Description of Faecalibacterium wellingii sp. nov. and two Faecalibacterium taiwanense strains, aiding to the reclassification of Faecalibacterium species.

OBJECTIVES: The genus Faecalibacterium is one of the most important butyrate producers in the human intestinal tract and has been widely linked to health. Recently, several different species have been described, but still more phylogroups have been identified, suggesting that additional species may exist. Four strains HTF-FT, HTF-128, HTF-75H and HTF-76H, representing two different phylogenetic clusters, are evaluated in this study. METHODS: Phylogenomic analysis was performed using whole-genome sequences and 16S rRNA gene sequences. Chemotaxonomic analysis was done based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Physiological and phenotypical characteristics of these strains were also determined. All characteristics of these strains were compared with other validly published species within the genus Faecalibacterium. RESULTS: On a genomic level, the strains HTF-FT and HTF-128 shared an average nucleotide identity (ANI) of <95.0 % and digital DNA-DNA hybridization (dDDH) of <70.0 with other Faecalibacterium species, while between HTF-FT and HTF-128 the ANI-value was 97.18 % and the dDDH was 76.8 %. HTF-75H and HTF-76H had an ANI and dDDH value of 100 % (99.96 %) and 100 % (99.99 %) respectively. Both HTF-75H and HTF-76H were closely related to Faecalibacterium taiwanense HLW78T. 16S rRNA gene and chemotaxonomic analysis were in accordance with the genomic data, confirming that HTF-FT and HTF-128 represent a novel Faecalibacterium species and HTF-75H and HTF-76H belong to F. taiwanense. CONCLUSIONS: Faecalibacterium strains HTF-FT (=DSM 117771T = NCIMB 15531T) and HTF-128 represent a novel species. The name Faecalibacterium wellingii with HTF-FT as type strain is proposed. Two novel isolates HTF-75H (=DSM 17770 = NCIMB 15530) and HTF-76H are described in this study and belong to the recently described Faecalibacterium taiwanense.

Faecalibacterium taiwanense sp. nov., isolated from human faeces.

Two Gram-stain-negative, straight rods, non-motile, asporogenous, catalase-negative and obligately anaerobic butyrate-producing strains, HLW78T and CYL33, were isolated from faecal samples of two healthy Taiwanese adults. Phylogenetic analyses of 16S rRNA and DNA mismatch repair protein MutL (mutL) gene sequences revealed that these two novel strains belonged to the genus Faecalibacterium. On the basis of 16S rRNA and mutL gene sequence similarities, the type strains Faecalibacterium butyricigenerans AF52-21T(98.3-98.1 % and 79.0-79.5 % similarity), Faecalibacterium duncaniae A2-165T(97.8-97.9 % and 70.9-80.1 %), Faecalibacterium hattorii APC922/41-1T(97.1-97.3 % and 80.3-80.5 %), Faecalibacterium longum CM04-06T(97.8-98.0% and 78.3 %) and Faecalibacterium prausnitzii ATCC 27768T(97.3-97.4 % and 82.7-82.9 %) were the closest neighbours to the novel strains HLW78T and CYL33. Strains HLW78T and CYL33 had 99.4 % both the 16S rRNA and mutL gene sequence similarities, 97.9 % average nucleotide identity (ANI), 96.3 % average amino acid identity (AAI), and 80.5 % digital DNA-DNA hybridization (dDDH) values, indicating that these two strains are members of the same species. Phylogenomic tree analysis indicated that strains HLW78T and CYL33 formed an independent robust cluster together with F. prausnitzii ATCC 27768T. The ANI, AAI and dDDH values between strain HLW78T and its closest neighbours were below the species delineation thresholds of 77.6-85.1 %, 71.4-85.2 % and 28.3-30.9 %, respectively. The two novel strains could be differentiated from the type strains of their closest Faecalibacterium species based on their cellular fatty acid compositions, which contained C18 : 1 ω7c and lacked C15 : 0 and C17 : 1 ω6c, respectively. Phenotypic, chemotaxonomic and genotypic test results demonstrated that the two novel strains HLW78T and CYL33 represented a single, novel species within the genus Faecalibacterium, for which the name Faecalibacterium taiwanense sp. nov. is proposed. The type strain is HLW78T (=BCRC 81397T=NBRC 116372T).

