Metaproteomics Analysis Reveals the Adaptation Process for the Chicken Gut Microbiota

The animal gastrointestinal tract houses a large microbial community, the gut microbiota, that confers many benefits to its host, such as protection from pathogens and provision of essential metabolites. Metagenomic approaches have defined the chicken fecal microbiota in other studies, but here, we wished to assess the correlation between the metagenome and the bacterial proteome in order to better understand the healthy chicken gut microbiota. Here, we performed high-throughput sequencing of 16S rRNA gene amplicons and metaproteomics analysis of fecal samples to determine microbial gut composition and protein expression. 16 rRNA gene sequencing analysis identified Clostridiales, Bacteroidaceae, and Lactobacillaceae species as the most abundant species in the gut. For metaproteomics analysis, peptides were generated by using the Fasp method and subsequently fractionated by strong anion exchanges. Metaproteomics analysis identified 3,673 proteins. Among the most frequently identified proteins, 380 proteins belonged to Lactobacillus spp., 155 belonged to Clostridium spp., and 66 belonged to Streptococcus spp. The most frequently identified proteins were heat shock chaperones, including 349 GroEL proteins, from many bacterial species, whereas the most abundant enzymes were pyruvate kinases, as judged by the number of peptides identified per protein (spectral counting). Gene ontology and KEGG pathway analyses revealed the functions and locations of the identified proteins. The findings of both metaproteomics and 16S rRNA sequencing analyses are discussed.     

Compositional Stability of a Salivary Bacterial Population against Supragingival Microbiota Shift following Periodontal Therapy

Supragingival plaque is permanently in contact with saliva. However, the extent to which the microbiota contributes to the salivary bacterial population remains unclear. We compared the compositional shift in the salivary bacterial population with that in supragingival plaque following periodontal therapy. Samples were collected from 19 patients with periodontitis before and after periodontal therapy (mean sample collection interval, 25.8±2.6 months), and their bacterial composition was investigated using barcoded pyrosequencing analysis of the 16S rRNA gene. Phylogenetic community analysis using the UniFrac distance metric revealed that the overall bacterial community composition of saliva is distinct from that of supragingival plaque, both pre- and post-therapy. Temporal variation following therapy in the salivary bacterial population was significantly smaller than in the plaque microbiota, and the post-therapy saliva sample was significantly more similar to that pre-therapy from the same individual than to those from other subjects. Following periodontal therapy, microbial richness and biodiversity were significantly decreased in the plaque microbiota, but not in the salivary bacterial population. The operational taxonomic units whose relative abundances changed significantly after therapy were not common to the two microbiotae. These results reveal the compositional stability of salivary bacterial populations against shifts in the supragingival microbiota, suggesting that the effect of the supragingival plaque microbiota on salivary bacterial population composition is limited.     

Iron in infectious diseases friend or foe?: The role of gut microbiota

Iron is a trace element involved in metabolic functions for all organisms, from microorganisms to mammalians. Iron deficiency is a prevalent health problem that affects billions of people worldwide, and iron overload could have some hazardous effect. The complex microbial community in the human body, also called microbiota, influences the host immune defence against infections. An imbalance in gut microbiota, dysbiosis, changes the host's susceptibility to infections by regulating the immune system. In recent years, the number of studies on the relationship between infectious diseases and microbiota has increased. Gut microbiota is affected by different parameters, including mode of delivery, hygiene habits, diet, drugs, and plasma iron levels during the lifetime. Gut microbiota may influence iron levels in the body, and iron overload and deficiency can also affect gut microbiota composition. Novel researches on microbiota shed light on the fact that the bidirectional interactions between gut microbiota and iron play a role in the pathogenesis of many diseases, especially infections. A better understanding of these interactions may help us to comprehend the pathogenesis of many infectious and metabolic diseases affecting people worldwide and following the development of more effective preventive and/or therapeutic strategies. In this review, we aimed to present the iron-mediated host-gut microbiota interactions, susceptibility to bacterial infections, and iron-targeted therapy approaches for infections.     

