Complex gut microbiota increases chickens' resistance to enteric pathogens. However, the principles of this phenomenon are not understood in detail. One of the possibilities for how to decipher the role of gut microbiota in chickens' resistance to enteric pathogens is to systematically characterise the gene expression of individual gut microbiota members colonising the chicken caecum. To reach this aim, newly hatched chicks were inoculated with bacterial species whose whole genomic sequence was known. Total protein purified from the chicken caecum was analysed by mass spectrometry, and the obtained spectra were searched against strain-specific protein databases generated from known genomic sequences. Campylobacter jejuni, Phascolarctobacterium sp. and Sutterella massiliensis did not utilise carbohydrates when colonising the chicken caecum. On the other hand, Bacteroides, Mediterranea, Marseilla, Megamonas, Megasphaera, Bifidobacterium, Blautia, Escherichia coli and Succinatimonas fermented carbohydrates. C. jejuni was the only motile bacterium, and Bacteroides mediterraneensis expressed the type VI secretion system. Classification of in vivo expression is key for understanding the role of individual species in complex microbial populations colonising the intestinal tract. Knowledge of the expression of motility, the type VI secretion system, and preference for carbohydrate or amino acid fermentation is important for the selection of bacteria for defined competitive exclusion products.

Veterinary Research Institute CZ6210 Brno Czech Republic

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Number of Chickens Worldwide from 1990 to 2022. [(accessed on 20 August 2023)]. Available online: https://www.statista.com/statistics/263962/number-of-chickens-worldwide-since-1990.

Kubasova T., Kollarcikova M., Crhanova M., Karasova D., Cejkova D., Sebkova A., Matiasovicova J., Faldynova M., Pokorna A., Cizek A., et al. Contact with adult hen affects development of caecal microbiota in newly hatched chicks. PLoS ONE. 2019;14:e0212446. doi: 10.1371/journal.pone.0212446. PubMed DOI PMC

Rantala M., Nurmi E. Prevention of the growth of Salmonella infantis in chicks by the flora of the alimentary tract of chickens. Br. Poult. Sci. 1973;14:627–630. doi: 10.1080/00071667308416073. PubMed DOI

Nurmi E., Rantala M. New aspects of Salmonella infection in broiler production. Nature. 1973;241:210–211. doi: 10.1038/241210a0. PubMed DOI

Methner U., Barrow P.A., Martin G., Meyer H. Comparative study of the protective effect against Salmonella colonisation in newly hatched SPF chickens using live, attenuated Salmonella vaccine strains, wild-type Salmonella strains or a competitive exclusion product. Int. J. Food Microbiol. 1997;35:223–230. doi: 10.1016/S0168-1605(96)01236-6. PubMed DOI

Medvecky M., Cejkova D., Polansky O., Karasova D., Kubasova T., Cizek A., Rychlik I. Whole genome sequencing and function prediction of 133 gut anaerobes isolated from chicken caecum in pure cultures. BMC Genom. 2018;19:561. doi: 10.1186/s12864-018-4959-4. PubMed DOI PMC

Glendinning L., Stewart R.D., Pallen M.J., Watson K.A., Watson M. Assembly of hundreds of novel bacterial genomes from the chicken caecum. Genome Biol. 2020;21:34. doi: 10.1186/s13059-020-1947-1. PubMed DOI PMC

Sergeant M.J., Constantinidou C., Cogan T.A., Bedford M.R., Penn C.W., Pallen M.J. Extensive microbial and functional diversity within the chicken cecal microbiome. PLoS ONE. 2014;9:e91941. doi: 10.1371/journal.pone.0091941. PubMed DOI PMC

Langille M.G., Zaneveld J., Caporaso J.G., McDonald D., Knights D., Reyes J.A., Clemente J.C., Burkepile D.E., Vega Thurber R.L., Knight R., et al. Predictive functional profiling of microbial communities using 16S rRNA marker gene sequences. Nat. Biotechnol. 2013;31:814–821. doi: 10.1038/nbt.2676. PubMed DOI PMC

Zhong Z.K., Wang C., Zhang H.D., Mi J.D., Liang J.B., Liao X.D., Wu Y.B., Wang Y. Sodium butyrate reduces ammonia emissions through glutamate metabolic pathways in cecal microorganisms of laying hens. Ecotox. Environ. Safe. 2022;233:113299. doi: 10.1016/j.ecoenv.2022.113299. PubMed DOI

