model indicated by higher mortality rate, more severe colon histology and enhanced gut-leakage (FITC-dextran assay, endotoxemia, serum BG and blood bacterial burdens) but did not affect weight loss and diarrhea. DSS+Candida induced higher pro-inflammatory cytokines both in blood and in intestinal tissue. Worsened systemic pro-inflammatory cytokine responses in DSS+Candida compared with DSS alone was possibly due to the more severe translocation of LPS, BG and bacteria (not fungemia) from gut into systemic circulation. Interestingly, bacteremia from Pseudomonas aeruginosa was more frequently isolated from DSS+Candida than DSS alone. In parallel, P. aeruginosa was also isolated from fecal culture in most of the mice in DSS+Candida group supported by prominent Gammaproteobacteria in fecal microbioata analysis. However, L. rhamnosus L34 attenuated both DSS+Candida and DSS model through the attenuation of gut local inflammation (cytokines and histology), gut-leakage severity, fecal dysbiosis (culture method and microbiome analysis) and systemic inflammation (serum cytokines). In conclusion, gut Candida in DSS model induced fecal bacterial dysbiosis and enhanced leaky-gut induced bacteremia. Probiotic treatment strategy aiming to reduce gut-fungi and fecal dysbiosis could attenuate disease severity. Investigation on gut fungi in patients with IBD is highly interesting.

An outbreak of citrus greening or Huanglongbing disease bacteria occurs in many areas. We sampled and identified an ongoing ~year 2020 orange tree endemic in northern Thailand as Candidatus Liberibacter asiaticus. We thereby developed a plant greening disease (C. Liberibacter asiaticus) detection assay using simple alkaline heat DNA lysis and loop-mediated isothermal amplification coupled hydroxynaphthol blue (AL-LAMP-HNB), and evaluated the developed assay for its feasibility as point-of-care detection on 65 plant leaf samples with 100-1×104 copies of C. Liberibacter asiaticus or mocked injection compared with commercial DNA lysis kit and PCR-GE. Our assay is sensitive to 5-8.9 copies of omp (equaling 0.0056-0.01 fg) compatible with PCR-GE limit of detection. This ultra sensitive limit of detection could allow the disease detection before clinical apparent state of disease when C. Liberibacter asiaticus infection number is few, i.e. fewer than 100 copies of C. Liberibacter asiaticus. The assay is also specific with 6 degenerate primers targeting every strain of C. Liberibacter asiaticus omp from GenBank database, rapid (40 min total assay time), inexpensive (~2-3 USD/reaction), does not require sophisticated instrumentation, and has comparable assay accuracy (93.85-100% accuracy, 100% specificity, and 89.74-100% sensitivity) to bacterial DNA extraction by a commercial kit followed by PCR and gel electrophoresis (92.31% accuracy, 100% specificity, and 87.18% sensitivity) based on the real sample tests. Hence, the technique could be used in local or laboratory resource-restricted settings. The test result could be read by naked eyes through the color change from violet (negative) to sky blue (positive) for a C. Liberibacter asiaticus-infected specimen. Furthermore, this assay uses safe chemical reagents and, thus, is safe for the users.

Clostridioides difficile is a major cause of diarrhea in patients with antibiotic administration. Lacticaseibacillus casei T21, isolated from a human gastric biopsy, was tested in a murine C. difficile infection (CDI) model and colonic epithelial cells (Caco-2 and HT-29). Daily administration of L. casei T21 [1 × 108 colony forming units (CFU)/dose] for 4 days starting at 1 day before C. difficile challenge attenuated CDI as demonstrated by a reduction in mortality rate, weight loss, diarrhea, gut leakage, gut dysbiosis, intestinal pathology changes, and levels of pro-inflammatory cytokines [interleukin (IL)-1β, tumor necrosis factor (TNF)-α, macrophage inflammatory protein 2 (MIP-2), and keratinocyte chemoattractant (KC)] in the intestinal tissue and serum. Conditioned media from L. casei T21 exerted biological activities that fight against C. difficile as demonstrated in colonic epithelial cells by the following: (i) suppression of gene expression and production of IL-8, an important chemokine involved in C. difficile pathogenesis, (ii) reduction in the expression of SLC11A1 (solute carrier family 11 member 1) and HuR (human antigen R), important genes for the lethality of C. difficile toxin B, (iii) augmentation of intestinal integrity, and (iv) up-regulation of MUC2, a mucosal protective gene. These results supported the therapeutic potential of L. casei T21 for CDI and the need for further study on the intervention capabilities of CDI.