Differential Effects of Antibiotic Therapy on the Structure and Function of Human Gut Microbiota

The human intestinal microbiota performs many essential functions for the host. Antimicrobial agents, such as antibiotics (AB), are also known to disturb microbial community equilibrium, thereby having an impact on human physiology. While an increasing number of studies investigate the effects of AB usage on changes in human gut microbiota biodiversity, its functional effects are still poorly understood. We performed a follow-up study to explore the effect of ABs with different modes of action on human gut microbiota composition and function. Four individuals were treated with different antibiotics and samples were taken before, during and after the AB course for all of them. Changes in the total and in the active (growing) microbiota as well as the functional changes were addressed by 16S rRNA gene and metagenomic 454-based pyrosequencing approaches. We have found that the class of antibiotic, particularly its antimicrobial effect and mode of action, played an important role in modulating the gut microbiota composition and function. Furthermore, analysis of the resistome suggested that oscillatory dynamics are not only due to antibiotic-target resistance, but also to fluctuations in the surviving bacterial community. Our results indicated that the effect of AB on the human gut microbiota relates to the interaction of several factors, principally the properties of the antimicrobial agent, and the structure, functions and resistance genes of the microbial community.     

Composition, diversity, and origin of the bacterial community in grass carp intestine

Gut microbiota has become an integral component of the host, and received increasing attention. However, for many domestic animals, information on the microbiota is insufficient and more effort should be exerted to manage the gastrointestinal bacterial community. Understanding the factors that influence the composition of microbial community in the host alimentary canal is essential to manage or improve the microbial community composition. In the present study, 16S rRNA gene sequence-based comparisons of the bacterial communities in the grass carp (Ctenopharyngodon idellus) intestinal contents and fish culture-associated environments are performed. The results show that the fish intestinal microbiota harbors many cellulose-decomposing bacteria, including sequences related to Anoxybacillus, Leuconostoc, Clostridium, Actinomyces, and Citrobacter. The most abundant bacterial operational taxonomic units (OTUs) in the grass carp intestinal content are those related to feed digestion. In addition, the potential pathogens and probiotics are important members of the intestinal microbiota. Further analyses show that grass carp intestine holds a core microbiota composed of Proteobacteria, Firmicutes, and Actinobacteria. The comparison analyses reveal that the bacterial community in the intestinal contents is most similar to those from the culture water and sediment. However, feed also plays significant influence on the composition of gut microbiota.     

七味白术散对菌群失调腹泻小鼠肠道细菌多样性的影响

【背景】肠道微生物与人体多项生理功能密切相关,我们课题组前期研究表明七味白术散对菌群失调腹泻有较好的疗效。【目的】探讨与七味白术散疗效相关肠道微生物,比较传统中药饮片与超微中药饮片的疗效,进一步明确七味白术散治疗菌群失调腹泻的机理,为临床应用提供科学依据。【方法】应用抗生素建立菌群失调小鼠腹泻模型,分别灌胃给药七味白术散传统汤剂和超微50%量七味白术散汤剂。治疗结束后,提取肠道内容物微生物宏基因组DNA,建立16S rRNA基因文库进行测序。【结果】肠道微生物中的群落由乳酸菌(Lactobacillus spp.)、屎肠球菌(Enterococcus feacium)、梭状芽孢杆菌(Clostridium spp.)、黏液真杆菌(Blautia producta)、毛螺旋菌(Anaerostipes spp.)、腐生性葡萄球菌(Staphylococcus saprophyticus)和不能培养的细菌组成,其中乳酸菌为优势菌,占全部细菌DNA克隆数的61.90%。经抗生素造模后Lactobacillus spp.数量明显减少,Enterococcus feacium成为优势菌种,Clostridium spp.、Blautia product、Anaerostipes spp.和Staphylococcus saprophyticus等在数量上开始有增长的趋势,而治疗结束后,传统七味白术散治疗组和超微50%量七味白术散治疗组的Lactobacillus spp.比例均有所恢复,且超微50%量七味白术散治疗组的乳酸菌比例与正常组最接近;通过建立聚类树和计算各组中各菌群的多样性(H)、丰富度(S)、均匀度(E)及优势度指数(D)可知,超微50%量七味白术散治疗组可与正常组聚为一类,且超微50%量七味白术散治疗组各项指数与正常组最为接近,说明超微50%量七味白术散治疗组肠道微生物的基因文库及多样性与正常组最接近,超微50%量七味白术散汤剂的治疗效果优于七味白术散传统汤剂。【结论】应用16S rRNA基因克隆文库技术进一步明确了正常小鼠肠道中细菌群落的组成,以及七味白术散对菌群失调腹泻小鼠肠道细菌群落的恢复效果,超微50%量七味白术散汤剂的治疗效果优于七味白术散传统汤剂。     

