Ectotherms have peculiar relationships with microorganisms. For instance, bacteria are recovered from the blood and internal organs of healthy teleosts. However, the presence of microbial communities in the healthy teleost brain has not been proposed. Here, we report a living bacterial community in the brain of healthy salmonids with bacterial loads comparable to those of the spleen and 1000-fold lower than in the gut. Brain bacterial communities share >50% of their diversity with gut and blood bacterial communities. Using culturomics, we obtained 54 bacterial isolates from the brains of healthy trout. Comparative genomics suggests that brain bacteria may have adaptations for niche colonization and polyamine biosynthesis. In a natural system, Chinook salmon brain microbiomes shift from juveniles to reproductively mature adults. Our study redefines the physiological relationships between the brain and bacteria in teleosts. This symbiosis may endow salmonids with a direct mechanism to sense and respond to environmental microbes.

Center for Evolutionary and Theoretical Immunology Department of Biology University of New Mexico Albuquerque NM 87108 USA

Department of Fisheries Wildlife and Conservation Sciences Oregon State University Corvallis OR 97331 USA

Faculty of Fisheries and Protection of Waters South Bohemian Research Center of Aquaculture and Biodiversity of Hydrocenoses Institute of Aquaculture and Protection of Waters University of South Bohemia České Budějovice Czech Republic

Fish Health Division University of Veterinary Medicine Vienna Austria

Institute of Parasitology Biology Centre of the Czech Academy of Sciences České Budějovice Czech Republic

Norwegian Veterinary Institute Thormøhlens Gate 53C 5006 Bergen Norway

Zobrazit více v PubMed

Cryan J. F., O’Riordan K. J., Cowan C. S., Sandhu K. V., Bastiaanssen T. F., Boehme M., Codagnone M. G., Cussotto S., Fulling C., Golubeva A. V., The microbiota-gut-brain axis. Physiol. Rev. 99, 1877–2013 (2019). PubMed

O’Riordan K. J., Collins M. K., Moloney G. M., Knox E. G., Aburto M. R., Fülling C., Morley S. J., Clarke G., Schellekens H., Cryan J. F., Short chain fatty acids: Microbial metabolites for gut-brain axis signalling. Mol. Cell. Endocrinol. 546, 111572 (2022). PubMed

Mohanta L., Das B. C., Patri M., Microbial communities modulating brain functioning and behaviors in zebrafish: A mechanistic approach. Microb. Pathog. 145, 104251 (2020). PubMed

R. D. Berg, Translocation of indigenous bacteria from the intestinal tract, in Human Intestinal Microflora in Health and Disease (Academic Press, 1983), chap. 15, pp. 333–352.

Geva-Zatorsky N., Sefik E., Kua L., Pasman L., Tan T. G., Ortiz-Lopez A., Yanortsang T. B., Yang L., Jupp R., Mathis D., Mining the human gut microbiota for immunomodulatory organisms. Cell 168, 928–943.e11 (2017). PubMed PMC

McPherson A. C., Pandey S. P., Bender M. J., Meisel M., Systemic immunoregulatory consequences of gut commensal translocation. Trends Immunol. 42, 137–150 (2021). PubMed PMC

Z. Hu, C.-A. McKenzie, C. Smith, J. G. Haas, R. Lathe, The remarkable complexity of the brain microbiome in health and disease. bioRxiv 2023.02.06.527297 (2023). 10.1101/2023.02.06.527297. DOI

Branton W. G., Ellestad K. K., Maingat F., Wheatley B. M., Rud E., Warren R. L., Holt R. A., Surette M. G., Power C., Brain microbial populations in HIV/AIDS: α-Proteobacteria predominate independent of host immune status. PLOS ONE 8, e54673–e54673 (2013). PubMed PMC

Prudencio M., Belzil V. V., Batra R., Ross C. A., Gendron T. F., Pregent L. J., Murray M. E., Overstreet K. K., Piazza-Johnston A. E., Desaro P., Distinct brain transcriptome profiles in C9orf72-associated and sporadic ALS. Nat. Neurosci. 18, 1175–1182 (2015). PubMed PMC

Alonso R., Pisa D., Fernández-Fernández A. M., Carrasco L., Infection of fungi and bacteria in brain tissue from elderly persons and patients with Alzheimer’s disease. Front. Aging Neurosci. 10, 159 (2018). PubMed PMC

Bullock G. L., Snieszko S. F., Bacteria in blood and kidney of apparently healthy hatchery trout. Trans. Am. Fish. Soc. 98, 268–271 (1969).