Understanding of the Site-Specific Microbial Patterns towards Accurate Identification for Patients with Diarrhea-Predominant Irritable Bowel Syndrome.

Fecal microbial community could not fully represent the intestinal microbial community. However, most studies analyzing diarrhea-dominant irritable bowel syndrome (IBS-D) were mainly based on fecal samples. We aimed to characterize the IBS-D microbial community patterns using samples at multiple intestinal sites. This study recruited 74 IBS-D patients and 20 healthy controls (HC). 22.34%, 8.51%, 14.89%, and 54.26% of them contributed to one, two, three, and four sites: duodenal mucosa (DM), duodenal lumen (DL), rectal mucosa (RM), and rectal lumen (RL) of intestinal samples, respectively. Then 16S rRNA gene analysis was performed on these 283 samples. The result showed that IBS-D microbial communities have specific patterns at each intestinal site differing from that of HC. Across hosts and sites, Bacillus, Burkholderia, and Faecalibacterium were the representative genera in duodenum of IBS-D, duodenum of HC, and rectum of HC, respectively. Samples from mucosa and lumen in rectum were highly distinguishable, regardless of IBS-D and HC. Additionally, IBS-D patients have lower microbial co-abundance network connectivity. Moreover, RM site-specific biomarker: Bacteroides used alone or together with Prevotella and Oscillospira in RM showed outstanding performance in IBS-D diagnosis. Furthermore, Bacteroides and Prevotella in RM were strongly related to the severity of abdominal pain, abdominal discomfort, and bloating in IBS-D patients. In summary, this study also confirmed fecal microbial community could not fully characterize intestinal microbial communities. Among these site-specific microbial communities, RM microbial community would be more applicable in the diagnosis of IBS-D. IMPORTANCE Microbial community varied from one site to another along the gastrointestinal tract, but current studies about intestinal microbial community in IBS-D were mainly based on fecal samples. Based on 283 intestinal samples collected from DM, DL, RM, and RL of HC and IBS-D, we found different intestinal sites had their site-specific microbial patterns in IBS-D. Notably, RM site-specific microbes Bacteroides, Prevotella, and Oscillospira could be used to discriminate IBS-D from HC accurately. Our findings could help clinicians realize the great potential of the intestinal microbial community in RM for better diagnosis of IBS-D patients.

The multiple effects of fecal microbiota transplantation on diarrhea-predominant irritable bowel syndrome (IBS-D) patients with anxiety and depression behaviors.

BACKGROUND: Anxiety and depression are complications in Irritable bowel syndrome (IBS) patients. In this study, we recruited 18 IBS patients with mild-modest anxiety and depression behaviors, and after the screening, we defined the FMT treatment group (n = 9) and the control group (n = 9). The IBS symptom severity scale (IBS-SSS), Hamilton Anxiety Rating Scale (HAM-A), Hamilton Depression Rating Scale (HAM-D), Irritable Bowel Syndrome Quality of Life (IBS-QOL) and Bristol stool scale (BSS) were evaluated one week before FMT (baseline), one-week-, one-month-, two-month-, and three-month-following FMT. Meanwhile, we determined the SCFAs in the patient's feces and serum and continued the metagenomic analysis of the microorganisms in the patient's feces. RESULTS: The results showed that the patient's anxiety and depression behavior gradually improved with FMT treatment. Moreover, the illness and quality of life had also been relieved significantly. The content of isovaleric acid and valeric acid was significantly reduced in the FMT group compared to the Col group. Metagenomic analysis showed that FMT treatment decreased the abundance of Faecalibacterium, Eubacterium and Escherichia. From KEGG functional analysis, we confirmed that the top five abundant pathways were "bacterial chemotaxis, "flagellar assembly", "glycine, serine and threonine metabolism", "apoptosis", and "bacterial invasion of epithelial cells". CONCLUSIONS: FMT treatment can effectively alleviate the anxiety and depression behaviors of IBS-D patients and reduce the IBS-SSS score, indicating that FMT can improve patients' symptoms. The high throughput sequencing results show that Bifidobacterium and Escherichia play the most critical role in the formation and recovery of IBS-D patients. The GC/MS data indicated that faeces isovaleric acid and valeric acid might be more suitable as a metabolic indicator of IBS-D remission. Trial registration ChiCTR, ChiCTR1900024924, Registered 3 August 2019, https://www.chictr.org.cn/showproj.aspx?proj=41676 .