Application of Methods for Identifying Broiler Chicken Gut Bacterial Species Linked with Increased Energy Metabolism

A high-throughput microbial profiling tool based on terminal restriction fragment length polymorphism was developed to monitor the poultry gut microbiota in response to dietary manipulations. Gut microbial communities from the duodena, jejuna, ilea, and ceca of 48 birds fed either a barley control diet or barley diet supplemented with exogenous enzymes for degrading nonstarch polysaccharide were characterized by using multivariate statistical methods. Analysis of samples showed that gut microbial communities varied significantly among gut sections, except between the duodenum and jejunum. Significant diet-associated differences in gut microbial communities were detected within the ileum and cecum only. The dissimilarity in bacterial community composition between diets was 73 and 66% within the ileum and cecum, respectively. Operational taxonomic units, representing bacterial species or taxonomically related groups, contributing to diet-associated differences were identified. Several bacterial species contributed to differences between diet-related gut microbial community composition, with no individual bacterial species contributing more than 1 to 5% of the total. Using canonical analysis of principal coordinates biplots, we correlated differences in gut microbial community composition within the ileum and cecum to improved performance, as measured by apparent metabolizable energy. This is the first report that directly links differences in the composition of the gut microbial community with improved performance, which implies that the presence of specific beneficial and/or absence of specific detrimental bacterial species may contribute to the improved performance in these birds.     

Deterministic Assembly of Complex Bacterial Communities in Guts of Germ-Free Cockroaches

The gut microbiota of termites plays important roles in the symbiotic digestion of lignocellulose. However, the factors shaping the microbial community structure remain poorly understood. Because termites cannot be raised under axenic conditions, we established the closely related cockroach Shelfordella lateralis as a germ-free model to study microbial community assembly and host-microbe interactions. In this study, we determined the composition of the bacterial assemblages in cockroaches inoculated with the gut microbiota of termites and mice using pyrosequencing analysis of their 16S rRNA genes. Although the composition of the xenobiotic communities was influenced by the lineages present in the foreign inocula, their structure resembled that of conventional cockroaches. Bacterial taxa abundant in conventional cockroaches but rare in the foreign inocula, such as Dysgonomonas and Parabacteroides spp., were selectively enriched in the xenobiotic communities. Donor-specific taxa, such as endomicrobia or spirochete lineages restricted to the gut microbiota of termites, however, either were unable to colonize germ-free cockroaches or formed only small populations. The exposure of xenobiotic cockroaches to conventional adults restored their normal microbiota, which indicated that autochthonous lineages outcompete foreign ones. Our results provide experimental proof that the assembly of a complex gut microbiota in insects is deterministic.     

Comparative assessment of human and farm animal faecal microbiota using real-time quantitative PCR

Pollution of the environment by human and animal faecal pollution affects the safety of shellfish, drinking water and recreational beaches. To pinpoint the origin of contaminations, it is essential to define the differences between human microbiota and that of farm animals. A strategy based on real-time quantitative PCR (qPCR) assays was therefore developed and applied to compare the composition of intestinal microbiota of these two groups. Primers were designed to quantify the 16S rRNA gene from dominant and subdominant bacterial groups. TaqMan^® probes were defined for the qPCR technique used for dominant microbiota. Human faecal microbiota was compared with that of farm animals using faecal samples collected from rabbits, goats, horses, pigs, sheep and cows. Three dominant bacterial groups ( Bacteroides/Prevotella, Clostridium coccoides and Bifidobacterium ) of the human microbiota showed differential population levels in animal species. The Clostridium leptum group showed the lowest differences among human and farm animal species. Human subdominant bacterial groups were highly variable in animal species. Partial least squares regression indicated that the human microbiota could be distinguished from all farm animals studied. This culture-independent comparative assessment of the faecal microbiota between humans and farm animals will prove useful in identifying biomarkers of human and animal faecal contaminations that can be applied to microbial source tracking methods.     

