Project description:Dysbiosis in the gut microbiome has been implicated in several diseases including auto-immune diseases, inflammatory diseases, cancers and mental disorders. Keratitis is an inflammatory disease of the eye significantly contributing to corneal blindness in the developing world. It would be worthwhile to investigate the possibility of dysbiosis in the gut microbiome being associated with Keratitis. Here, we have analyzed fungal and bacterial populations in stool samples through high-throughput sequencing of the ITS2 region for fungi and V3-V4 region of 16S rRNA gene for bacteria in healthy controls (HC, n = 31) and patients with fungal keratitis (FK, n = 32). Candida albicans (2 OTUs), Aspergillus (1 OTU) and 3 other denovo-OTUs were enriched in FK samples and an unclassified denovo-OTU was enriched in HC samples. However, the overall abundances of these 'discriminatory' OTUs were very low (< 0.001%) and not indicative of significant dysbiosis in the fungal community inhabiting the gut of FK patients. In contrast, the gut bacterial richness and diversity in FK patients was significantly decreased when compared to HC. 52 OTUs were significantly enriched in HC samples whereas only 5 OTUs in FK. The OTUs prominently enriched in HC were identified as Faecalibacterium prausnitzii, Bifidobacterium adolescentis, Lachnospira, Mitsuokella multacida, Bacteroides plebeius, Megasphaera and Lachnospiraceae. In FK samples, 5 OTUs affiliated to Bacteroides fragilis, Dorea, Treponema, Fusobacteriaceae, and Acidimicrobiales were significantly higher in abundance. The functional implications are that Faecalibacterium prausnitzii, an anti-inflammatory bacterium and Megasphaera, Mitsuokella multacida and Lachnospira are butyrate producers, which were enriched in HC patients, whereas Treponema and Bacteroides fragilis, which are pathogenic were abundant in FK patients, playing a potential pro-inflammatory role. Heatmap, PCoA plots and functional profiles further confirm the distinct patterns of gut bacterial composition in FK and HC samples. Our study demonstrates dysbiosis in the gut bacterial microbiomes of FK patients compared to HC. Further, based on inferred functions, it appears that dysbiosis in the gut of FK subjects is strongly associated with the disease phenotype with decrease in abundance of beneficial bacteria and increase in abundance of pro-inflammatory and pathogenic bacteria.

Project description:BackgroundColorectal cancer is a leading cause of cancer-related deaths worldwide. The human gut microbiome has become an active area of research for understanding the initiation, progression, and treatment of colorectal cancer. Despite multiple studies having found significant alterations in the carriage of specific bacteria within the gut microbiome of colorectal cancer patients, no single bacterium has been unequivocally connected to all cases. Whether alterations in species carriages are the cause or outcome of cancer formation is still unclear, but what is clear is that focus should be placed on understanding changes to the bacterial community structure within the cancer-associated gut microbiome.ResultsBy applying a novel set of analyses on 252 previously published whole-genome shotgun sequenced fecal samples from healthy and late-stage colorectal cancer subjects, we identify taxonomic, functional, and structural changes within the cancer-associated human gut microbiome. Bacterial association networks constructed from these data exhibited widespread differences in the underlying bacterial community structure between healthy and colorectal cancer associated gut microbiomes. Within the cancer-associated ecosystem, bacterial species were found to form associations with other species that are taxonomically and functionally dissimilar to themselves, as well as form modules functionally geared towards potential changes in the tumor-associated ecosystem. Bacterial community profiling of these samples revealed a significant increase in species diversity within the cancer-associated gut microbiome, and an elevated relative abundance of species classified as originating from the oral microbiome including, but not limited to, Fusobacterium nucleatum, Peptostreptococcus stomatis, Gemella morbillorum, and Parvimonas micra. Differential abundance analyses of community functional capabilities revealed an elevation in functions linked to virulence factors and peptide degradation, and a reduction in functions involved in amino-acid biosynthesis within the colorectal cancer gut microbiome.ConclusionsWe utilize whole-genome shotgun sequenced fecal samples provided from a large cohort of late-stage colorectal cancer and healthy subjects to identify a number of potentially important taxonomic, functional, and structural alterations occurring within the colorectal cancer associated gut microbiome. Our analyses indicate that the cancer-associated ecosystem influences bacterial partner selection in the native microbiota, and we highlight specific oral bacteria and their associations as potentially relevant towards aiding tumor progression.