Detection of cell-free microbial DNA using a contaminant-controlled analysis framework.

BACKGROUND: The human microbiome plays an important role in cancer. Accumulating evidence indicates that commensal microbiome-derived DNA may be represented in minute quantities in the cell-free DNA of human blood and could possibly be harnessed as a new cancer biomarker. However, there has been limited use of rigorous experimental controls to account for contamination, which invariably affects low-biomass microbiome studies. RESULTS: We apply a combination of 16S-rRNA-gene sequencing and droplet digital PCR to determine if the specific detection of cell-free microbial DNA (cfmDNA) is possible in metastatic melanoma patients. Compared to matched stool and saliva samples, the absolute concentration of cfmDNA is low but significantly above the levels detected from negative controls. The microbial community of plasma is strongly influenced by laboratory and reagent contaminants introduced during the DNA extraction and sequencing processes. Through the application of an in silico decontamination strategy including the filtering of amplicon sequence variants (ASVs) with batch dependent abundances and those with a higher prevalence in negative controls, we identify known gut commensal bacteria, such as Faecalibacterium, Bacteroides and Ruminococcus, and also other uncharacterised ASVs. We analyse additional plasma samples, highlighting the potential of this framework to identify differences in cfmDNA between healthy and cancer patients. CONCLUSIONS: Together, these observations indicate that plasma can harbour a low yet detectable level of cfmDNA. The results highlight the importance of accounting for contamination and provide an analytical decontamination framework to allow the accurate detection of cfmDNA for future biomarker studies in cancer and other diseases.

Gut microbial composition difference between pediatric ALL survivors and siblings.

Acute lymphoblastic leukemia (ALL) is the most common childhood cancer with high cure rates leading to rising numbers of long-term survivors. Adult survivors of childhood ALL are at increased risk of obesity, cardiovascular disease, and other chronic illnesses. We hypothesize that ALL therapy is associated with long-term gut microbiome alterations that contribute to predisposition to chronic medical conditions. We conducted a pilot study to test whether differences can be detected between stool microbiota of pediatric ALL survivors and their siblings. Stool samples were collected from 38 individuals under age 19 who were at least 1 year after completion of therapy for ALL. Stool samples collected from 16 healthy siblings served as controls. 16S ribosomal RNA gene sequencing was performed on the stool samples. Comparing microbiota of survivors to sibling controls, no statistically significant differences were found in alpha or beta diversity. However, among the top 10 operational taxonomic units (OTUs) from component 1 in sparse partial least squares discriminant analysis (sPLS-DA) with different relative abundance in survivors versus siblings, OTUs mapping to the genus Faecalibacterium were depleted in survivors. Differences in gut microbial composition were found between pediatric survivors of childhood ALL and their siblings. Specifically, the protective Faecalibacterium is depleted in survivors, which is reminiscent of gut microbiota alteration found in adult survivors of childhood ALL and reported in obesity, suggesting that microbiota alterations in pediatric ALL survivors start in childhood and may play a role in predisposition to chronic illness in later years of survivorship.

Alterations in gut bacterial and fungal microbiomes are associated with bacterial Keratitis, an inflammatory disease of the human eye.