Although the impacts of Saccharomyces cerevisiae on cancers are mentioned, data on its use in mice with cyclic GMP-AMP synthase deficiency (cGAS-/-) are even rarer. Here, 12 weeks of oral administration of S. cerevisiae protected cGAS-/- mice from azoxymethane (AOM)-induced colon cancers, partly through dysbiosis attenuation (fecal microbiome analysis). In parallel, a daily intralesional injection of a whole glucan particle (WGP; the beta-glucan extracted from S. cerevisiae) attenuated the growth of subcutaneous tumor using MC38 (murine colon cancer cell line) in cGAS-/- mice. Interestingly, the incubation of fluorescent-stained MC38 with several subtypes of macrophages, including M1 (using Lipopolysaccharide; LPS), M2 (IL-4), and tumor-associated macrophages (TAM; using MC38 supernatant activation), could not further reduce the tumor burdens (fluorescent intensity) compared with M0 (control culture media). However, WGP enhanced tumoricidal activities (fluorescent intensity), the genes of M1 pro-inflammatory macrophage polarization (IL-1β and iNOS), and Dectin-1 expression and increased cell energy status (extracellular flux analysis) in M0, M2, and TAM. In M1, WGP could not increase tumoricidal activities, Dectin-1, and glycolysis activity, despite the upregulated IL-1β. In conclusion, S. cerevisiae inhibited the growth of colon cancers through dysbiosis attenuation and macrophage energy activation, partly through Dectin-1 stimulation. Our data support the use of S. cerevisiae for colon cancer protection.

Global warming has caused elevated seawater temperature and coral bleaching, including events on shallow reefs in the upper Gulf of Thailand (uGoT). Previous studies have reported an association between loss of zooxanthellae and coral bleaching. However, studies on the microbial diversity of prokaryotes and eukaryotes (microbiome) as coral holobionts are also important and this information is still limited in the uGoT. To address this shortcoming, this report provided baseline information on the prokaryotic (bacteria and archaea) and eukaryotic microbes of healthy and bleached colonies of four prevalent corals Acropora humilis, Acropora millepora, Platygyra sinensis, and Porites lutea and surrounding seawater and sediments, using 16S and 18S rRNA gene next-generation sequencing. Both prokaryotic and eukaryotic microbes showed isolated community profiles among sample types (corals, sediment, and seawater) (ANOSIM: P < 0.001, R = 0.51 for prokaryotic profiles and P < 0.001, R = 0.985 for eukaryotic microbe profiles). Among coral species, P. sinensis showed the most diverse prokaryotic community compared with the others (ANOSIM: P < 0.001, R = 0.636), and P. lutea showed the most diverse eukaryotic microbes (P = 0.014, R = 0.346). Healthy and bleached corals had some different microbiomes in species and their prevalences. For instance, the significant increase of Alphaproteobacteria in P. sinensis resulted in reduced prokaryotic community evenness and altered potential metabolic profiles (i.e., increased amino acid metabolism and genetic information processing and transcription, but decreased prokaryotic functions in cell motility, signaling, and transduction). For eukaryotic microbes, the loss of the algal Symbiodinium (colloquially known as zooxanthellae) in bleached corals such as P. lutea resulted in increased Chromista and Protista and, hence, clearly distinct eukaryotic microbe (including fungi) communities in healthy vs. bleached colonies of corals. Bleached corals were enriched in bacterial pathogens (e.g., Acinetobacter, Helicobacter, Malassesia, and Aspergillus) and decreased coral-beneficial prokaryotic and eukaryotic microbes (e.g., Rhizobiales and Symbiodinium). Additionally, this study identified microbiome species in bleached P. lutea that might help bleaching recovery (e.g., high abundance of Rhizobiales, Oceanospirillales, Flavobacteriales, and Alteromonadales). Overall, our coral-associated microbiome analyses identified altered diversity patterns of bacteria, archaea, fungi, and eukaryotic microbes between healthy and bleached coral species that are prevalent in the uGoT. This knowledge supports our ongoing efforts to manipulate microbial diversity as a means of reducing the negative impacts of thermal bleaching events in corals inhabiting the uGoT.