Polansky O., Sekelova Z., Faldynova M., Sebkova A., Sisak F., Rychlik I. Important metabolic pathways and biological processes expressed by chicken cecal microbiota. Appl. Environ. Microbiol. 2015;82:1569–1576. doi: 10.1128/AEM.03473-15. PubMed DOI PMC

Kubasova T., Kollarcikova M., Crhanova M., Karasova D., Cejkova D., Sebkova A., Matiasovicova J., Faldynova M., Sisak F., Babak V., et al. Gut anaerobes capable of chicken caecum colonisation. Microorganisms. 2019;7:597. doi: 10.3390/microorganisms7120597. PubMed DOI PMC

Papouskova A., Rychlik I., Harustiakova D., Cizek A. Research Note: A mixture of Bacteroides spp. and other probiotic intestinal anaerobes reduces colonization by pathogenic E. coli strain O78:H4-ST117 in newly hatched chickens. Poult. Sci. 2023;102:102529. doi: 10.1016/j.psj.2023.102529. PubMed DOI PMC

Kalmokoff M.L., Austin J.W., Cyr T.D., Hefford M.A., Teather R.M., Selinger L.B. Physical and genetic characterization of an outer-membrane protein (OmpM1) containing an N-terminal S-layer-like homology domain from the phylogenetically Gram-positive gut anaerobe Mitsuokella multacida. Anaerobe. 2009;15:74–81. doi: 10.1016/j.anaerobe.2009.01.001. PubMed DOI

Takatsuka Y., Kamio Y. Molecular dissection of the Selenomonas ruminantium cell envelope and lysine decarboxylase involved in the biosynthesis of a polyamine covalently linked to the cell wall peptidoglycan layer. Biosci. Biotechnol. Biochem. 2004;68:1–19. doi: 10.1271/bbb.68.1. PubMed DOI

Coyne M.J., Roelofs K.G., Comstock L.E. Type VI secretion systems of human gut Bacteroidales segregate into three genetic architectures, two of which are contained on mobile genetic elements. BMC Genom. 2016;17:58. doi: 10.1186/s12864-016-2377-z. PubMed DOI PMC

Fandi K.G., Ghazali H.M., Yazid A.M., Raha A.R. Purification and N-terminal amino acid sequence of fructose-6-phosphate phosphoketolase from Bifidobacterium longum BB536. Lett. Appl. Microbiol. 2001;32:235–239. doi: 10.1046/j.1472-765X.2001.00895.x. PubMed DOI

Raut M.P., Couto N., Karunakaran E., Biggs C.A., Wright P.C. Deciphering the unique cellulose degradation mechanism of the ruminal bacterium Fibrobacter succinogenes S85. Sci. Rep. 2019;9:16542. doi: 10.1038/s41598-019-52675-8. PubMed DOI PMC

Li R.M., Feng Y.G., Liu S.Y., Qi K.A., Cui Q., Liu Y.J. Inducing effects of cellulosic hydrolysate components of lignocellulose on cellulosome synthesis in Clostridium thermocellum. Microb. Biotechnol. 2018;11:905–916. doi: 10.1111/1751-7915.13293. PubMed DOI PMC

Videnska P., Sedlar K., Lukac M., Faldynova M., Gerzova L., Cejkova D., Sisak F., Rychlik I. Succession and replacement of bacterial populations in the caecum of egg laying hens over their whole life. PLoS ONE. 2014;9:e115142. doi: 10.1371/journal.pone.0115142. PubMed DOI PMC

Yu X., Niu S., Tie K., Zhang Q., Deng H., Gao C., Yu T., Lei L., Feng X. Characteristics of the intestinal flora of specific pathogen free chickens with age. Microb. Pathog. 2019;132:325–334. PubMed

Gao P., Ma C., Sun Z., Wang L., Huang S., Su X., Xu J., Zhang H. Feed-additive probiotics accelerate yet antibiotics delay intestinal microbiota maturation in broiler chicken. Microbiome. 2017;5:91. doi: 10.1186/s40168-017-0315-1. PubMed DOI PMC

Stanley D., Geier M.S., Hughes R.J., Denman S.E., Moore R.J. Highly variable microbiota development in the chicken gastrointestinal tract. PLoS ONE. 2013;8:e84290. doi: 10.1371/journal.pone.0084290. PubMed DOI PMC

Kubasova T., Seidlerova Z., Rychlik I. Ecological adaptations of gut microbiota members and their consequences for use as a new generation of probiotics. Int. J. Mol. Sci. 2021;22:5471. doi: 10.3390/ijms22115471. PubMed DOI PMC