Ofek T., Lalzar M., Izhaki I., Halpern M., Intestine and spleen microbiota composition in healthy and diseased tilapia. Anim. Microbiome 4, 50 (2022). PubMed PMC

Fronton F., Ferchiou S., Caza F., Villemur R., Robert D., St-Pierre Y., Insights into the circulating microbiome of Atlantic and Greenland halibut populations: The role of species-specific and environmental factors. Sci. Rep. 13, 5971 (2023). PubMed PMC

Berczi I., Bertok L., Bereznai T., Comparative studies on the toxicity of Escherichia coli lipopolysaccharide endotoxin in various animal species. Can. J. Microbiol. 12, 1070–1071 (1966). PubMed

Wedemeyer G., Ross A. J., Smith L., Some metabolic effects of bacterial endotoxins in salmonid fishes. J. Fish. Board Can. 26, 115–122 (1969).

Link C. D., Is there a brain microbiome? Neurosci. Insights 16, doi.org/10.1177/26331055211018709 (2021). PubMed PMC

Eisenhofer R., Minich J. J., Marotz C., Cooper A., Knight R., Weyrich L. S., Contamination in low microbial biomass microbiome studies: Issues and recommendations. Trends Microbiol. 27, 105–117 (2019). PubMed

Mangul S., Yang H. T., Strauli N., Gruhl F., Porath H. T., Hsieh K., Chen L., Daley T., Christenson S., Wesolowska-Andersen A., ROP: Dumpster diving in RNA-sequencing to find the source of 1 trillion reads across diverse adult human tissues. Genome Biol. 19, 36 (2018). PubMed PMC

Lusk R. W., Diverse and widespread contamination evident in the unmapped depths of high throughput sequencing data. PLOS ONE 9, e110808 (2014). PubMed PMC

Elkjaer M. L., Simon L., Frisch T., Bente L.-M., Kacprowski T., Thomassen M., Reynolds R., Baumbach J., Röttger R., Illes Z., Hypothesis of a potential BrainBiota and its relation to CNS autoimmune inflammation. Front. Immunol. 13, 1043579 (2022). PubMed PMC

Rasmussen J. A., Villumsen K. R., Duchêne D. A., Puetz L. C., Delmont T. O., Sveier H., von Gersdorff Jørgensen L., Præbel K., Martin M. D., Bojesen A. M., Gilbert M. T. P., Kristiansen K., Limborg M. T., Genome-resolved metagenomics suggests a mutualistic relationship between Mycoplasma and salmonid hosts. Commun. Biol. 4, 579 (2021). PubMed PMC

Wang J., Li Y., Jaramillo-Torres A., Einen O., Jakobsen J. V., Krogdahl Å., Kortner T. M., Exploring gut microbiota in adult Atlantic salmon (Salmo salar L.): Associations with gut health and dietary prebiotics. Anim. Microbiome 5, 47 (2023). PubMed PMC

Rasmussen J. A., Kiilerich P., Madhun A. S., Waagbø R., Lock E.-J. R., Madsen L., Gilbert M. T. P., Kristiansen K., Limborg M. T., Co-diversification of an intestinal Mycoplasma and its salmonid host. ISME J. 17, 682–692 (2023). PubMed PMC

Bozzi D., Rasmussen J. A., Carøe C., Sveier H., Nordøy K., Gilbert M. T. P., Limborg M. T., Salmon gut microbiota correlates with disease infection status: Potential for monitoring health in farmed animals. Anim. Microbiome 3, 30 (2021). PubMed PMC

Jin Y., Angell I. L., Sandve S. R., Snipen L. G., Olsen Y., Rudi K., Atlantic salmon raised with diets low in long-chain polyunsaturated n-3 fatty acids in freshwater have a Mycoplasma-dominated gut microbiota at sea. Aquac. Environ. Interact. 11, 31–39 (2019).