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Klebsiella pneumoniae is an opportunistic pathogen and a commensal organism that is possibly enhanced in several conditions with gut dysbiosis, and frequently detectable together with Candida overgrowth. Here, K. pneumoniae with or without Candida albicans was daily orally administered for 3 months in 0.8% dextran sulfate solution-induced mucositis mice and also tested in vitro. As such, Candida worsened Klebsiella-DSS-colitis as demonstrated by mortality, leaky gut (FITC-dextran assay, bacteremia, endotoxemia, and serum beta-glucan), gut dysbiosis (increased Deferribacteres from fecal microbiome analysis), liver pathology (histopathology), liver apoptosis (activated caspase 3), and cytokines (in serum and in the internal organs) when compared with Klebsiella-administered DSS mice. The combination of heat-killed Candida plus Klebsiella mildly facilitated inflammation in enterocytes (Caco-2), hepatocytes (HepG2), and THP-1-derived macrophages as indicated by supernatant cytokines or the gene expression. The addition of heat-killed Candida into Klebsiella preparations upregulated TLR-2, reduced Occludin (an intestinal tight junction molecule), and worsened enterocyte integrity (transepithelial electrical resistance) in Caco-2 and enhanced casp8 and casp9 (apoptosis genes) in HepG2 when compared with heat-killed Klebsiella alone. In conclusion, Candida enhanced enterocyte inflammation (partly through TLR-2 upregulation and gut dysbiosis) that induced gut translocation of endotoxin and beta-glucan causing hyper-inflammatory responses, especially in hepatocytes and macrophages.

Being ubiquitous, fungi are common opportunistic pathogens to humans that can lead to invasive and life-threatening infections in immunocompromised individuals. Eukaryote-resembling cell membrane and filamentous branches make the fungal diagnosis difficult. This study therefore developed a ready-to-use ITS1 loop-mediated isothermal amplification combined with hydroxynaphthol blue (LAMP-HNB) for rapid, sensitive and specific colorimetric detection of universal fungi in all phyla. The ITS1 LAMP-HNB could identify every evolutionary phylum of fungi according to sequence analyses. We tested a total of 30 clinically relevant fungal isolates (representing three major human pathogenic phyla of fungi, namely Zygomycota, Ascomycota and Basidiomycota) and 21 non-fungal isolates, and the ITS1 LAMP-HNB properly identified all isolates, with a detection limit of as low as 4.6 ag (9.6 copies), which was identical to ITS1 and 18S rDNA PCR. The assays were also validated on the feasibility of point-of-care diagnostic with real food (dry peanuts, chili and garlics) and blood samples. Furthermore, the shelf life of our ready-to-use ITS1 LAMP activity (≥50%) was more than 40 days at 30 °C with 3–5% polyvinyl alcohol or glycerol additive. The results supported the ready-to-use ITS1 LAMP-HNB for simple detection of fungi contamination with high sensitivity in local and resource-constrained areas to prevent opportunistic fungal species infections.

The southeast Andaman Sea 52-m off the west coast of Phang Nga province, Thailand, is located in the Indian Ocean, representing a hotspot for marine biodiversity of the world. This study utilized metagenomics combined 16S rRNA gene (V3–V4) sequencing, and firstly revealed the microbiota and their metabolism potentials of this site at an epipelagic depth (150-m depth, TC150M), including comparison with its pelagic depth (30-m depth, TC30M) as well as other nearby Thailand and global ocean sites. Between TC150M and TC30M, the TC150M microbial metagenomics was an eight-fold higher, and the microbiota comprised, for examples more abundant Bacteroidetes while fewer Proteobacteria, than the TC30M. The microbial metabolic potentials of the TC150M were statistically higher in replication repair and metabolisms of amino acids, lipids, nucleotides, and xenobiotics biodegradation, etc. Following comparative microbiota analyses between three Andaman Sea sites and two Gulf of Thailand sites, the relatively great proportions of Bacteroidetes, Nitrospirae, Gemmatimonadetes, and Chlorobi characterized the southeast Andaman Sea. Nevertheless, the microbiota representing Thailand marine sites remained distinguished from the global ocean sites where beta diversities were close. Thai maritime sites showed proportionally higher Proteobacteria, Bacteroides, Nitrospirae, Gemmatimonadetes, and Chlorobi. Thus, the Thai marine microbiota database helps better understand our global ocean microbiota and microbial metabolic potentials. Here, the microbial metabolism potentials between Thailand and the global ocean sites of relatively close microbiota databases encompose the similar functions yet in statistically different frequencies. Our research provided the first preliminary marine microbiome comparison between the epipelagic and pelagic sea levels of the southeast Andaman Sea, Thailand.