Project description:Carotid atherosclerosis (CAS) is a reflection of systemic atherosclerosis and the main pathological processes of cardiovascular disease (CVD), namely, carotid intima-media thickening, carotid plaque formation, and carotid stenosis. Accumulating evidence indicates that the gut microbiota plays an important role in CVD and gut-brain disorders, but the associations of the composition and metabolites of the gut microbiome with CAS have not been studied comprehensively. We performed a gut microbiome genome-wide association study in 31 patients with CAS and 51 healthy controls using whole-genome shotgun sequencing. We found that several risk factors (waist circumference, body mass index, diastolic blood pressure, systolic blood pressure, fasting blood glucose, glycated hemoglobin A1c, total cholesterol, triglyceride, and low-density lipoprotein cholesterol) and inflammatory markers (white blood cell count and absolute value of neutrophils) were significantly higher in the CAS group than in the control group. In addition, 21 species and 142 pathways were enriched in the CAS group, and 10 species and 1 pathway were enriched in the control group. Specifically, Bacteroides eggerthii, Escherichia coli, and Klebsiella pneumoniae were the most abundant species in the CAS group, whereas Parabacteroides unclassified, Prevotella copri, Bacteroides sp 3_1_19, and Haemophilus parainfluenzae were the most abundant species in the control group. Finally, we found that most gut microbes and microbial pathways that were enriched in the CAS group had significant positive correlations with clinical characteristics, whereas the microbes and pathways enriched in healthy controls had significant negative correlations with clinical characteristics excluding high-density lipoprotein cholesterol. In addition, the associations between gut microbes and some microbial pathways (short-chain fatty acid, lipopolysaccharide, and menaquinol biosynthesis) were identified. Our results indicate the existence of a cyclic pathway that elevates the circulating concentrations of trimethylamine-N-oxide in patients with CAS but reduces its concentrations in healthy controls.

Project description:Microbial keratitis is a major cause of blindness worldwide. An excessive host inflammatory response can occur even after adequate antimicrobial treatment. This results in tissue damage with corneal thinning and even perforation, which may require corneal transplantation. In this study we investigated the pathways involved in the pathophysiology of this disease by comparing the human transcriptome profile of tissue from culture-proven bacterial and fungal keratitis (n=7 and n=8 respectively) with normal non-infected cadaveric corneal tissue (C, n=12) using Illumina HT12 v4 microarrays. The causative organisms were Streptococcus pneumoniae (n=6) and Pseudomonas aeruginosa (n=1) for bacterial keratitis (BK). Fungal keratitis (FK) was caused by Fusarium sp. (n=5), Aspergillus sp. (n=2, A. flavus and terreus) and Lasiodiplodia sp. (n=1). Differential expression (DE) analysis revealed 2310 significantly altered probes in the BK v C comparison, and 1813 probes for FK v C. The most highly upregulated gene in both comparisons was MMP9 with fold changes (FC) of 64 (fdr-adjusted p<6 x10-11) for FK v C and 89 for BK v C (fdr-adjusted p<4 x10-11) respectively. Network co-expression analyses revealed the defense response, inflammatory response and extracellular matrix mechanisms to be the main functional pathways involved. Microarray results were validated by performing real-time quantitative PCR (RTqPCR) for 46 DE genes using RNA extracted from the same samples. There was a high correlation between log2 FC values from microarray and RTqPCR. Further studies are needed to evaluate the most highly differentially expressed genes as possible biomarkers of disease progression or therapeutic targets. Case - control study design. Corneal ulcer tissue from 8 bacterial and 9 fungal ulcers was excised at the time of corneal transplantation surgery and immediately preserved in RNALater. Non-infected corneal tissue from 13 cadaver corneas were the control tissue. Transcriptome profile generated using Illumina HT12 v4 beadchips. Differential expression analysis was performed with pairwise comparisons: bacterial ulcers versus controls, fungal ulcers versus controls and bacterial versus fungal ulcers. Microarray results validated with RTqPCR.

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Correction: Alterations in the gut bacterial microbiome in fungal Keratitis patients.

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