Microbiota of the Gut-Lymph Node Axis: Depletion of Mucosa-Associated Segmented Filamentous Bacteria and Enrichment of Methanobrevibacter by Colistin Sulfate and Linco-Spectin in Pigs

Abstract

Microorganisms are translocated from the gut to lymphatic tissues via immune cells, thereby challenging and training the mammalian immune system. Antibiotics alter the gut microbiome and consecutively might also affect the corresponding translocation processes, resulting in an imbalanced state between the intestinal microbiota and the host. Hence, understanding the variant effects of antibiotics on the microbiome of gut-associated tissues is of vital importance for maintaining metabolic homeostasis and animal health. In the present study, we analyzed the microbiome of (i) pig feces, ileum, and ileocecal lymph nodes under the influence of antibiotics (Linco-Spectin and Colistin sulfate) using 16S rRNA gene sequencing for high-resolution community profiling and (ii) ileocecal lymph nodes in more detail with two additional methodological approaches, i.e., cultivation of ileocecal lymph node samples and (iii) metatranscriptome sequencing of a single lymph node sample. Supplementation of medicated feed showed a local effect on feces and ileal mucosa-associated microbiomes. Pigs that received antibiotics harbored significantly reduced amounts of segmented filamentous bacteria (SFB) along the ileal mucosa ( p = 0.048; 199.17-fold change) and increased amounts of Methanobrevibacter , a methanogenic Euryarchaeote in fecal samples ( p = 0.005; 20.17-fold change) compared to the control group. Analysis of the porcine ileocecal lymph node microbiome exposed large differences between the viable and the dead fraction of microorganisms and the microbiome was altered to a lesser extent by antibiotics compared with feces and ileum. The core microbiome of lymph nodes was constituted mainly of Proteobacteria . RNA-sequencing of a single lymph node sample unveiled transcripts responsible for amino acid and carbohydrate metabolism as well as protein turnover, DNA replication and signal transduction. The study presented here is the first comparative study of microbial communities in feces, ileum, and its associated ileocecal lymph nodes. In each analyzed site, we identified specific phylotypes susceptible to antibiotic treatment that can have profound impacts on the host physiological and immunological state, or even on global biogeochemical cycles. Our results indicate that pathogenic bacteria, e.g., enteropathogenic Escherichia coli , could escape antibiotic treatment by translocating to lymph nodes. In general ileocecal lymph nodes harbor a more diverse and active community of microorganisms than previously assumed.

Alterations of the Viable Ileal Microbiota of the Gut Mucosa-Lymph Node Axis in Pigs Fed Phytase and Lactic Acid-Treated Cereals

“…In taking into account that mucosaassociated bacteria utilize host mucin as the substrate (24), greater luminal substrate availability may explain the higher bacterial richness and total bacterial gene copy number in ileal digesta than in the mucosa. In contrasting previous studies (25)(26)(27), Clostridiaceae and Lactobacillaceae were the predominant taxa at all three ileal sites, implying the importance of the luminal bacteria for colonization of the mucosa and translocation into ICLNs in the present pigs. Previously, Kelly et al (25) reported a predominance of Helicobacteraceae and Campylobacteraceae at the ileal mucosa, which were less abundant in the current study.…”

“…Specific bacterial subsampling or translocation was also obvious for Methanobacteriaceae, whose abundance largely increased from digesta to the mucosa and into the ICLNs. Although this is in line with our recent findings (27), like for many other bacteria found in ICLNs, little is known about their metabolic role and impact on the host. Theoretically, Methanobacteriaceae may have benefited from cross-feeding relationships with eubacteria, as they obtain their energy for growth mostly from the reduction of CO 2 and formate with H 2 , or, in the case of Methanosphaera, from the reduction of methanol with H 2 (32).…”

“…Previously, Kelly et al (25) reported a predominance of Helicobacteraceae and Campylobacteraceae at the ileal mucosa, which were less abundant in the current study. Likewise, Mann et al (12) and Zwirzitz et al (27) found that the microbiome of ICLNs was mainly composed of Proteobacteria. Moreover, high abundances of Lactobacillus species in porcine ICLNs have been reported before (9).…”

“…Lactobacilli have been reported to hinder pathogens to adhere to the mucosa and to proliferate, with benefits for host gut health (29). Although their presence in ICLNs has been repeatedly described (9,27), little is known about whether Lactobacillus species exert similar effects in the ICLNs. Interestingly, bacterial taxa that are known to interact with lactobacilli in the gut lumen via cross-feeding, such as Lachnospiraceae and Veillonellaceae (30), were also enriched in ICLNs.…”

Bacteriological, cytological, and molecular investigation of Corynebacterium pseudotuberculosis, mycobacteria, and other bacteria in caseous lymphadenitis and healthy lymph nodes of slaughtered sheep

“…At present, molecular methods, e.g., PCR, genome sequencing, and MALDI-TOF MS enable definitive diagnosis, along with geographical distribution studies, virulence profile, and genetic diversity investigation of Corynebacterium species from domestic animals worldwide, including C. pseudotuberculosis strains [2,7,8,[31][32][33][34]. In the coming years, large-scale sequencing has seen used to investigate the microbiota of lymph nodes from humans [35] and animals (pigs) [36], with the aim to diagnose and differentiate agents/diseases; however, this next generation sequencing method has not yet been applied in the investigation of lymph nodes from sheep with and without lesions compatible with CL.…”