Indigenous bacteria and bacterial metabolic products in the gastrointestinal tract of broiler chickens

The gastrointestinal tract is a dynamic ecosystem containing a complex microbial community. In this paper, the indigenous intestinal bacteria and the microbial fermentation profile particularly short chain fatty acids (SCFA), lactate, and ammonia concentrations are reviewed. The intestinal bacterial composition changes with age. The bacterial density of the small intestine increases with age and comprises of lactobacilli, streptococci, enterobacteria, fusobacteria and eubacteria. Strict anaerobes (anaerobic gram-positive cocci, Eubacterium spp., Clostridium spp., Lactobacillus spp., Fusobacterium spp. and Bacteroides) are predominating caecal bacteria in young broilers. Data from culture-based studies showed that bifidobacteria could not be isolated from young birds, but were recovered from four-week-old broilers. Caecal lactobacilli accounted for 1.5-24% of the caecal bacteria. Gene sequencing of caecal DNA extracts showed that the majority of bacteria belonged to Clostridiaceae. Intestinal bacterial community is influenced by the dietary ingredients, nutrient levels and physical structure of feed. SCFA and other metabolic products are affected by diet formulation and age. Additional studies are required to know the bacterial metabolic activities together with the community analysis of the intestinal bacteria. Feed composition and processing have great potential to influence the activities of intestinal bacteria towards a desired direction in order to support animal health, well-being and microbial safety of broiler meat.     

Cultivable bacterial diversity from the human colon

Knowledge of the composition of the colonic microbiota is important for our understanding of how the balance of these microbes is influenced by diet and the environment, and which bacterial groups are important in maintaining gut health or promoting disease. Molecular methodologies have advanced our understanding of the composition and diversity of the colonic microbiota. Importantly, however, it is the continued isolation of bacterial representatives of key groups that offers the best opportunity to conduct detailed metabolic and functional studies. This also permits bacterial genome sequencing which will accelerate the linkage to functionality. Obtaining new human colonic bacterial isolates can be challenging, because most of these are strict anaerobes and many have rather exact nutritional and physical requirements. Despite this many new species are being isolated and described that occupy distinct niches in the colonic microbial community. This review focuses on these under-studied yet important gut anaerobes.     

Effects of two probiotic Lactobacillus strains on jejunal and cecal microbiota of broiler chicken under acute heat stress condition as revealed by molecular analysis of 16S rRNA genes

We examined the effects of probiotic Lactobacillus strains of Lactobacillus agilis JCM 1048 and Lactobacillus salivarius subsp. salicinius JCM 1230 on jejunal and cecal microbiota of broiler chicken under heat stress condition using terminal restriction fragment length polymorphism (T-RFLP) analysis. The jejunal bacterial community was limited to a few bacterial groups, mostly Lactobacillus spp. A relatively abundant and higher prevalence of Lactobacillus spp. were observed in the jejunal and cecal microbiota of the probiotic chickens compared with those of the control chickens under heat stress condition. In general, the probiotic strains did not significantly affect the abundance of L. agilis and L. salivarius in chicken intestine but clearly contributed to increasing their prevalence in the probiotic chickens. The probiotic Lactobacillus strains enriched the diversity of Lactobacillus flora in chicken jejunum and cecum by increasing the abundance and prevalence of Lactobacillus spp. inhabiting the intestine. The richness of Lactobacillus species tended to be similar among the jejunal and cecal microbiota. The bacterial community of cecum was complex and age-dependent. The major components of the cecal microbiota were clostridia and lactobacilli. The Clostridium subcluster XIVa was the most predominant group in chicken cecum. Probiotic Lactobacillus strains restored the microbial balance and maintained the natural stability of indigenous bacterial microbiota following heat stress-induced changes.     

Differences between the normal vaginal bacterial community of baboons and that of humans

Humans and baboons (Papio spp.) share considerable anatomical and physiological similarities in their reproductive tracts. Given the similarities, it is reasonable to expect that the normal vaginal microbial composition (microbiota) of baboons would be similar to that of humans. We have used a 16S rRNA phylogenetic approach to assess the composition of the baboon vaginal microbiota in a set of nine animals from a captive facility and six from the wild. Results show that although Gram‐positive bacteria dominate in baboons as they do in humans, there are major differences between the vaginal microbiota of baboons and that of humans. In contrast to humans, the species of Gram‐positive bacteria (Firmicutes) were taxa other than Lactobacillus species. In addition, some groups of Gram‐negative bacteria that are not normally abundant in humans were found in the baboon samples. A further level of difference was also seen even within the same bacterial phylogenetic group, as baboon strains tended to be more phylogenetically distinct from human strains than human strains were with each other. Finally, results of our analysis suggests that co‐evolution of microbes and their hosts cannot account for the major differences between the microbiota of baboons and that of humans because divergences between the major bacterial genera were too ancient to have occurred since primates evolved. Instead, the primate vaginal tracts appear to have acquired discrete subsets of bacteria from the vast diversity of bacteria available in the environment and established a community responsive to and compatible with host species physiology. Am. J. Primatol. 73:119–126, 2011. © 2010 Wiley‐Liss, Inc.     

