경제동물 분변 균총 연구를 위한 시료 채취 조건의 평가
Evaluating Sampling Condition from Farm Animal for Fecal Microbiome Study
0
조회
0
다운로드
0
인용
0
참고문헌
키워드
초록
When researchers performed intestinal microbiota analysis using fecal samples, sample preparation processes are not standardized as of now. In particular, contrary to lab conditions, there are various factors hard to be controlled when sampling livestock feces on the farm. In this study, we wanted to know the influences of sample preparation conditions on fecal microbiota. We designed an experiment considering various sample preparation conditions (sample origin: rectum and ground; transporting condition: lysis buffer-treated, dry ice, ice, and room temperature; delayed times to extract DNA: 0, 1, 6, and 24 h) that can occur during microbiota analysis using cattle feces. First, we investigated the influences of sample origin, and microbial diversity (observed OTUs, p<0.001) was significantly higher when fecal samples were obtained from the ground than from the rectum and the principal coordinates analysis plot showed that samples were divided into two groups by origin. When we investigated the influences of transporting and storage conditions, observed OTUs (p<0.05) was significantly higher when samples were transported at room temperature than other conditions, and microbiota was affected by transporting and storage conditions. Finally, we investigated the effects of delayed time before DNA extraction from frozen fecal samples on microbial composition. Although this factor did not have a significant influence on microbial diversity, multidimensional scaling revealed its impact on microbial composition. Our findings will contribute to establishing an optimal procedure for microbiota analysis using farm-housing livestock feces.
Abstract
When researchers performed intestinal microbiota analysis using fecal samples, sample preparation processes are not standardized as of now. In particular, contrary to lab conditions, there are various factors hard to be controlled when sampling livestock feces on the farm. In this study, we wanted to know the influences of sample preparation conditions on fecal microbiota. We designed an experiment considering various sample preparation conditions (sample origin: rectum and ground; transporting condition: lysis buffer-treated, dry ice, ice, and room temperature; delayed times to extract DNA: 0, 1, 6, and 24 h) that can occur during microbiota analysis using cattle feces. First, we investigated the influences of sample origin, and microbial diversity (observed OTUs, p<0.001) was significantly higher when fecal samples were obtained from the ground than from the rectum and the principal coordinates analysis plot showed that samples were divided into two groups by origin. When we investigated the influences of transporting and storage conditions, observed OTUs (p<0.05) was significantly higher when samples were transported at room temperature than other conditions, and microbiota was affected by transporting and storage conditions. Finally, we investigated the effects of delayed time before DNA extraction from frozen fecal samples on microbial composition. Although this factor did not have a significant influence on microbial diversity, multidimensional scaling revealed its impact on microbial composition. Our findings will contribute to establishing an optimal procedure for microbiota analysis using farm-housing livestock feces.
참고문헌 (35)
- Ahmed, I., "Proposal of Lysinibacillus boronitolerans gen. nov. sp. nov., and transfer of Bacillus fusiformis to Lysinibacillus fusiformis comb. nov. and Bacillus sphaericus to Lysinibacillus sphaericus comb. nov", Int. J. Syst. Evol. Microbiol, 57(5), pp.1117~1125 (2007)
- Ainonen, S., "Antibiotics at birth and later antibiotic courses : effects on gut microbiota", Pediatr. Res, 91(1), pp.154~162 (2022)
- Arumugam, M., "Enterotypes of the human gut microbiome", Nature, 473(7346), pp.174~180 (2011)
- Bahl, M. I., "Freezing fecal samples prior to DNA extraction affects the Firmicutes to Bacteroidetes ratio determined by downstream quantitative PCR analysis", FEMS Microbiol. Lett, 329(2), pp.193~197 (2012)
- Bergmann, G. T., "Seasonal Shifts in Diet and Gut Microbiota of the American Bison (Bison bison)", PLoS One, 10(11), pp.e0142409~ (2015)
- Birkl, P., "Differences in cecal microbiome of selected high and low feather-pecking laying hens", Poult. Sci, 97(9), pp.3009~3014 (2018)
