Rodent Gut Bacteria Coexisting with an Insect Gut Virus in Parasitic Cysts: Metagenomic Evidence of Microbial Translocation and Co-adaptation in Spatially-Confined Niches

Abstract

In medicine, parasitic cysts or cysticerci (fluid-filled cysts, larval stage of tapeworms) are believed to be sterile (no bacteria), and therein, the treatment of cysticerci infestations of deep extra-intestinal tissues (e.g., brain) relies almost exclusively on the use of antiparasitic medications, and rarely antibiotics. To date, however, it is unclear why common post-treatment complications include abscessation. This study quantified the microbial composition of parasitic cyst contents in a higher-order rodent host, using multi-kingdom shotgun metagenomics, to improve our understanding of gut microbial translocation and adaptation strategies in wild environments. Analysis was conducted on DNA from two hepatic parasitic cysts (Hydatigera (Taeenia) taeniaeformis) in an adult vole mouse (Microtus arvalis), and from feces, liver, and peritoneal fluid of three other vole family members living in a vegetable garden in Ohio, USA. Bacterial metagenomics revealed the presence of gut commensal/opportunistic species, including Parabacteroides distasonis, Klebsiella variicola, Enterococcus faecium, and Lactobacillus acidophilus, inhabiting the cysts. Parabacteroides distasonis and other species were also present outside the cyst in the peritoneal fluid. Remarkably, viral metagenomics revealed various murine viral species, but unexpectedly, it detected an insect-origin virus from the army moth (Pseudaletia/Mythimna unipuncta) known as Mythimna unipuncta granulovirus A (MyunGV-A) in both cysts, and in one fecal and one peritoneal sample from two different voles, indicating survival of the insect virus and adaption in voles. Metagenomics also revealed a significantly lower probability of fungal detection in the cysts compared to other samples (peritoneal fluid, p<0.05; and feces p<0.05), with single taxon detection in each cyst for Malassezia and Pseudophaeomoniella oleicola. The samples with a higher probability of fungi were the peritoneal fluid. In conclusion, commensal/pathobiont bacterial species can inhabit parasitic tapeworm cysts, which needs to be considered during therapeutic decisions of cysticerci or other chronic disease scenarios where immune privileged and spatially restricted ecosystems with limited nutrients and minimal presence of immune cells could facilitate microbial adaptation, such as within gut wall cavitating micropathologies in Crohn's disease.

Keywords: Bacteroidota; CAVFT; Cavernous fistulous tracts; Hydatigera taeniaeformis; Klebsiella; Mythimna unipuncta granulovirus A; P distasonis; cysticercosis; metacestode.

Figure 1.. Overview of family of voles in this study and identification of the hepatic parasitic cyst as the larval stage of Hydatigera taeniaeformis tapeworm.

A) Contextualization of the clinical and ecological relevance of human cysticercosis, exemplified with Taenia solium. (CDC public domain image). B) Samples collected from the voles in this report. C) Schematic of origin of samples within metacestode (parasitic-cystic structure) for analysis and photomicrograph, illustrating distinctive microscopic segmentation of the parasite as indicative of an immature strobilocerus metacestode. Not detailed the parasite lacks visualized hooks and the overall size suggests that this cyst-like structure is an immature strobilocerus. D) Chromatogram after Sanger sequencing used for tapeworm identification as Hydatigera taeniaeformis illustrates pure DNA in the cyst samples tested in this study.

Figure 2.. Metagenomic analysis identifies reproducible commensal Bacteroidota and opportunistic Pseudomonadota (Enterobacteriaceae) in the parasitic cysts.

A) Relative abundance across samples. B) Comparison of bacteria in both cysts demonstrates consistent pattern of abundance among similar bacteria, including K. variicola, P. distasonis, and Bacteroidales.

Figure 3.. Metagenomic analysis of viruses revealed MyunGV-A in Hydatygera larval cysts and peritoneal fluid of wild voles.

A) Hierarchical clustering of samples based on abundance. Euclidean distances. B) Abundance bar plot. MyunGV-A in both cysts was 100%, with similar findings in the peritoneal fluid and feces of two voles.

Figure 4.. Overview of Hydatigera taeniaeformis lifecyle and metacestode larval cystic stages in rodents and our theory of how MyunGV-A viral DNA could reach and co-adapt with bacteria in tapeworm cysts.

A) Left panel. H. taeniaeformis responsible for cyst formation in the peritoneal cavity in rodents may serve as a conducive environment for the retention and replication of MyunGV-A and bacteria. The cysts may offer protection against external factors and create a microenvironment suitable for these microorganisms, facilitating evasion of the immune system. B) Right panel. Presumptive theory for acquisition of the insect virus by voles is presumed to have occurred indirectly through dietary sources e.g., ingestion of insects or vegetation carrying MyunGV-A contaminants. Steps in proposed cycle: i., MyunGV-A replicates in the midgut epithelial cells of armyworms; ii., vole consumes armyworm moth; iii., MyunGV-A is absorbed by via the brush border of the intestinal epithelial cells and into the bloodstream, disseminating to the cysts, or follows the H. taeniaeformis as it migrates through the gut wall to extra-intestinal tissues to start the cystic stage of the tapeworm.