Lowrey L., Woodhams D. C., Tacchi L., Salinas I., Topographical mapping of the rainbow trout (Oncorhynchus mykiss) microbiome reveals a diverse bacterial community with antifungal properties in the skin. Appl. Environ. Microbiol. 81, 6915–6925 (2015). PubMed PMC

Y. Ding, A. Fernandez-Montero, A. Mani, E. Casadei, Y. Shibasaki, F. Takizawa, R. Miyazawa, I. Salinas, J. O. Sunyer, Secretory IgM (sIgM) is an ancient master regulator of microbiota homeostasis and metabolism. bioRxiv 2023.02.26.530119 (2023). 10.1101/2023.02.26.530119. DOI

Brown R. M., Wiens G. D., Salinas I., Analysis of the gut and gill microbiome of resistant and susceptible lines of rainbow trout (Oncorhynchus mykiss). Fish Shellfish Immunol. 86, 497–506 (2019). PubMed PMC

Lyons P. P., Turnbull J. F., Dawson K. A., Crumlish M., Phylogenetic and functional characterization of the distal intestinal microbiome of rainbow trout Oncorhynchus mykiss from both farm and aquarium settings. J. Appl. Microbiol. 122, 347–363 (2017). PubMed

C. Udoye, “Analytical challenges and characterization of skin and gut microbiota of Atlantic salmon fry in a commercial smolt production facility,” thesis, Norwegian University of Science and Technology, 2018).

L. T. Lowrey, “The microbiome of rainbow trout (Oncorhynchus mykiss),” thesis, University of New Mexico (2014).

Dehler C. E., Secombes C. J., Martin S. A. M., Environmental and physiological factors shape the gut microbiota of Atlantic salmon parr (Salmo salar L.). Aquaculture 467, 149–157 (2017). PubMed PMC

Ferchiou S., Caza F., Villemur R., Labonne J., St-Pierre Y., Skin and blood microbial signatures of sedentary and migratory trout (Salmo trutta) of the Kerguelen Islands. Fishes 8, 174 (2023).

Couch C. E., Neal W. T., Herron C. L., Kent M. L., Schreck C. B., Peterson J. T., Gut microbiome composition associates with corticosteroid treatment, morbidity, and senescence in Chinook salmon (Oncorhynchus tshawytscha). Sci. Rep. 13, 2567 (2023). PubMed PMC

Penny H. A., Domingues R. G., Krauss M. Z., Melo-Gonzalez F., Lawson M. A. E., Dickson S., Parkinson J., Hurry M., Purse C., Jegham E., Rhythmicity of intestinal IgA responses confers oscillatory commensal microbiota mutualism. Sci. Immunol. 7, eabk2541 (2022). PubMed PMC

Ellison A. R., Wilcockson D., Cable J., Circadian dynamics of the teleost skin immune-microbiome interface. Microbiome 9, 222 (2021). PubMed PMC

Holloway A. C., Reddy P. K., Sheridan M. A., Leatherland J. F., Diurnal rhythms of plasma growth hormone, somatostatin, thyroid hormones, cortisol and glucose concentrations in rainbow trout, Oncorhynchus mykiss, during progressive food deprivation. Biol. Rhythm Res. 25, 415–432 (1994).

Polakof S., Ceinos R. M., Fernández-Durán B., Míguez J. M., Soengas J. L., Daily changes in parameters of energy metabolism in brain of rainbow trout: Dependence on feeding. Comp. Biochem. Physiol. A Mol. Integr. Physiol. 146, 265–273 (2007). PubMed

Reddy P. K., Leatherland J. F., Does the time of feeding affect the diurnal rhythms of plasma hormone and glucose concentration and hepatic glycogen content of rainbow trout? Fish Physiol. Biochem. 13, 133–140 (1994). PubMed

Yang Y., Nguyen M., Khetrapal V., Sonnert N. D., Martin A. L., Chen H., Kriegel M. A., Palm N. W., Within-host evolution of a gut pathobiont facilitates liver translocation. Nature 607, 563–570 (2022). PubMed PMC

Bledsoe J. W., Peterson B. C., Swanson K. S., Small B. C., Ontogenetic characterization of the intestinal microbiota of channel catfish through 16S rRNA gene sequencing reveals insights on temporal shifts and the influence of environmental microbes. PLOS ONE 11, e0166379 (2016). PubMed PMC

Sugita H., Nakamura T., Deguchi Y., Identification of Plesiomonas shigelloides isolated from freshwater fish with the microplate hybridization method. J. Food Prot. 56, 949–953 (1993). PubMed

Skrodenytė-Arbačiauskienė V., Sruoga A., Butkauskas D., Skrupskelis K., Phylogenetic analysis of intestinal bacteria of freshwater salmon Salmo salar and sea trout Salmo trutta trutta and diet. Fish. Sci. 74, 1307–1314 (2008).

