Spatial And Temporal Heterogeneity Of Microbiome Of Silurus Meridionalis

Trillions of microorganisms in nature consist of the important part of biosphere.Of them,numerous microboes colonize in/on the body surfaces of animals and exert a strong infunce on driving animals' environmental adaptations and their evolutionary processes.The symbiotic behavior is a result of long-term interactions,which has been a cutting-edge research hotspot in biology and ecology.In last decade,the development and combination of high-thruoughput sequencing and bioimformatics that overcome molecular technological limitations make it possible to study microbial community assembagles of animals and their functions to the hosts,and to further explore mechanisms underlying by which animals and microbiota communicate and interact.The major efforts of human gut have revealed that the microboes are affected by a series of factors spanning from host to environment such as host species and diet.This provides insights into host-microbiota relationship.It is essential to conduct fundmental work to explore microbial establishements and successions of animals.In China,the most rapid growth sector in agriculture is aquaculture,where fish farming is the most important part.China has the developed techniques for fish farming,yet much knowledge is unknown about gut microbiota community and the microbial ecology of commonly farmed fish species.The majority of studies focused on gut microbiota based on traditional methods.Meanwhile,few studies have investigated fish gut microbiota closely associated with factors in aquaculture such as individual development,food processing and antibiotic use,and so on.Beside gut,little is understoond about microbiota in other body regions such as gill and skin which both contain a mocusal layer like gut.In this study,we used a commercially important carnivorous fish,southern catfish(Silurus meridionalis),to study the differences of fish gut microbiome at multiple temporal scales and the spatial heterogeneity of fish microbiome at mulpile organ scales,combining with grass crap(Ctenopharyngodon idellus)and C57BL/6 mice for better elaborating microbial variations and dynamics and their dirvers and influcing factors.To explore the role of individual development in shaping gut microbiata by excluding dietary effects,we conducted a long-term analysis of gut microbiota associated with host age at 8,18,35,65 and 125 day post fertilization(dpf)and investigated microbial relationships among host,food and water environment at 8,35 and 125 dpf.The results showed that gut microbial diversity in southern catfish tended to increase linearly as host aged.Despite similar microbial communities in food and rearing water during the host development,gut microbiota significantly underwent dynamic changes.Compared to that at the early stages,gut microibota reached another stable status at 65 and 125 dpf.Gut microbiota had lower microbial diversity and dramatically differed from environmental microbiota.The gut was dominated by Tenericutes and Fusobacteria which both markedly varied with host age,whereas Spirochaetes and Bacteroidetes were persistently enriched in food and rearing water,respectively.These results provide direct evidence that host itself can shape gut microbial assemblages across development,which is independent on dietary effects.The knowledge enhances the understanding of gut microbial establishment in the developing fish and provides a useful resource for such studies of fish-or egg-associated microbiota in aquaculture.In order to further understanding the establishement of topographical mapping of fish,we conbined carnivorous southern catfish(stomach,gut,skin and gill)with herbivorous grass carp(foregut,midgut,hindgut,skin and gill)to study spatial distribution of fish microibome by body region,and compared their micorbome with rearing water microbiome.Although southern catfish and grass carp were subjected to the same envivronmental conditions,their micoribome markedly differed.For the same tissue,the two cohabiting fish had marked differences in microbial community.For each fish species,the separations of microbial community by body region were observed.The same type of interal body regions(digestive tract)or external body regions(skin and gill)showed more similar microbial community structure than that in different type of body reigions.Based on RMT(random matrix theory)model,we further constructed molecular ecology networks of the microbome of southern catfish and grass carp.The results revealed the significant differences in network topology and preporties between southern catfish and grass carp,demonstrating different microbial interactions in the two fish species and diversified interplays among microibota.Compared to the rearing water microbome,however,the internal body regions for southern catfish and grass carp harbored different microbiome,even though the fish directly contacted with the rearing water.These results demostrate that the microbiome of not only digestive tract but also skin and gill is shaped by host species and the microbial differences among body regions may be associated with their divergent niche habitats.Subsequently,we measured p H of digestive tract(stomach