ABSTRACT The impact of gut fungi and (1→3)-β-d-glucan (BG), a major fungal cell wall component, on uremia was explored by Candida albicans oral administration in bilateral nephrectomy (BiNx) mice because of the prominence of C. albicans in the human intestine but not in mice. As such, BiNx with Candida administration (BiNx-Candida) enhanced intestinal injury (colon cytokines and apoptosis), gut leakage (fluorescein isothiocyanate [FITC]-dextran assay, endotoxemia, serum BG, and bacteremia), systemic inflammation, and liver injury at 48 h postsurgery compared with non-Candida BiNx mice. Interestingly, uremia-induced enterocyte apoptosis was severe enough for gut translocation of viable bacteria, as indicated by culture positivity for bacteria in blood, mesenteric lymph nodes (MLNs), and other organs, which was more severe in BiNx-Candida than in non-Candida BiNx mice. Candida induced alterations in the gut microbiota of BiNx mice as indicated by (i) the higher fungal burdens in the feces of BiNx-Candida mice than in sham-Candida mice by culture methods and (ii) increased Bacteroides with decreased Firmicutes and reduced bacterial diversity in the feces of BiNx-Candida mice compared with non-Candida BiNx mice by fecal microbiome analysis. In addition, lipopolysaccharide plus BG (LPS+BG), compared with each molecule alone, induced high supernatant cytokine levels, which were enhanced by uremic mouse serum in both hepatocytes (HepG2 cells) and macrophages (RAW264.7 cells). Moreover, LPS+BG, but not each molecule alone, reduced the glycolysis capacity and mitochondrial function in HepG2 cells as determined by extracellular flux analysis. Additionally, a probiotic, Lactobacillus rhamnosus L34 (L34), attenuated disease severity only in BiNx-Candida mice but not in non-Candida BiNx mice, as indicated by liver injury and serum cytokines through the attenuation of gut leakage, the fecal abundance of fungi, and fecal bacterial diversity but not fecal Gram-negative bacteria. In conclusion, Candida enhanced BiNx severity through the worsening of gut leakage and microbiota alterations that resulted in bacteremia, endotoxemia, and glucanemia. IMPORTANCE The impact of fungi in the intestine on acute uremia was demonstrated by the oral administration of Candida albicans in mice with the removal of both kidneys. Because fungi in the mouse intestine are less abundant than in humans, a Candida-administered mouse model has more resemblance to patient conditions. Accordingly, acute uremia, without Candida, induced intestinal mucosal injury, which resulted in the translocation of endotoxin, a major molecule of gut bacteria, from the intestine into blood circulation. In acute uremia with Candida, intestinal injury was more severe due to fungi and the alteration in intestinal bacteria (increased Bacteroides with decreased Firmicutes), leading to the gut translocation of both endotoxin from gut bacteria and (1→3)-β-d-glucan from Candida, which synergistically enhanced systemic inflammation in acute uremia. Both pathogen-associated molecules were delivered to the liver and induced hepatocyte inflammatory responses with a reduced energy production capacity, resulting in acute uremia-induced liver injury. In addition, Lactobacillus rhamnosus attenuated intestinal injury through reduced gut Candida and improved intestinal bacterial conditions.

Because of a possible impact of capsaicin in the high concentrations on enterocyte injury (cytotoxicity) and bactericidal activity on probiotics, Lactobacillus rhamnosus L34 (L34) and Lactobacillus rhamnosus GG (LGG), the probiotics derived from Thai and Caucasian population, respectively, were tested in the chili-extract administered C57BL/6 mice and in vitro experiments. In comparison with placebo, 2 weeks administration of the extract from Thai chili in mice caused loose feces and induced intestinal permeability defect as indicated by FITC-dextran assay and the reduction in tight junction molecules (occludin and zona occludens-1) using fluorescent staining and gene expression by quantitative real-time polymerase chain reaction (qRT-PCR). Additionally, the chili extracts also induced the translocation of gut pathogen molecules; lipopolysaccharide (LPS) and (1→3)-β-d-glucan (BG) and fecal dysbiosis (microbiome analysis), including reduced Firmicutes, increased Bacteroides, and enhanced total Gram-negative bacteria in feces. Both L34 and LGG attenuated gut barrier defect (FITC-dextran, the fluorescent staining and gene expression of tight junction molecules) but not improved fecal consistency. Additionally, high concentrations of capsaicin (0.02-2 mM) damage enterocytes (Caco-2 and HT-29) as indicated by cell viability test, supernatant cytokine (IL-8), transepithelial electrical resistance (TEER) and transepithelial FITC-dextran (4.4 kDa) but were attenuated by Lactobacillus condition media (LCM) from both probiotic-strains. The 24 h incubation with 2 mM capsaicin (but not the lower concentrations) reduced the abundance of LGG (but not L34) implying a higher capsaicin tolerance of L34. However, Lactobacillus rhamnosus fecal abundance, using qRT-PCR, of L34 or LGG after 3, 7, and 20 days of the administration in the Thai healthy volunteers demonstrated the similarity between both strains. In conclusion, high dose chili extracts impaired gut permeability and induced gut dysbiosis but were attenuated by probiotics. Despite a better capsaicin tolerance of L34 compared with LGG in vitro, L34 abundance in feces was not different to LGG in the healthy volunteers. More studies on probiotics with a higher intake of chili in human are interesting.