Larsen A. M., Mohammed H. H., Arias C. R., Characterization of the gut microbiota of three commercially valuable warmwater fish species. J. Appl. Microbiol. 116, 1396–1404 (2014). PubMed

Al-Hisnawi A., Ringø E., Davies S. J., Waines P., Bradley G., Merrifield D. L., First report on the autochthonous gut microbiota of brown trout (Salmo trutta Linnaeus). Aquacult. Res. 46, 2962–2971 (2015).

Wu S., Gao T., Zheng Y., Wang W., Cheng Y., Wang G., Microbial diversity of intestinal contents and mucus in yellow catfish (Pelteobagrus fulvidraco). Aquaculture 303, 1–7 (2010).

McDonald R., Schreier H. J., Watts J. E. M., Phylogenetic analysis of microbial communities in different regions of the gastrointestinal tract in Panaque nigrolineatus, a wood-eating fish. PLOS ONE 7, e48018 (2012). PubMed PMC

Duman M., García Valdés E., Ay H., Altun S., Saticioglu I. B., Description of a Novel Fish Pathogen, Plesiomonas shigelloides subsp. oncorhynchi, Isolated from Rainbow trout (Oncorhynchus mykiss): First genome analysis and comparative genomics. Fishes 8, 179 (2023).

Chen H., Zhao Y., Chen K., Wei Y., Luo H., Li Y., Liu F., Zhu Z., Hu W., Luo D., Isolation, identification, and investigation of pathogenic bacteria from common carp (Cyprinus carpio) naturally infected with Plesiomonas shigelloides. Front. Immunol. 13, 872896 (2022). PubMed PMC

Nisha R. G., Rajathi V., Manikandan R., Prabhu N. M., Isolation of Plesiomonas shigelloides from infected cichlid fishes using 16S rRNA characterization and its control with probiotic Pseudomonas sp. Acta Sci. Vet. 42, 1–7 (2014).

Yin Z., Zhang S., Wei Y., Wang M., Ma S., Yang S., Wang J., Yuan C., Jiang L., Du Y., Horizontal gene transfer clarifies taxonomic confusion and promotes the genetic diversity and pathogenicity of Plesiomonas shigelloides. mSystems 5, e00448-20 (2020). PubMed PMC

Nakamura A., Kurihara S., Takahashi D., Ohashi W., Nakamura Y., Kimura S., Onuki M., Kume A., Sasazawa Y., Furusawa Y., Symbiotic polyamine metabolism regulates epithelial proliferation and macrophage differentiation in the colon. Nat. Commun. 12, 2105 (2021). PubMed PMC

Michael A. J., Polyamines in eukaryotes, bacteria, and archaea. J. Biol. Chem. 291, 14896–14903 (2016). PubMed PMC

Krysenko S., Wohlleben W., Polyamine and ethanolamine metabolism in bacteria as an important component of nitrogen assimilation for survival and pathogenicity. Med. Sci. 10, 40 (2022). PubMed PMC

Hamana K., Akiba T., Uchino F., Matsuzaki S., Distribution of spermine in bacilli and lactic acid bacteria. Can. J. Microbiol. 35, 450–455 (1989). PubMed

Hamana K., Tanaka T., Hosoya R., Niitsu M., Itoh T., Cellular polyamines of the acidophilic, thermophilic and thermoacidophilic archaebacteria, Acidilobus, Ferroplasma, Pyrobaculum, Pyrococcus, Staphylothermus, Thermococcus, Thermodiscus and Vulcanisaeta. J. Gen. Appl. Microbiol. 49, 287–293 (2003). PubMed

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 Bioinformatics 11, 119 (2010). PubMed PMC

Schroeder S., Hofer S. J., Zimmermann A., Pechlaner R., Dammbrueck C., Pendl T., Marcello G. M., Pogatschnigg V., Bergmann M., Muller M., Dietary spermidine improves cognitive function. Cell Rep. 35, 108985 (2021). PubMed