Karasova D., Faldynova M., Matiasovicova J., Sebkova A., Crhanova M., Kubasova T., Seidlerova Z., Prikrylova H., Volf J., Zeman M., et al. Host species adaptation of obligate gut anaerobes is dependent on their environmental survival. Microorganisms. 2022;10:1085. doi: 10.3390/microorganisms10061085. PubMed DOI PMC

Avershina E., Larsen M.G., Aspholm M., Lindback T., Storro O., Oien T., Johnsen R., Rudi K. Culture dependent and independent analyses suggest a low level of sharing of endospore-forming species between mothers and their children. Sci. Rep. 2020;10:1832. doi: 10.1038/s41598-020-58858-y. PubMed DOI PMC

Nayfach S., Rodriguez-Mueller B., Garud N., Pollard K.S. An integrated metagenomics pipeline for strain profiling reveals novel patterns of bacterial transmission and biogeography. Genome Res. 2016;26:1612–1625. doi: 10.1101/gr.201863.115. PubMed DOI PMC

Mishra S., Imlay J.A. An anaerobic bacterium, Bacteroides thetaiotaomicron, uses a consortium of enzymes to scavenge hydrogen peroxide. Mol. Microbiol. 2013;90:1356–1371. doi: 10.1111/mmi.12438. PubMed DOI PMC

Kint N., Alves Feliciano C., Martins M.C., Morvan C., Fernandes S.F., Folgosa F., Dupuy B., Texeira M., Martin-Verstraete I. How the anaerobic enteropathogen Clostridioides difficile tolerates low O(2) tensions. mBio. 2020;11:e01559-20. doi: 10.1128/mBio.01559-20. PubMed DOI PMC

Ponpium P., Ratanakhanokchai K., Kyu K.L. Isolation and properties of a cellulosome-type multienzyme complex of the thermophilic Bacteroides sp. strain P-1. Enzym. Microb. Technol. 2000;26:459–465. doi: 10.1016/S0141-0229(99)00195-7. PubMed DOI

Chatzidaki-Livanis M., Geva-Zatorsky N., Comstock L.E. Bacteroides fragilis type VI secretion systems use novel effector and immunity proteins to antagonize human gut Bacteroidales species. Proc. Natl. Acad. Sci. USA. 2016;113:3627–3632. doi: 10.1073/pnas.1522510113. PubMed DOI PMC

Verster A.J., Ross B.D., Radey M.C., Bao Y., Goodman A.L., Mougous J.D., Borenstein E. The landscape of yype VI secretion across human gut microbiomes reveals its role in community composition. Cell Host Microbe. 2017;22:411–419.e4. doi: 10.1016/j.chom.2017.08.010. PubMed DOI PMC

Laanbroek H.J., Stal L.H., Veldkamp H. Utilization of hydrogen and formate by Campylobacter spec. under aerobic and anaerobic conditions. Arch. Microbiol. 1978;119:99–102. doi: 10.1007/BF00407935. PubMed DOI

Velayudhan J., Kelly D.J. Analysis of gluconeogenic and anaplerotic enzymes in Campylobacter jejuni: An essential role for phosphoenolpyruvate carboxykinase. Microbiology. 2002;148:685–694. doi: 10.1099/00221287-148-3-685. PubMed DOI

Guccione E., Leon-Kempis Mdel R., Pearson B.M., Hitchin E., Mulholland F., van Diemen P.M., Stevens M.P., Kelly D.J. Amino acid-dependent growth of Campylobacter jejuni: Key roles for aspartase (AspA) under microaerobic and oxygen-limited conditions and identification of AspB (Cj0762), essential for growth on glutamate. Mol. Microbiol. 2008;69:77–93. doi: 10.1111/j.1365-2958.2008.06263.x. PubMed DOI

Stoakes E., Savva G.M., Coates R., Tejera N., Poolman M.G., Grant A.J., Wain J., Singh D. Substrate utilisation and energy metabolism in non-growing Campylobacter jejuni M1cam. Microorganisms. 2022;10:1355. doi: 10.3390/microorganisms10071355. PubMed DOI PMC

Gonzalez-Rodriguez I., Gaspar P., Sanchez B., Gueimonde M., Margolles A., Neves A.R. Catabolism of glucose and lactose in Bifidobacterium animalis subsp. lactis, studied by 13C nuclear magnetic resonance. Appl. Environ. Microbiol. 2013;79:7628–7638. PubMed PMC

Fushinobu S. Unique sugar metabolic pathways of bifidobacteria. Biosci. Biotechnol. Biochem. 2010;74:2374–2384. doi: 10.1271/bbb.100494. PubMed DOI