Although bacteria-free DNA in blood during systemic infection is mainly derived from bacterial death, translocation of the DNA from the gut into the blood circulation (gut translocation) is also possible. Hence, several mouse models with experiments on macrophages were conducted to explore the sources, influences, and impacts of bacteria-free DNA in sepsis. First, bacteria-free DNA and bacteriome in blood were demonstrated in cecal ligation and puncture (CLP) sepsis mice. Second, administration of bacterial lysate (a source of bacterial DNA) in dextran sulfate solution (DSS)-induced mucositis mice elevated blood bacteria-free DNA without bacteremia supported gut translocation of free DNA. The absence of blood bacteria-free DNA in DSS mice without bacterial lysate implies an impact of the abundance of bacterial DNA in intestinal contents on the translocation of free DNA. Third, higher serum cytokines in mice after injection of combined bacterial DNA with lipopolysaccharide (LPS), when compared to LPS injection alone, supported an influence of blood bacteria-free DNA on systemic inflammation. The synergistic effects of free DNA and LPS on macrophage pro-inflammatory responses, as indicated by supernatant cytokines (TNF-α, IL-6, and IL-10), pro-inflammatory genes (NFκB, iNOS, and IL-1β), and profound energy alteration (enhanced glycolysis with reduced mitochondrial functions), which was neutralized by TLR-9 inhibition (chloroquine), were demonstrated. In conclusion, the presence of bacteria-free DNA in sepsis mice is partly due to gut translocation of bacteria-free DNA into the systemic circulation, which would enhance sepsis severity. Inhibition of the responses against bacterial DNA by TLR-9 inhibition could attenuate LPS-DNA synergy in macrophages and might help improve sepsis hyper-inflammation in some situations.

A chronic kidney disease (CKD) causes uremic toxin accumulation and gut dysbiosis, which further induces gut leakage and worsening CKD. Lipopolysaccharide (LPS) of Gram-negative bacteria and (1➔3)-β-D-glucan (BG) of fungi are the two most abundant gut microbial molecules. Due to limited data on the impact of intestinal fungi in CKD mouse models, the influences of gut fungi and Lacticaseibacillus rhamnosus L34 (L34) on CKD were investigated using oral C. albicans-administered 5/6 nephrectomy (5/6Nx) mice. At 16 weeks post-5/6Nx, Candida-5/6Nx mice demonstrated an increase in proteinuria, serum BG, serum cytokines (tumor necrotic factor-α; TNF-α and interleukin-6), alanine transaminase (ALT), and level of fecal dysbiosis (Proteobacteria on fecal microbiome) when compared to non-Candida-5/6Nx. However, serum creatinine, renal fibrosis, or gut barrier defect (FITC-dextran assay and endotoxemia) remained comparable between Candida- versus non-Candida-5/6Nx. The probiotics L34 attenuated several parameters in Candida-5/6Nx mice, including fecal dysbiosis (Proteobacteria and Bacteroides), gut leakage (fluorescein isothiocyanate (FITC)-dextran), gut-derived uremic toxin (trimethylamine-N-oxide; TMAO) and indoxyl sulfate; IS), cytokines, and ALT. In vitro, IS combined with LPS with or without BG enhanced the injury on Caco-2 enterocytes (transepithelial electrical resistance and FITC-dextran permeability) and bone marrow-derived macrophages (supernatant cytokines (TNF-α and interleukin-1 β; IL-1β) and inflammatory genes (TNF-α, IL-1β, aryl hydrocarbon receptor, and nuclear factor-κB)), compared with non-IS activation. These injuries were attenuated by the probiotics condition media. In conclusion, Candida administration worsens kidney damage in 5/6Nx mice through systemic inflammation, partly from gut dysbiosis-induced uremic toxins, which were attenuated by the probiotics. The additive effects on cell injury from uremic toxin (IS) and microbial molecules (LPS and BG) on enterocytes and macrophages might be an important underlying mechanism.

Fat reduction and anti-inflammation are commonly claimed properties of probiotics. Lactiplantibacillus plantarum and Enterococcus faecium were tested in high fat-induced obesity mice and in vitro experiments. After 16 weeks of probiotics, L. plantarum dfa1 outperforms E. faecium dfa1 on the anti-obesity property as indicated by body weight, regional fat accumulation, serum cholesterol, inflammatory cytokines (in blood and colon tissue), and gut barrier defect (FITC-dextran assay). With fecal microbiome analysis, L. plantarum dfa1 but not E. faecium dfa1 reduced fecal abundance of pathogenic Proteobacteria without an alteration in total Gram-negative bacteria when compared with non-probiotics obese mice. With palmitic acid induction, the condition media from both probiotics similarly attenuated supernatant IL-8, improved enterocyte integrity and down-regulated cholesterol absorption-associated genes in Caco-2 cell (an enterocyte cell line) and reduced supernatant cytokines (TNF-α and IL-6) with normalization of cell energy status (extracellular flux analysis) in bone-marrow-derived macrophages. Due to the anti-inflammatory effect of the condition media of both probiotics on palmitic acid-activated enterocytes was neutralized by amylase, the active anti-inflammatory molecules might, partly, be exopolysaccharides. As L. plantarum dfa1 out-performed E. faecium dfa1 in anti-obesity property, possibly through the reduced fecal Proteobacteria, with a similar anti-inflammatory exopolysaccharide; L. plantarum is a potentially better option for anti-obesity than E. faecium.