Gut microbiota composition of Japanese macaques associates with extent of human encroachment

In recent decades, human–wildlife interaction and associated anthropogenic food provisioning has been increasing and becoming more severe due to fast population growth and urban development. Noting the role of the gut microbiome in host physiology like nutrition and health, it is thus essential to understand how human–wildlife interactions and availability of anthropogenic food in habitats can affect an animal's gut microbiome. This study, therefore, set out to examine the gut microbiota of Japanese macaques (Macaca fuscata) with varying accessibility to anthropogenic food and the possibility of using gut microbiota as indicator for macaques’ reliance on anthropogenic food. Using 16S ribosomal RNA gene sequencing, we described the microbial composition of Japanese macaques experiencing different types of human disturbance and anthropogenic food availability—captive, provisioned, crop‐raiding, and wild. In terms of alpha diversity, our results showed that observed richness of gut microbiota did not differ significantly between disturbance types but among collection sites, whereas Shannon diversity index differed by both disturbance types and sites. In terms of beta diversity, captive populations harbored the most distinctive gut microbial composition, and had the greatest difference compared with wild populations. Whereas for provisioned and crop‐raiding groups, the macaques exhibited intermediate microbiota between wild and captive. We identified several potential bacterial taxa at different taxonomic ranks whose abundance potentially could help in assessing macaques’ accessibility to anthropogenic food. This study revealed the flexibility of the gut microbiome of Japanese macaques and provided possible indices based on the gut microbiome profile in assessing macaques’ accessibility to/reliance on anthropogenic foods.     

Understanding the effects of diet on bacterial metabolism in the large intestine

Recent analyses of ribosomal RNA sequence diversity have demonstrated the extent of bacterial diversity in the human colon, and have provided new tools for monitoring changes in the composition of the gut microbial community. There is now an excellent opportunity to correlate ecological niches and metabolic activities with particular phylogenetic groups among the microbiota of the human gut. Bacteria that associate closely with particulate material and surfaces in the gut include specialized primary degraders of insoluble substrates, including resistant starch, plant structural polysaccharides and mucin. Butyrate-producing bacteria found in human faeces belong mainly to the clostridial clusters IV and XIVa. In vitro and in vivo evidence indicates that a group related to Roseburia and Eubacterium rectale plays a major role in mediating the butyrogenic effect of fermentable dietary carbohydrates. Additional cluster XIVa species can convert lactate to butyrate, while some members of the clostridial cluster IX convert lactate to propionate. The metabolic outputs of the gut microbial community depend not only on available substrate, but also on the gut environment, with pH playing a major role. Better understanding of the colonic microbial ecosystem will help to explain and predict the effects of dietary additives, including nondigestible carbohydrates, probiotics and prebiotics.     

Forest gene diversity is correlated with the composition and function of soil microbial communities

The growing field of community and ecosystem genetics indicates that plant genotype and genotypic variation are important for structuring communities and ecosystem processes. Little is known, however, regarding the effects of stand gene diversity on soil communities and processes under field conditions. Utilizing natural genetic variation occurring in Populus spp. hybrid zones, we tested the hypothesis that stand gene diversity structures soil microbial communities and influences soil nutrient pools. We found significant unimodal patterns relating gene diversity to soil microbial community composition, microbial exoenzyme activity of a carbon-acquiring enzyme, and availability of soil nitrogen. Multivariate analyses indicate that this pattern is due to the correlation between gene diversity, plant secondary chemistry, and the composition of the microbial community that impacts the availability of soil nitrogen. Together, these data from a natural system indicate that stand gene diversity may affect soil microbial communities and soil processes in ways similar to species diversity (i.e., unimodal patterns). Our results further demonstrate that the effects of plant genetic diversity on other organisms may be mediated by plant functional trait variation.     