Dysbiosis, or imbalance in the gut microbiome, has been implicated in auto-immune, inflammatory, neurological diseases as well as in cancers. More recently it has also been shown to be associated with ocular diseases. In the present study, the association of gut microbiome dysbiosis with bacterial Keratitis, an inflammatory eye disease which significantly contributes to corneal blindness, was investigated. Bacterial and fungal gut microbiomes were analysed using fecal samples of healthy controls (HC, n = 21) and bacterial Keratitis patients (BK, n = 19). An increase in abundance of several antiinflammatory organisms including Dialister, Megasphaera, Faecalibacterium, Lachnospira, Ruminococcus and Mitsuokella and members of Firmicutes, Veillonellaceae, Ruminococcaceae and Lachnospiraceae was observed in HC compared to BK patients in the bacterial microbiome. In the fungal microbiome, a decrease in the abundance of Mortierella, Rhizopus, Kluyveromyces, Embellisia and Haematonectria and an increase in the abundance of pathogenic fungi Aspergillus and Malassezia were observed in BK patients compared to HC. In addition, heatmaps, PCoA plots and inferred functional profiles also indicated significant variations between the HC and BK microbiomes, which strongly suggest dysbiosis in the gut microbiome of BK patients. This is the first study demonstrating the association of gut microbiome with the pathophysiology of BK and thus supports the gut-eye axis hypothesis. Considering that Keratitis affects about 1 million people annually across the globe, the data could be the basis for developing alternate strategies for treatment like use of probiotics or fecal transplantation to restore the healthy microbiome as a treatment protocol for Keratitis.

Description of two novel members of the family Erysipelotrichaceae: Ileibacterium valens gen. nov., sp. nov. and Dubosiella newyorkensis, gen. nov., sp. nov., from the murine intestine, and emendation to the description of Faecalibaculum rodentium.

To better characterize murine intestinal microbiota, a large number (187) of Gram-positive-staining, rod- and coccoid-shaped, and facultatively or strictly anaerobic bacteria were isolated from small and large intestinal contents from mice. Based on 16S rRNA gene sequencing, a total 115 isolates formed three phylogenetically distinct clusters located within the family Erysipelotrichaceae. Group 1, as represented by strain NYU-BL-A3T, was most closely related to Allobaculum stercoricanis, with 16S rRNA gene sequence similarity values of 87.7 %. A second group, represented by NYU-BL-A4T, was most closely related to Faecalibaculum rodentium, with 86.6 % 16S rRNA gene sequence similarity. A third group had a nearly identical 16S rRNA gene sequence (99.9 %) compared with the recently described Faecalibaculum rodentium, also recovered from a laboratory mouse; however, this strain had a few differences in biochemical characteristics, which are detailed in an emended description. The predominant (>10 %) cellular fatty acids of strain NYU-BL-A3T were C16 : 0 and C18 : 0, and those of strain NYU-BL-A4T were C10 : 0, C16 : 0, C18 : 0 and C18 : 1ω9c. The two groups could also be distinguished by multiple biochemical reactions, with the group represented by NYU-BL-A4T being considerably more active. Based on phylogenetic, biochemical and chemotaxonomic criteria, two novel genera are proposed, Ileibacterium valens gen. nov., sp. nov. with NYU-BL-A3T (=ATCC TSD-63T=DSM 103668T) as the type strain and Dubosiella newyorkensis gen. nov., sp. nov. with NYU-BL-A4T (=ATCC TSD-64T=DSM 103457T) as the type strain.

Microbiological profiles of sputum and gastric juice aspirates in Cystic Fibrosis patients.

Gastro-Oesophageal Reflux (GOR) is a key problem in Cystic Fibrosis (CF), but the relationship between lung and gastric microbiomes is not well understood. We hypothesised that CF gastric and lung microbiomes are related. Gastric and sputum cultures were obtained from fifteen CF patients receiving percutaneous endoscopic gastrostomy feeding. Non-CF gastric juice data was obtained through endoscopy from 14 patients without lung disease. Bacterial and fungal isolates were identified by culture. Molecular bacterial profiling used next generation sequencing (NGS) of the 16S rRNA gene. Cultures grew bacteria and/or fungi in all CF gastric juice and sputa and in 9/14 non-CF gastric juices. Pseudomonas aeruginosa(Pa) was present in CF sputum in 11 patients, 4 had identical Pa strains in the stomach. NGS data from non-CF gastric juice samples were significantly more diverse compared to CF samples. NGS showed CF gastric juice had markedly lower abundance of normal gut bacteria; Bacteroides and Faecalibacterium, but increased Pseudomonas compared with non-CF. Multivariate partial least squares discriminant analysis demonstrated similar bacterial profiles of CF sputum and gastric juice samples, which were distinct from non-CF gastric juice. We provide novel evidence suggesting the existence of an aerodigestive microbiome in CF, which may have clinical relevance.