In this study, coral disease was first reported in the coral hatchery in Thailand. Disease were usually found on corals aged two to five years old during the months of November to December of each year. To identify bacterial strains, culture-based methods for strain isolation and molecular techniques of the 16S rRNA gene analysis were used. The resuts showed that the dominant genera of bacteria in diseased corals were Vibrio spp. (comprising 41.01% of the isolates). The occurrence of the disease in the coral hatchery can have a significant effect on the health and survival of juvenile corals before being transplanted to natural reefs for restoration.

ABSTRACT Clinical persistence of Chlamydia trachomatis (Ct) sexually transmitted infections (STIs) is a major public health concern. In vitro persistence is known to develop through interferon gamma (IFN-γ) induction of indoleamine 2,3-dioxygenase (IDO), which catabolizes tryptophan, an essential amino acid for Ct replication. The organism can recover from persistence by synthesizing tryptophan from indole, a substrate for the enzyme tryptophan synthase. The majority of Ct strains, except for reference strain B/TW-5/OT, contain an operon comprised of α and β subunits that encode TrpA and TrpB, respectively, and form a functional αββα tetramer. However, trpA mutations in ocular Ct strains, which are responsible for the blinding eye disease known as trachoma, abrogate tryptophan synthesis from indole. We examined serial urogenital samples from a woman who had recurrent Ct infections over 4 years despite antibiotic treatment. The Ct isolates from each infection episode were genome sequenced and analyzed for phenotypic, structural, and functional characteristics. All isolates contained identical mutations in trpA and developed aberrant bodies within intracellular inclusions, visualized by transmission electron microscopy, even when supplemented with indole following IFN-γ treatment. Each isolate displayed an altered αββα structure, could not synthesize tryptophan from indole, and had significantly lower trpBA expression but higher intracellular tryptophan levels compared with those of reference Ct strain F/IC-Cal3. Our data indicate that emergent mutations in the tryptophan operon, which were previously thought to be restricted only to ocular Ct strains, likely resulted in in vivo persistence in the described patient and represents a novel host-pathogen adaptive strategy for survival. IMPORTANCE Chlamydia trachomatis (Ct) is the most common sexually transmitted bacterium with more than 131 million cases occurring annually worldwide. Ct infections are often asymptomatic, persisting for many years despite treatment. In vitro recovery from persistence occurs when indole is utilized by the organism’s tryptophan synthase to synthesize tryptophan, an essential amino acid for replication. Ocular but not urogenital Ct strains contain mutations in the synthase that abrogate tryptophan synthesis. Here, we discovered that the genomes of serial isolates from a woman with recurrent, treated Ct STIs over many years were identical with a novel synthase mutation. This likely allowed long-term in vivo persistence where active infection resumed only when tryptophan became available. Our findings indicate an emerging adaptive host-pathogen evolutionary strategy for survival in the urogenital tract that will prompt the field to further explore chlamydial persistence, evaluate the genetics of mutant Ct strains and fitness within the host, and their implications for disease pathogenesis.

Tha Wang coast in Sichang island, Chonburi province, Thailand, serves a major pier, cargo transportation and is populated with anthropogenic activities. Hence, monitoring of environmental status, in part by microbial diversity, is essential. Previously, the coastal marine microbiota of this area in a dry season (February 2011) had been reported, but the database for a wet season is none. As marine microbiota in several sites were reported fluctuate between dry and wet seasons, specifically in the 2011 that one of the largest flood in Thailand history even occurred from July 2011 to January 2012; the present study thereby first revealed the coastal Sichang water microbiota in the 2011 wet season (July 2011) using metagenomic derived 16S rRNA gene sequencing. Wet and dry seasons showed differences in prokaryotic community richness and evenness: community similarity indices by Jaccard, Lennon, and Yue and Clayton were 2.06–58.28%. This resulted in differences in metabolic potentials, for instances the lower metabolisms for energy and nutrient cycling but the greater in pathogenesis in wet season. Our data were also compared against the flooding water microbiota nearby to Chonburi province to observe any possible correlation; and the coastal Sichang was found distinct given diverse sources of flooding water and local contamination might affect the diversity. In conclusion, this wet-season microbiota supported the better knowledge on the seasonal microbiota of coastal Sichang island.