Schwarz C., Benson G. S., Horn N., Wurdack K., Grittner U., Schilling R., Märschenz S., Köbe T., Hofer S. J., Magnes C., Effects of spermidine supplementation on cognition and biomarkers in older adults with subjective cognitive decline: A randomized clinical trial. JAMA Netw. Open 5, e2213875 (2022). PubMed PMC

Yang Y., Chen S., Zhang Y., Lin X., Song Y., Xue Z., Qian H., Wang S., Wan G., Zheng X., Induction of autophagy by spermidine is neuroprotective via inhibition of caspase 3-mediated Beclin 1 cleavage. Cell Death Dis. 8, e2738 (2017). PubMed PMC

Sandusky-Beltran L. A., Kovalenko A., Ma C., Calahatian J. I. T., Placides D. S., Watler M. D., Hunt J. B., Darling A. L., Baker J. D., Blair L. J., Spermidine/spermine-N1-acetyltransferase ablation impacts tauopathy-induced polyamine stress response. Alzheimer’s Res. Ther. 11, 58 (2019). PubMed PMC

Glantz L., Nates J. L., Trembovler V., Bass R., Shohami E., Polyamines induce blood-brain barrier disruption and edema formation in the rat. J. Basic Clin. Physiol. Pharmacol. 7, 1–10 (1996). PubMed

Camak D. T., Osborne M. J., Turner T. F., Population genomics and conservation of Gila Trout (Oncorhynchus gilae). Conserv. Genet. 22, 729–743 (2021).

Peters M. B., Turner T. F., Genetic variation of the major histocompatibility complex (MHC class II β gene) in the threatened Gila trout, Oncorhynchus gilae gilae. Conserv. Genet. 9, 257–270 (2008).

Wares J. P., Alò D., Turner T. F., A genetic perspective on management and recovery of federally endangered trout (Oncorhynchus gilae) in the American Southwest. Can. J. Fish. Aquat. Sci. 61, 1890–1899 (2004).

B. E. Riddell, R. D. Brodeur, A. V. Bugaev, P. Moran, J. M. Murphy, J. A. Orsi, M. Trudel, L. A. Weitkamp, B. K. Wells, A. C. Wertheimer, Ocean ecology of Chinook salmon, in Ocean Ecology of Pacific Salmon and Trout (American Fisheries Society, 2018), pp. 555–696.

Schroeder R. K., Whitman L. D., Cannon B., Olmsted P., Juvenile life-history diversity and population stability of spring Chinook salmon in the Willamette River basin, Oregon. Can. J. Fish. Aquat. Sci. 73, 921–934 (2016).

Couch C. E., Colvin M. E., Chitwood R. L., Peterson J. T., Schreck C. B., Scope of the cortisol stress response in Chinook salmon during maturation. Fish. Res. 254, 106416 (2022).

Corbett S. C., Moser M. L., Dittman A. H., Experimental evaluation of adult spring Chinook salmon radio-tagged during the late stages of spawning migration. N. Am. J. Fish. Manag. 32, 853–858 (2012).

Dolan B. P., Fisher K. M., Colvin M. E., Benda S. E., Peterson J. T., Kent M. L., Schreck C. B., Innate and adaptive immune responses in migrating spring-run adult chinook salmon, Oncorhynchus tshawytscha. Fish Shellfish Immunol. 48, 136–144 (2016). PubMed

Maule A. G., Schrock R., Slater C., Fitzpatrick M. S., Schreck C. B., Immune and endocrine responses of adult chinook salmon during freshwater immigration and sexual maturation. Fish Shellfish Immunol. 6, 221–233 (1996).

T. A. Maldonado, Distribution of Neurodegeneration, Amyloid Precursor Protein, β-Amyloid Peptide and Neuroprotection in the Brain of Migrating and Senescent Kokanee Salmon (Univ. of Colorado Boulder, 2000).

Llewellyn M. S., McGinnity P., Dionne M., Letourneau J., Thonier F., Carvalho G. R., Creer S., Derome N., The biogeography of the atlantic salmon (Salmo salar) gut microbiome. ISME J. 10, 1280–1284 (2016). PubMed PMC

R. Mann, C. Caudill, M. Keefer, A. Roumasset, C. Schreck, M. Kent, “Migration behavior and spawning success of spring Chinook salmon in Fall Creek and the North Fork Middle Fork Willamette River: Relationships among fate, fish condition, and environmental factors 2010” (Technical report 2015-2).