- Caporaso, J. G., "PyNAST : a flexible tool for aligning sequences to a template alignment", Bioinformatics, 26(2), pp.266~267 (2010)
- Dankwa, A. S., "Bacterial communities in the rumen and feces of lactating Holstein dairy cows are not affected when fed reduced-fat dried distillers' grains with solubles", Animal, 15(7), pp.100281~ (2021)
- de Brito, L. C. N., "The apical root canal system microbial communities determined by next-generation sequencing", Sci. Rep, 10(1), pp.1~8 (2020)
- DeSantis, T. Z., "Greengenes, a chimera-checked 16S rRNA gene database and workbench compatible with ARB", Appl. Environ. Microbiol, 72(7), pp.5069~5072 (2006)
- Drew, G. C., "Microbial evolution and transitions along the parasite-mutualist continuum", Nat. Rev. Microbiol, 19(10), pp.623~638 (2021)
- Edgar, R. C., "Search and clustering orders of magnitude faster than BLAST", Bioinformatics, 26(19), pp.2460~2461 (2010)
- Garrett, W. S., "Cancer and the microbiota", Science, 348(6230), pp.80~86 (2015)
- Han, G. G., "Relationship between the microbiota in different sections of the gastrointestinal tract, and the body weight of broiler chickens", Springerplus, 5(1), pp.1~9 (2016)
- Han, G. G., "Tracing of the fecal microbiota of commercial pigs at five growth stages from birth to shipment", Sci. Rep, 8(1), pp.1~9 (2018)
- Huang, S., "The day-to-day stability of the ruminal and fecal microbiota in lactating dairy cows", Microbiologyopen, 9(5), pp.e990~ (2020)
- Jin, J., "High-throughput identification and quantification of single bacterial cells in the microbiota", Nat. Commun, 13(1), pp.863~ (2022)
- Kaltenegger, A., "Feeding dairy cows bakery by-products enhanced nutrient digestibility, but affected fecal microbial composition and pH in a dose-dependent manner", J. Dairy. Sci, 104(7), pp.7781~7793 (2021)
- Kennedy, K. M., "Fetal meconium does not have a detectable microbiota before birth", Nat. Microbiol, 6(7), pp.865~873 (2021)
- Kim, H. S., "Longitudinal evaluation of fecal microbiota transplantation for ameliorating calf diarrhea and improving growth performance", Nat. Commun, 12(1), pp.161~ (2021)
- Krishnamurthi, S., "Re-examination of the taxonomic position of Bacillus silvestris Rheims et al. 1999 and proposal to transfer it to Solibacillus gen. nov as Solibacillus silvestris comb. nov", Int. J. Syst. Evol. Micr, 59, pp.1054~1058 (2009)
- Lauber, C. L., "Effect of storage conditions on the assessment of bacterial community structure in soil and human-associated samples", FEMS Microbiol. Lett, 307(1), pp.80~86 (2010)
- Magoc, T., "FLASH : fast length adjustment of short reads to improve genome assemblies", Bioinformatics, 27(21), pp.2957~2963 (2011)
- McCormack, U. M., "Exploring a possible link between the intestinal microbiota and feed efficiency in pigs", Appl. Environ. Microb, 83(15), pp.e00380~17 (2017)
- Mote, R. S., "Response of Beef Cattle Fecal Microbiota to Grazing on Toxic Tall Fescue", Appl. Environ. Microb, 85(15), pp.e00032~19 (2019)
- Nicco, C., "From donor to patient : collection, preparation and cryopreservation of fecal samples for fecal microbiota transplantation", Diseases, 8(2), pp.9~ (2020)
- 오주경, "Protective effects of Bacillus subtilis against Salmonella infection in the microbiome of Hy-Line Brown layers", Animal Bioscience, 30(9), pp.1332~1339 (2017), 아세아·태평양축산학회[DOI]
- Qiao, H. X., "Astragalus affects fecal microbial composition of young hens as determined by 16S rRNA sequencing", AMB Express, 8(1), pp.70~ (2018)
- Ravussin, Y., "Responses of gut microbiota to diet composition and weight loss in lean and obese mice", Obesity, 20(4), pp.738~747 (2012)
- Song, E. S., "Comparison of Fecal microbiota between german holstein dairy cows with and without left-sided displacement of the abomasum", J. Clin. Microbiol, 54(4), pp.1140~1143 (2016)
- Thaiss, C. A., "The microbiome and innate immunity", Nature, 535(7610), pp.65~74 (2016)
- Uchiyama, J., "Examination of the fecal microbiota in dairy cows infected with bovine leukemia virus", Vet. Microbiol, 240, pp.108547~ (2020)
- Varga, A., "How to apply FMT more effectively, conveniently and flexible-A comparison of FMT methods", Front. Cell. Infect. Mi, 11, pp.494~ (2021)
- Weinroth, M. D., "Investigation of tylosin in feed of feedlot cattle and effects on liver abscess prevalence, and fecal and soil microbiomes and resistomes", J. Anim. Sci, 97(11), pp.4567~4578 (2019)
- Yatsunenko, T., "Human gut microbiome viewed across age and geography", Nature, 486(7402), pp.222~227 (2012)