Gut fungi may influence the course of Clostridium difficile infection either positively or negatively for the host. Fungi are not prominent in the mouse gut, and C. albicans, the major human gastrointestinal commensal yeast, is in low abundance or absent in mice. Bifidobacterium is one of the probiotics that may attenuate the severity of C. difficile infection. Inflammatory synergy between C. albicans and C. difficile, in gut, may provide a state that more closely resembles human infection and be more suitable for testing probiotic effects. We performed fecal mycobiota analysis and administered C. albicans at 1 day prior to C. difficile dosing. Fecal eukaryotic 18S rDNA analysis demonstrated the presence of Ascomycota, specifically, Candida spp., after oral antibiotics, despite negative fecal fungal culture. C. albicans administration enhanced the severity of the C. difficile infection model as determined by mortality rate, weight loss, gut leakage (FITC-dextran assay), and serum and intestinal tissue cytokines. This occurred without increased fecal C. difficile or bacteremia, in comparison with C. difficile gavage alone. Candida lysate with C. difficile increased IL-8 production from HT-29 and Caco-2 human intestinal epithelial cell-lines. Bifidobacterium attenuated the disease severity of the C. difficile plus Candida model. The reduced severity was associated with decreased Candida burdens in feces. In conclusion, gut C. albicans worsened C. difficile infection, possibly through exacerbation of inflammation. Hence, a mouse model of Clostridium difficile infection with C. albicans present in the gut may better model the human patient condition. Gut fungal mycobiome investigation in patients with C. difficile is warranted and may suggest therapeutic targets.

Reefs at Ko Samae San (S), Khao Ma Cho (K), and Ko Tao Mo (T), in the Gulf of Thailand (GoT) represent a biodiversity hotspot, and bacteria play significant roles in maintaining the health of these coral reefs and their biogeochemical cycles. Therefore, this study analyzed bacterial communities (microbiota) from healthy corals and nearby seawater and sediment, using B-RISA and 16S rRNA gene sequencing. Sampling was done in one dry and one wet season to provide an initial assessment of variation with environmental conditions. The most prevalent coral species were Porites lutea, Platygyra sinensis, Acropora humilis, and Acropora millepora. The B-RISA and the sequencing results were correlated, which increased confidence the results. The microbiota varied among corals, seawater, and sediment and between the wet and dry seasons. Percentages of bacteria with known functions varied among sample types and seasons, and their relative abundances supported previously reported essential functions, such as prevention of disease (e.g., Pseudoalteromonas, Psychrobacter, and Cobetia were more abundant on corals in the dry season). Pearson's correlations and multiple factor regressions identified dissolved oxygen (DO), temperature, salinity, and density as significant influences on the microbiota. The equations estimated the relative abundance of a community comprising 147 bacterial genera, as well as the relative abundance of Pseudomonas, Clostridium, Verrucomicrobium, and Epulopiscium (R2 ≥ 0.721). These results represent the first descriptions of microbiota from corals, and surrounding seawater and sediments in the upper GoT.

Reef sites of Ko Samae San (S), Khao Ma Cho (K) and Ko Tao Mo (T) in the upper Gulf of Thailand have abundant corals and represent a hotspot of marine biodiversity. Coral reefs serve as major networks of food and energy, where bacteria, microbial eukaryotes (fungi) and small eukaryotes play significant roles as primary producers that convert inorganic compounds to organic compounds, degraders of toxic substances, and recyclers. These functions sustain food and energy supplies. Advances in metagenomics and next-generation sequencing can provide knowledge of diversity without limitations imposed by media and other conditions associated with laboratory cultures. Scientists have researched bacterial diversity of coral sites; however, a database for fungi and small eukaryotes from Thailand’s sites with abundant corals is lacking. The present study combined fungal ribosomal intergenic spacer analysis (F-RISA) and 18S rRNA gene sequencing to unveil the first culture-independent microbial and small eukaryotes from these sites at two times and across four species of coral (Porites lutea, Platygyra sinensis, Acropora humilis, and Acropora millepora), seawater and sediment. Results showed that the small eukaryotic communities on corals were distinct from communities in the surrounding seawater and sediment. The communities were relatively similar at the three sites and during the two periods of time. Pearson’s correlations indicated the community diversity were associated with water quality (e.g., dissolved oxygen concentrations and density of water).

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.

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.