Nervino S., Polley T., Peterson J. T., Schreck C. B., Kent M. L., Alexander J. D., Intestinal lesions and parasites associated with senescence and prespawn mortality in Chinook Salmon (Oncorhynchus tshawytscha). J. Fish Dis. 7, e13876 (2024). PubMed

Varatharaj A., Galea I., The blood-brain barrier in systemic inflammation. Brain Behav. Immun. 60, 1–12 (2017). PubMed

Elwood E., Lim Z., Naveed H., Galea I., The effect of systemic inflammation on human brain barrier function. Brain Behav. Immun. 62, 35–40 (2017). PubMed PMC

Sun Y., Koyama Y., Shimada S., Inflammation from peripheral organs to the brain: How does systemic inflammation cause neuroinflammation? Front. Aging Neurosci. 14, 903455 (2022). PubMed PMC

C. Lobsinger, “Energy expenditure during breeding competition between feral Chinook salmon (Oncorphynchus tshawytscha) and native Atlantic salmon (Salmo salar),” thesis, University of British Columbia (2004).

Maldonado T. A., Jones R. E., Norris D. O., Distribution of β-amyloid and amyloid precursor protein in the brain of spawning (senescent) salmon: A natural, brain-aging model. Brain Res. 858, 237–251 (2000). PubMed

Vojtechova I., Machacek T., Kristofikova Z., Stuchlik A., Petrasek T., Infectious origin of Alzheimer’s disease: Amyloid beta as a component of brain antimicrobial immunity. PLOS Pathog. 18, e1010929 (2022). PubMed PMC

Gosztyla M. L., Brothers H. M., Robinson S. R., Alzheimer’s amyloid-β is an antimicrobial peptide: A review of the evidence. J. Alzheimers Dis. 62, 1495–1506 (2018). PubMed

Kumar D. K. V., Choi S. H., Washicosky K. J., Eimer W. A., Tucker S., Ghofrani J., Lefkowitz A., McColl G., Goldstein L. E., Tanzi R. E., Moir R. D., Amyloid-β peptide protects against microbial infection in mouse and worm models of Alzheimer’s disease. Sci. Transl. Med. 8, 340ra72 (2016). PubMed PMC

Maldonado T. A., Jones R. E., Norris D. O., Timing of neurodegeneration and beta-amyloid (Aβ) peptide deposition in the brain of aging kokanee salmon. J. Neurobiol. 53, 21–35 (2002). PubMed

Maldonado T. A., Jones R. E., Norris D. O., Intraneuronal amyloid precursor protein (APP) and appearance of extracellular β-amyloid peptide (Aβ) in the brain of aging kokanee salmon. J. Neurobiol. 53, 11–20 (2002). PubMed

Fung T. C., Bessman N. J., Hepworth M. R., Kumar N., Shibata N., Kobuley D., Wang K., Ziegler C. G. K., Goc J., Shima T., Umesaki Y., Sartor R. B., Sullivan K. V., Lawley T. D., Kunisawa J., Kiyono H., Sonnenberg G. F., Lymphoid-tissue-resident commensal bacteria promote members of the IL-10 cytokine family to establish mutualism. Immunity 44, 634–646 (2016). PubMed PMC

Obata T., Goto Y., Shibata N., Sato S., Kunisawa J., Kiyono H., Indigenous opportunistic bacteria inhabit mammalian gut-associated lymphoid tissues for mucosal antibody-mediated symbiosis. Biol. Sci. 107, 7419–7424 (2010). PubMed PMC

Novoa B., Bowman T. V., Zon L., Figueras A., LPS response and tolerance in the zebrafish (Danio rerio). Fish Shellfish Immunol. 26, 326–331 (2009). PubMed PMC

E. E. Adade, R. J. Stevick, D. Pérez-Pascual, J.-M. Ghigo, A. M. Valm, Gnotobiotic zebrafish microbiota display inter-individual variability affecting host physiology. bioRxiv 2023.02.01.526612 (2023). 10.1101/2023.02.01.526612. DOI

Zhang M., Shan C., Tan F., Limbu S. M., Chen L., Du Z.-Y., Gnotobiotic models: Powerful tools for deeply understanding intestinal microbiota-host interactions in aquaculture. Aquaculture 517, 734800 (2020).