and gut)of southern catfish and studied the dynamics of gut microbiota by collecting microbial samples at 3,12 and 24 h after feeding.Stomach p H was acidic and gut p H was weak alkaline,and both of them were significantly reduced during food digestion.Compared to the gut,the stomach differently harbored microbial community.Firmicutes was the most abundant taxon in the stomach,whereas Fusobacteria dominated prevailed in the intestine.Firmicutes markedly increased and Fusobacteria decreased in the gastrointestinal tract after feeding.At 24 h,Proteobacteria further decreased with a significant increase of Bacteroidetes in the gut.The microbial community of gastrointestinal tract changed over the food digestion,but the microbial diversity did not,and a higher microbial diversity was consistently observed in the stomach than the gut.Based on KEGG ortholog function predictions by Phylogenetic Investigationof Communities by Reconstruction of Unobserved States(PICRUSt),we found that the stomach and intestine had significantly different functional categories,such as energy metabolism,glycan biosynthesis and metabolism,carbohydrate metabolism,and metabolism of cofactors and vitamins.The temporal differences in functional pathways were also found in the gastrointestinal tract.These results indicate that temporal dynamics of microbial community in the digestive tract of fish,suggesting the importance of sampling time for gut micoribal analysis.Furhter,we targeted southern catfish and C57BL/6 as experimental animals to explore the effect of thermal processing of food on gut microbiota,and to further compare the differences in microbial response between the two hosts by providing two foods(grass carp fillets and stone moroko(Pseudorasbora parva))with thermal and non-thermal processing.In this study,thermal processing did not significantly affect nutritional composition of the foods.However,southern catfish fed thermally processed grass carp fillets had significantly different profliles of amino acids but not fatty acids,compared to the corresponding counterparts.There were no differences in overall microbial composition and structure in the pairwise thermally and non-thermally processed food(TF and NF),but the differential communities between food and gut.Both fish and mice fed TF had significantly lower gut microbial diversity than those fed NF.Moreover,TF triggered the changes in their microbial communities.Comparative host studies further indicated host species determined gut microbial assemblies,even if fed with the same food.Fusobacteria was the most abundant phylum in the fish,and Bacteroidetes and Firmicutes dominated in the mice.The response of several dominated gut microbiota in the fish and mice to TF was taxonomically opposite at the phylum level,and those further found at the genus level.Our results reveal that thermal processing of food strongly contributes to the reduction of gut microbial diversity and differentially drives microbial alterations in a host-dependent manner.This study is the first time to reveal the effect of thermal processing of food on gut microbiota and opens a window of opportunity to understand the gut microbial community variations,and provides new insights into the microbial assemblages associated with the use of processing techniques in food preparation in humans and domesticated animals.Lastly,the effects of antibiotic on the gut microbiome of southern catfish and the growth performance were investigated by a therapeutic florfenicol treatment conducted with a dose of 20 mg/kg body weight per day for a week followed by a one-week recovery period.The results showed that gut microbiome was significantly affected by the florfenicol treatment.Meanwhile,the reduction of body weight and feed intake,and the increase of feed conversion ratio(FCR)were observed.However,gut microbiome and FCR gradually recovered to the control group level after the treatment was stopped for one week.Using PICRUSt,it was discovered that functional genes of gut microbiome changed with significant increases of altered pathways during a week of the florfenicol treatment and marked decreases of those for the antibiotic withdraw in the following week.Nevertheless,no impact on the death ratio and gut histological structure was found during the whole experimental period.This study indicates that florfenicol in aquaculture could affect the gut microbiome and growth of fish but be related to reversible alterations of them as a therapeutic strategy,potentially suggesting that the short-term use of florfenicol might have no long-term effects on the gut health.Moreover,it seems to be infeasible to treat florfenicol as a growth promoter to modeluate fish gut homeostasis for promoting fish growth in healthy aquaculture.In summary,our data indicate the temporal dynamics and stabilities of gut microbiota in southern catfish and the significant differences in the fish microbiome at the organ scales.The dynamics in microbiota could be contributed to its adaptation to environment(such as the dynamic food digestion and food processing),which is shaped by exogenous(antibiotic,food and the processing)and endogenous(such as ontogenesis,host species and niche differentiation of tissues)factors.