Candida albicans is the most common fungus in the human intestinal microbiota but not in mice. To make a murine sepsis model more closely resemble human sepsis and to explore the role of intestinal C. albicans, in the absence of candidemia, in bacterial sepsis, live- or heat-killed C. albicans was orally administered to mice at 3h prior to cecal ligation and puncture (CLP). A higher mortality rate of CLP was demonstrated with Candida-administration (live- or heat-killed) prior to CLP. Fecal Candida presented only in experiments with live-Candida administration. Despite the absence of candidemia, serum (1→3)-β-D-glucan (BG) was higher in CLP with Candida-administration than CLP-controls (normal saline administration) at 6h and/or 18h post-CLP. Interestingly, fluconazole attenuated the fecal Candida burden and improved survival in mice with live-Candida administration, but not CLP-control. Microbiota analysis revealed increased Bacteroides spp. and reduced Lactobacillus spp. in feces after Candida administration. Additionally, synergy in the elicitation of cytokine production from bone marrow-derived macrophages, in vitro, was demonstrated by co-exposure to heat-killed E. coli and BG. In conclusion, intestinal abundance of fungi and/or fungal-molecules was associated with increased bacterial sepsis-severity, perhaps through enhanced cytokine elicitation induced by synergistic responses to molecules from gut-derived bacteria and fungi. Conversely, reducing intestinal fungal burdens decreased serum BG and attenuated sepsis in our model.

Chlamydia trachomatis is the leading cause of preventable blindness and bacterial sexually transmitted diseases worldwide. Plaque assays have been used to clonally segregate laboratory-adapted C. trachomatis strains from mixed infections, but no assays have been reported to segregate clones from recent clinical samples. We developed a novel shotgun cell culture harvest assay for this purpose because we found that recent clinical samples do not form plaques. Clones were strain-typed by using outer membrane protein A and 16S rRNA sequences. Surprisingly, ocular trachoma reference strain A/SA-1 contained clones of Chlamydophila abortus. C. abortus primarily infects ruminants and pigs and has never been identified in populations where trachoma is endemic. Three clonal variants of reference strain Ba/Apache-2 were also identified. Our findings reflect the importance of clonal isolation in identifying constituents of mixed infections containing new or emerging strains and of viable clones for research to more fully understand the dynamics of in vivo strain-mixing, evolution, and disease pathogenesis.

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.

ABSTRACT Chlamydia trachomatis is an obligate intracellular bacterium that causes a diversity of severe and debilitating diseases worldwide. Sporadic and ongoing outbreaks of lymphogranuloma venereum (LGV) strains among men who have sex with men (MSM) support the need for research on virulence factors associated with these organisms. Previous analyses have been limited to single genes or genomes of laboratory-adapted reference strain L2/434 and outbreak strain L2b/UCH-1/proctitis. We characterized an unusual LGV strain, termed L2c, isolated from an MSM with severe hemorrhagic proctitis. L2c developed nonfusing, grape-like inclusions and a cytotoxic phenotype in culture, unlike the LGV strains described to date. Deep genome sequencing revealed that L2c was a recombinant of L2 and D strains with conserved clustered regions of genetic exchange, including a 78-kb region and a partial, yet functional, toxin gene that was lost with prolonged culture. Indels (insertions/deletions) were discovered in an ftsK gene promoter and in the tarp and hctB genes, which encode key proteins involved in replication, inclusion formation, and histone H1-like protein activity, respectively. Analyses suggest that these indels affect gene and/or protein function, supporting the in vitro and disease phenotypes. While recombination has been known to occur for C. trachomatis based on gene sequence analyses, we provide the first whole-genome evidence for recombination between a virulent, invasive LGV strain and a noninvasive common urogenital strain. Given the lack of a genetic system for producing stable C. trachomatis mutants, identifying naturally occurring recombinants can clarify gene function and provide opportunities for discovering avenues for genomic manipulation. IMPORTANCE Lymphogranuloma venereum (LGV) is a prevalent and debilitating sexually transmitted disease in developing countries, although there are significant ongoing outbreaks in Australia, Europe, and the United States among men who have sex with men (MSM). Relatively little is known about LGV virulence factors, and only two LGV genomes have been sequenced to date. We isolated an LGV strain from an MSM with severe hemorrhagic proctitis that was morphologically unique in tissue culture compared with other LGV strains. Bioinformatic and statistical analyses identified the strain as a recombinant of L2 and D strains with highly conserved clustered regions of genetic exchange. The unique culture morphology and, more importantly, disease phenotype could be traced to the genes involved in recombination. The findings have implications for bacterial species evolution and, in the case of ongoing LGV outbreaks, suggest that recombination is a mechanism for strain emergence that results in significant disease pathology.