Microbial changes in stool, saliva, serum, and urine before and after anti-TNF-α therapy in patients with inflammatory bowel diseases.

Inflammatory bowel diseases (IBD), including Crohn's disease and ulcerative colitis, are chronic immune-mediated intestinal inflammatory disorders associated with microbial dysbiosis at multiple sites, particularly the gut. Anti-tumor necrosis factor-α (TNF-α) agents are important treatments for IBD. We investigated whether microbiome changes at multiple sites can predict the effectiveness of such treatment in IBD. Stool, saliva, serum, and urine biosamples were collected from 19 IBD patients before (V1) and 3 months after (V2) anti-TNF-α treatment, and 19 healthy subjects (control). Microbiota analysis was performed using extracellular vesicles (EVs; all four sample types) and next-generation sequencing (NGS; stool and saliva). The stool, using NGS analysis, was the only sample type in which α-diversity differed significantly between the IBD and control groups at V1 and V2. Relative to non-responders, responders to anti-TNF-α treatment had significantly higher levels of Firmicutes (phylum), Clostridia (class), and Ruminococcaceae (family) in V1 stool, and Prevotella in V1 saliva. Non-responders had significantly higher V2 serum and urine levels of Lachnospiraceae than responders. Finally, Acidovorax caeni was detected in all V1 sample types in responders, but was not detected in non-responders. Microbiome changes at multiple sites may predict the effectiveness of anti-TNF-α treatment in IBD, warranting further research.

Measuring the absolute abundance of the microbiome by adding yeast containing 16S rRNA gene from a hyperthermophile.

High-throughput sequencing (HTS) of 16S rRNA gene amplicons provides compositional information regarding the microbial community, but not the absolute abundance of the bacteria. We aimed to develop a standardized method for quantifying the absolute abundance of bacteria in microbiome studies. To demonstrate the utility of our approach, we quantified the number of bacteria from the compositional data of the fecal and cecal microbiomes. The 16S rRNA gene of a hyperthermophile, Thermus aquaticus, was cloned into Pichia pastoris (yeast) genome, and an equivalent amount of the yeast was added to the stool and cecal samples of mice before DNA extraction. 16S rRNA gene library construction and HTS were performed after DNA extraction. The absolute abundances of bacteria were calculated using T. aquaticus reads. The average relative abundances of T. aquaticus in the five stool and five cecal samples were 0.95% and 0.33%, respectively, indicating that the number of bacteria in a cecum sample is 2.9 times higher than that in a stool sample. The method proposed for quantifying the absolute abundance of the bacterial population in this study is expected to overcome the limitation of showing only compositional data in most microbiome studies.

Laboratory Aspects of Donor Screening for Fecal Microbiota Transplantation at a Korean Fecal Microbiota Bank.

Fecal microbiota transplantation (FMT) is a widely accepted alternative therapy for Clostridioides difficile infection and other gastrointestinal disorders. Thorough donor screening is required as a safety control measure to minimize transmission of infectious agents in FMT. We report the donor screening process and outcomes at a fecal microbiota bank in Korea. From August 2017 to June 2020, the qualification of 62 individuals as FMT donors was evaluated using clinical assessment and laboratory tests. Forty-six (74%) candidates were excluded after clinical assessment; high body mass index (>25) was the most common reason for exclusion, followed by atopy, asthma, and allergy history. Four of the remaining 16 (25%) candidates failed to meet laboratory test criteria, resulting in a 19% qualification rate. FMT donor re-qualification was conducted monthly as an additional safety control measure, and only three (5%) candidates were eligible for repeated donation. As high prevalence of multidrug-resistant organisms (55%) and Helicobacter pylori (44%) were detected in qualified donors during the screening, a urea breath test was added to the existing protocol. The present results emphasize the importance of implementing a donor re-qualification system to minimize risk factors not identified during initial donor screening.

Comparative Microbiome Analysis of Three Species of Laboratory-Reared Periplaneta Cockroaches.

Cockroaches inhabit various habitats, which will influence their microbiome. Although the microbiome can be influenced by the diet and environmental factors, it can also differ between species. Therefore, we conducted 16S rDNAtargeted high-throughput sequencing to evaluate the overall bacterial composition of the microbiomes of 3 cockroach species, Periplaneta americana, P. japonica, and P. fuliginosa, raised in laboratory for several generations under the same conditions. The experiments were conducted using male adult cockroaches. The number of operational taxonomic units (OTUs) was not significantly different among the 3 species. With regard to the Shannon and Pielou indexes, higher microbiome values were noted in P. americana than in P. japonica and P. fuliginosa. Microbiome composition was also evaluated, with endosymbionts accounting for over half of all OTUs in P. japonica and P. fuliginosa. Beta diversity analysis further showed that P. japonica and P. fuliginosa had similar microbiome composition, which differed from that of P. americana. However, we also identified that P. japonica and P. fuliginosa host distinct OTUs. Thus, although microbiome compositions may vary based on multiple conditions, it is possible to identify distinct microbiome compositions among different Periplaneta cockroach species, even when the individuals are reared under the same conditions.