Melancon E., Canny S. G. D. L. T., Sichel S., Kelly M., Wiles T., Rawls J., Eisen J., Guillemin K., Best practices for germ-free derivation and gnotobiotic zebrafish husbandry. Methods Cell Biol. 138, 61–100 (2017). PubMed PMC

Pérez-Pascual D., Vendrell-Fernández S., Audrain B., Bernal-Bayard J., Patiño-Navarrete R., Petit V., Rigaudeau D., Ghigo J.-M., Gnotobiotic rainbow trout (Oncorhynchus mykiss) model reveals endogenous bacteria that protect against Flavobacterium columnare infection. PLOS Pathog. 17, e1009302 (2021). PubMed PMC

de la Torre Canny S. G., Nordgård C. T., Mathisen A. J. H., Lorentsen E. D., Vadstein O., Bakke I., A novel gnotobiotic experimental system for Atlantic salmon (Salmo salar L.) reveals a microbial influence on mucosal barrier function and adipose tissue accumulation during the yolk sac stage. Front. Cell. Infect. Microbiol. 12, 1068302 (2023). PubMed PMC

Kim D., Hofstaedter C. E., Zhao C., Mattei L., Tanes C., Clarke E., Lauder A., Sherrill-Mix S., Chehoud C., Kelsen J., Optimizing methods and dodging pitfalls in microbiome research. Microbiome 5, 52 (2017). PubMed PMC

Noh K., Liu X., Wei C., Optimizing transcardial perfusion of small molecules and biologics for brain penetration and biodistribution studies in rodents. Biopharm. Drug Dispos. 44, 71–83 (2023). PubMed

Perdiguero P., Martín-Martín A., Benedicenti O., Díaz-Rosales P., Morel E., Muñoz-Atienza E., García-Flores M., Simón R., Soleto I., Cerutti A., Teleost IgD+ IgM− B cells mount clonally expanded and mildly mutated intestinal IgD responses in the absence of lymphoid follicles. Cell Rep. 29, 4223–4235.e5 (2019). PubMed PMC

de Nies L., Busi S. B., Tsenkova M., Halder R., Letellier E., Wilmes P., Evolution of the murine gut resistome following broad-spectrum antibiotic treatment. Nat. Commun. 13, 2296 (2022). PubMed PMC

Phelps C. M., Shapira J. H., Laughlin C. R., Meisel M., Detection of viable commensal bacteria in murine melanoma tumors by culturomics. STAR Protocols 4, 102492 (2023). PubMed PMC

Mitchell K. R., Takacs-Vesbach C. D., A comparison of methods for total community DNA preservation and extraction from various thermal environments. J. Ind. Microbiol. Biotechnol. 35, 1139–1147 (2008). PubMed

Bolyen E., Rideout J. R., Dillon M. R., Bokulich N. A., Abnet C. C., Al-Ghalith G. A., Alexander H., Alm E. J., Arumugam M., Asnicar F., Bai Y., Bisanz J. E., Bittinger K., Brejnrod A., Brislawn C. J., Brown C. T., Callahan B. J., Caraballo-Rodríguez A. M., Chase J., Cope E. K., Silva R. D., Diener C., Dorrestein P. C., Douglas G. M., Durall D. M., Duvallet C., Edwardson C. F., Ernst M., Estaki M., Fouquier J., Gauglitz J. M., Gibbons S. M., Gibson D. L., Gonzalez A., Gorlick K., Guo J., Hillmann B., Holmes S., Holste H., Huttenhower C., Huttley G. A., Janssen S., Jarmusch A. K., Jiang L., Kaehler B. D., Kang K. B., Keefe C. R., Keim P., Kelley S. T., Knights D., Koester I., Kosciolek T., Kreps J., Langille M. G. I., Lee J., Ley R., Liu Y.-X., Loftfield E., Lozupone C., Maher M., Marotz C., Martin B. D., Donald D. M., McIver L. J., Melnik A. V., Metcalf J. L., Morgan S. C., Morton J. T., Naimey A. T., Navas-Molina J. A., Nothias L. F., Orchanian S. B., Pearson T., Peoples S. L., Petras D., Preuss M. L., Pruesse E., Rasmussen L. B., Rivers A., Robeson M. S. II, Rosenthal P., Segata N., Shaffer M., Shiffer A., Sinha R., Song S. J., Spear J. R., Swafford A. D., Thompson L. R., Torres P. J., Trinh P., Tripathi A., Turnbaugh P. J., Ul-Hasan S., van der Hooft J. J. J., Vargas F., Vázquez-Baeza Y., Vogtmann E., von Hippel M., Walters W., Wan Y., Wang M., Warren J., Weber K. C., Williams C. H. D., Willis A. D., Xu Z. Z., Zaneveld J. R., Zhang Y., Zhu Q., Knight R., Caporaso J. G., Reproducible, interactive, scalable and extensible microbiome data science using QIIME 2. Nat. Biotechnol. 37, 852–857 (2019). PubMed PMC