How Microbes Shape Their Communities? A Microbial Community Model Based on Functional Genes

Exploring the mechanisms of maintaining microbial community structure is important to understand biofilm development or microbiota dysbiosis. In this paper, we propose a functional gene-based composition prediction(FCP) model to predict the population structure composition within a microbial community. The model predicts the community composition well in both a low-complexity community as acid mine drainage(AMD) microbiota, and a complex community as human gut microbiota. Furthermore, we define community structure shaping(CSS) genes as functional genes crucial for shaping the microbial community. We have identified CSS genes in AMD and human gut microbiota samples with FCP model and find that CSS genes change with the conditions. Compared to essential genes for microbes, CSS genes are significantly enriched in the genes involved in mobile genetic elements, cell motility, and defense mechanisms, indicating that the functions of CSS genes are focused on communication and strategies in response to the environment factors. We further find that it is the minority, rather than the majority, which contributes to maintaining community structure. Compared to health control samples, we find that some functional genes associated with metabolism of amino acids, nucleotides, and lipopolysaccharide are more likely to be CSS genes in the disease group. CSS genes may help us to understand critical cellular processes and be useful in seeking addable gene circuitries to maintain artificial self-sustainable communities. Our study suggests that functional genes are important to the assembly of microbial communities.     

Effects of host genetics and environment on egg‐associated microbiotas in brown trout (Salmo trutta)

Recent studies found fish egg‐specific bacterial communities that changed over the course of embryogenesis, suggesting an interaction between the developing host and its microbiota. Indeed, single‐strain infections demonstrated that the virulence of opportunistic bacteria is influenced by environmental factors and host immune genes. However, the interplay between a fish embryo host and its microbiota has not been studied yet at the community level. To test whether host genetics affects the assemblage of egg‐associated bacteria, adult brown trout (Salmo trutta) were sampled from a natural population. Their gametes were used for full‐factorial in vitro fertilizations to separate sire from dam effects. In total, 2520 embryos were singly raised under experimental conditions that differently support microbial growth. High‐throughput 16S rRNA amplicon sequencing was applied to characterize bacterial communities on milt and fertilized eggs across treatments. Dam and sire identity influenced embryo mortality, time until hatching and composition of egg‐associated microbiotas, but no link between bacterial communities on milt and on fertilized eggs could be found. Elevated resources increased embryo mortality and modified bacterial communities with a shift in their putative functional potential. Resource availability did not significantly affect any parental effects on embryo performance. Sire identity affected bacterial diversity that turned out to be a significant predictor of hatching time: embryos associated with high bacterial diversity hatched later. We conclude that both host genetics and the availability of resources define diversity and composition of egg‐associated bacterial communities that then affect the life history of their hosts.     

Species-specific effects of epigeic earthworms on microbial community structure during first stages of decomposition of organic matter

Background

Epigeic earthworms are key organisms in organic matter decomposition because of the interactions they establish with microorganisms. The earthworm species and the quality and/or substrate availability are expected to be major factors influencing the outcome of these interactions. Here we tested whether and to what extent the epigeic earthworms Eisenia andrei, Eisenia fetida and Perionyx excavatus, widely used in vermicomposting, are capable of altering the microbiological properties of fresh organic matter in the short-term. We also questioned if the earthworm-induced modifications to the microbial communities are dependent on the type of substrate ingested.

Methodology/Principal Findings

To address these questions we determined the microbial community structure (phospholipid fatty acid profiles) and microbial activity (basal respiration and microbial growth rates) of three types of animal manure (cow, horse and rabbit) that differed in microbial composition, after being processed by each species of earthworm for one month. No differences were found between earthworm-worked samples with regards to microbial community structure, irrespective of type of manure, which suggests the existence of a bottleneck effect of worm digestion on microbial populations of the original material consumed. Moreover, in mesocosms containing cow manure the presence of E. andrei resulted not only in a decrease in bacterial and fungal biomass, but also in a reduced bacterial growth rate and total microbial activity, while no such reduction was found with E. fetida and P. excavatus.

Conclusions/Significance

Our results point to the species of earthworm with its associated gut microbiota as a strong determinant of the process shaping the structure of microbial communities in the short-term. This must nonetheless be weighed against the fact that further knowledge is necessary to evaluate whether the changes in the composition of microbiota in response to the earthworm species is accompanied by a change in the microbial community diversity and/or function.