Introduction. Colorectal cancer (CRC) is one of the most common cancers worldwide. Multiple risk factors are involved in CRC development, including age, genetics, lifestyle, diet and environment. Of these, the role of the gut microbiota in cancer biology is increasingly recognized. Hypothesis/Gap Statement. Micro-organisms have been widely detected in stool samples, but few mucosal samples have been detected and sequenced in depth. Aim. Analysis of cultured mucosal bacteria from colorectal cancer and adjacent normal mucosal tissues with metagenomics sequencing. Methodology. Twenty-eight paired tumour and non-tumour tissues from 14 patients undergoing surgery for CRC were analysed. We removed the influence of eukaryotic cells via culture. The composition of mucosal microbiota in intestinal mucosa were detected and analysed with metagenomic sequencing. Results. Compared with non-cultured mucosal sample, 80 % bacteria species could be detected after culture. Moreover, after culture, additional 30 % bacteria could be detected, compared with non-cultured samples. Since after culture it was difficult to estimate the original abundance of microbiome, we focused on the identification of the CRC tissue-specific species. There were 298 bacterial species, which could only be cultured and detected in CRC tissues. Myroides odoratimimus and Cellulophaga baltica could be isolated from all the tumour samples of 14 CRC patients, suggesting that these species may be related to tumour occurrence and development. Further functional analysis indicated that bacteria from CRC tissues showed more active functions, including basic metabolism, signal transduction and survival activities. Conclusion. We used a new method based on culture to implement information on prokaryotic taxa, and related functions, which samples were from colorectal tissues. This method is suitable for removing eukaryotic contamination and detecting micro-organisms from other tissues.

Introduction. The human oocyte microenvironment is follicular fluid, which is important for follicle growth, ovulation and maturation of the oocyte. The micro-organisms present in follicular fluid could be a predictor of in vitro fertilization outcomes. Hypothesis/Gap Statement. Women with follicular fluid colonized with micro-organisms can be asymptomatic, but the presence of some genera in the follicular fluid correlates with in vitro fertilization. Aim. To confirm the existence of micro-organisms in follicular fluid, and to profile the micro-organisms present in follicular fluid sampled from women undergoing in vitro fertilization with different outcomes. Methodology. Women undergoing in vitro fertilization (n=163) were divided into different subgroups according to their in vitro fertilization outcomes. Their follicular fluid samples were collected, and among them, 157 samples were analysed by 16S rDNA sequencing, and 19 samples were analysed using culturomics. Results. The culturomics results suggested that the 19 follicular fluid samples were not sterile. The isolation rates for Streptococcus , Finegoldia and Peptoniphilus were >50 % in the 19 samples. Linear discriminant analysis effect size analysis showed differential bacteria abundance according to the pregnancy rate, the rate of normal fertilization, the rate of high-quality embryos and the rate of available oocytes. The sequencing results showed that micro-organisms could be detected in all 157 samples. Pseudomonas , Lactobacillus , Comamonas , Streptococcus and Acinetobacter were detected in all of the samples, but with a wide range of relative abundance. Pseudomonas , Lactobacillus , Ralstonia and Vibrio constituted a notable fraction of the microbiota. Conclusions. Follicular fluid is not sterile. Micro-organisms in follicular fluid could be a predictor of in vitro fertilization outcomes.