Strobel H.J. Vitamin B12-dependent propionate production by the ruminal bacterium Prevotella ruminicola 23. Appl. Environ. Microbiol. 1992;58:2331–2333. doi: 10.1128/aem.58.7.2331-2333.1992. PubMed DOI PMC

Xu Z., Liu Y., Ye B.C. PccD regulates branched-chain amino acid degradation and exerts a negative effect on erythromycin production in Saccharopolyspora erythraea. Appl. Environ. Microb. 2018;84:e00049-18. doi: 10.1128/AEM.00049-18. PubMed DOI PMC

Shetty S.A., Marathe N.P., Lanjekar V., Ranade D., Shouche Y.S. Comparative genome analysis of Megasphaera sp. reveals niche specialization and its potential role in the human gut. PLoS ONE. 2013;8:e79353. doi: 10.1371/journal.pone.0079353. PubMed DOI PMC

Maki J.J., Looft T. Megasphaera stantonii sp. nov., a butyrate-producing bacterium isolated from the cecum of a healthy chicken. Int. J. Syst. Evol. Microbiol. 2018;68:3409–3415. doi: 10.1099/ijsem.0.002991. PubMed DOI

Kelly D.J. The physiology and metabolism of the human gastric pathogen Helicobacter pylori. Adv. Microb. Physiol. 1998;40:137–189. PubMed

Balle B.S., Poole R.K. Requirement for ubiquinone downstream of cytochrome(s) b in the oxygen-terminated respiratory chains of Escherichia coli K-12 revealed using a null mutant allele of ubiCA. Microbiology. 1998;144:361–373. doi: 10.1099/00221287-144-2-361. PubMed DOI

Li H., Durbin R. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics. 2009;25:1754–1760. doi: 10.1093/bioinformatics/btp324. PubMed DOI PMC

Li H., Handsaker B., Wysoker A., Fennell T., Ruan J., Homer N., Marth G., Abecasis G., Durbin R., 1000 Genome Project Data Processing Subgroup The sequence alignment/map format and SAMtools. Bioinformatics. 2009;25:2078–2079. doi: 10.1093/bioinformatics/btp352. PubMed DOI PMC

Li D., Luo R., Liu C.M., Leung C.M., Ting H.F., Sadakane K., Yamashita H., Lam T.W. MEGAHIT v1.0: A fast and scalable metagenome assembler driven by advanced methodologies and community practices. Methods. 2016;102:3–11. doi: 10.1016/j.ymeth.2016.02.020. PubMed DOI

Alneberg J., Bjarnason B.S., de Bruijn I., Schirmer M., Quick J., Ijaz U.Z., Lahti L., Loman N.J., Andersson A.F., Quince C. Binning metagenomic contigs by coverage and composition. Nat. Methods. 2014;11:1144–1146. doi: 10.1038/nmeth.3103. PubMed DOI

Wu Y.W., Simmons B.A., Singer S.W. MaxBin 2.0: An automated binning algorithm to recover genomes from multiple metagenomic datasets. Bioinformatics. 2016;32:605–607. doi: 10.1093/bioinformatics/btv638. PubMed DOI

Kang D.D., Li F., Kirton E., Thomas A., Egan R., An H., Wang Z. MetaBAT 2: An adaptive binning algorithm for robust and efficient genome reconstruction from metagenome assemblies. PeerJ. 2019;7:e7359. doi: 10.7717/peerj.7359. PubMed DOI PMC

Seppey M., Manni M., Zdobnov E.M. BUSCO: Assessing genome assembly and annotation completeness. Methods Mol. Biol. 2019;1962:227–245. PubMed

Uritskiy G.V., DiRuggiero J., Taylor J. MetaWRAP-a flexible pipeline for genome-resolved metagenomic data analysis. Microbiome. 2018;6:158. doi: 10.1186/s40168-018-0541-1. PubMed DOI PMC

Seemann T. Prokka: Rapid prokaryotic genome annotation. Bioinformatics. 2014;30:2068–2069. doi: 10.1093/bioinformatics/btu153. PubMed DOI

Hyatt D., Chen G.L., Locascio P.F., Land M.L., Larimer F.W., Hauser L.J. Prodigal: Prokaryotic gene recognition and translation initiation site identification. BMC Bioinform. 2010;11:119. doi: 10.1186/1471-2105-11-119. PubMed DOI PMC

Probiotic Mixtures Consisting of Representatives of Bacteroidetes and Selenomonadales Increase Resistance of Newly Hatched Chicks to Salmonella Enteritidis Infection