Callahan B. J., McMurdie P. J., Rosen M. J., Han A. W., Johnson A. J. A., Holmes S. P., DADA2: High-resolution sample inference from Illumina amplicon data. Nat. Methods 13, 581–583 (2016). PubMed PMC

Quast C., Pruesse E., Yilmaz P., Gerken J., Schweer T., Yarza P., Peplies J., Glöckner F. O., The SILVA ribosomal RNA gene database project: Improved data processing and web-based tools. Nucleic Acids Res. 41, D590–D596 (2012). PubMed PMC

J. E. Bisanz, qiime2R: Importing QIIME2 Artifacts and Associated Data Into R Sessions (2018); R Package version 0.99.

Knights D., Kuczynski J., Charlson E. S., Zaneveld J., Mozer M. C., Collman R. G., Bushman F. D., Knight R., Kelley S. T., Bayesian community-wide culture-independent microbial source tracking. Nat. Methods 8, 761–763 (2011). PubMed PMC

Liu Y., Elworth R. A. L., Jochum M. D., Aagaard K. M., Treangen T. J., De novo identification of microbial contaminants in low microbial biomass microbiomes with Squeegee. Nat. Commun. 13, 6799 (2022). PubMed PMC

Zemb O., Achard C. S., Hamelin J., De Almeida M., Gabinaud B., Cauquil L., Verschuren L. M. G., Godon J., Absolute quantitation of microbes using 16S rRNA gene metabarcoding: A rapid normalization of relative abundances by quantitative PCR targeting a 16S rRNA gene spike-in standard. Microbiology Open 9, e977 (2020). PubMed PMC

Leger A., Leonardi T., pycoQC, interactive quality control for Oxford nanopore sequencing. J. Open Source Softw. 4, 1236 (2019).

Shafin K., Pesout T., Lorig-Roach R., Haukness M., Olsen H. E., Bosworth C., Armstrong J., Tigyi K., Maurer N., Koren S., Sedlazeck F. J., Marschall T., Mayes S., Costa V., Zook J. M., Liu K. J., Kilburn D., Sorensen M., Munson K. M., Vollger M. R., Monlong J., Garrison E., Eichler E. E., Salama S., Haussler D., Green R. E., Akeson M., Phillippy A., Miga K. H., Carnevali P., Jain M., Paten B., Nanopore sequencing and the Shasta toolkit enable efficient de novo assembly of eleven human genomes. Nat. Biotechnol. 38, 1044–1053 (2020). PubMed PMC

Seemann T., Prokka: Rapid prokaryotic genome annotation. Bioinformatics 30, 2068–2069 (2014). PubMed

Deorowicz S., Debudaj-Grabysz A., Gudyś A., FAMSA: Fast and accurate multiple sequence alignment of huge protein families. Sci. Rep. 6, 33964 (2016). PubMed PMC

Eren A. M., Esen Ö. C., Quince C., Vineis J. H., Morrison H. G., Sogin M. L., Delmont T. O., Anvi’o: An advanced analysis and visualization platform for ‘omics data. PeerJ 3, e1319 (2015). PubMed PMC

Nguyen L.-T., Schmidt H. A., Von Haeseler A., Minh B. Q., IQ-TREE: A fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Mol. Biol. Evol. 32, 268–274 (2015). PubMed PMC

Jelsbak L., Thomsen L. E., Wallrodt I., Jensen P. R., Olsen J. E., Polyamines are required for virulence in Salmonella enterica serovar Typhimurium. PLOS ONE 7, e36149 (2012). PubMed PMC