The microbiota’s influence on host (patho) physiology has gained interest in the context of Gulf War Illness (GWI), a chronic disorder featuring dysregulation of the gut–brain–immune axis. This study examined short- and long-term effects of GWI-related chemicals on gut health and fecal microbiota and the potential benefits of Lacto-N-fucopentaose-III (LNFPIII) treatment in a GWI model. Male C57BL/6J mice were administered pyridostigmine bromide (PB; 0.7 mg/kg) and permethrin (PM; 200 mg/kg) for 10 days with concurrent LNFPIII treatment (35 μg/mouse) in a short-term study (12 days total) and delayed LNFPIII treatment (2×/week) beginning 4 months after 10 days of PB/PM exposure in a long-term study (9 months total). Fecal 16S rRNA sequencing was performed on all samples post-LNFPIII treatment to assess microbiota effects of GWI chemicals and acute/delayed LNFPIII administration. Although PB/PM did not affect species composition on a global scale, it affected specific taxa in both short- and long-term settings. PB/PM elicited more prominent long-term effects, notably, on the abundances of bacteria belonging to Lachnospiraceae and Ruminococcaceae families and the genus Allobaculum. LNFPIII improved a marker of gut health (i.e., decreased lipocalin-2) independent of GWI and, importantly, increased butyrate producers (e.g., Butyricoccus, Ruminococcous) in PB/PM-treated mice, indicating a positive selection pressure for these bacteria. Multiple operational taxonomic units correlated with aberrant behavior and lipocalin-2 in PB/PM samples; LNFPIII was modulatory. Overall, significant and lasting GWI effects occurred on specific microbiota and LNFPIII treatment was beneficial.

The American cockroach, Periplaneta americana, is a cosmopolitan insect notorious for thriving among humans undeterred by attempts to eliminate it. The traits that contribute to its ubiquity as an opportunistic pest, such as long lifespan, expansive neurosensory capacity, and nutritional flexibility, also make P. americana an excellent invertebrate model organism as evidenced by its long history in neuroscience and physiological research. Current genetic resources available for P. americana highlight its large, complex genome and richly diverse transcriptional capabilities, but fall short of producing a complete, chromosome-level genome. Here, we present a high-quality de novo genome assembly of a laboratory-raised adult female P. americana using a combination of high fidelity PacBio long reads and Hi-C sequencing. The final 3.23 Gb genome was assembled with chromosomal resolution into 17 scaffolds, consistent with previous karyotype analysis, and has a scaffold N50 of 188.1 Mb and genome Benchmarking Universal Single-Copy Ortholog (BUSCO) score of 99.7%. This assembly includes a chromosome that was missing from the previous reference genome for this species. Protein prediction and annotation were performed via the NCBI Eukaryotic Genome Annotation Pipeline, which identified 16,750 protein-coding genes and generated an annotation BUSCO score of 97.8%. Ortholog comparisons with available Blattodea assemblies highlight the expanded chemosensory and immune capabilities of P. americana compared to termite relatives. This genome assembly is a valuable tool for facilitating future research on the biology and evolution of this remarkable insect.

1 The macronutrient composition of a host’s diet shapes its gut microbial community, with dietary fiber in particular escaping host digestion to serve as a potent carbon source for gut microbiota. Despite widespread recognition of fiber’s importance to microbiome health, nutritional research often fails to differentiate hyper-processed fibers from cell-matrix derived intrinsic fibers, limiting our understanding of how individual polysaccharides influence the gut community. We use the American cockroach ( Periplaneta americana ) as a model system to dissect the response of complex gut microbial communities to diet modifications that are impossible to test in traditional host models. Here, we designed synthetic diets from lab-grade, purified ingredients to identify how the cockroach microbiome responds to six different carbohydrates (chitin, methylcellulose, microcrystalline cellulose, pectin, starch, xylan) in otherwise balanced diets. We show via 16S rRNA gene profiling that these synthetic diets reduce bacterial diversity and alter the phylogenetic composition of cockroach gut microbiota in a fiber-dependent manner, regardless of the vitamin and protein content of the diet. Comparisons with cockroaches fed whole-food diets reveal that synthetic diets induce blooms in common cockroach-associated taxa and subsequently fragment previously stable microbial correlation networks. Our research leverages an unconventional microbiome model system and customizable lab-grade artificial diets to shed light on how purified polysaccharides, as opposed to nutritionally complex intrinsic fibers, exert substantial influence over a normally stable gut community.

1 The American cockroach, Periplaneta americana , is a cosmopolitan insect notorious for thriving among humans undeterred by attempts to eliminate it. The traits that contribute to its ubiquity as an opportunistic pest, such as long lifespan, expansive neurosensory capacity, and nutritional flexibility, also make P. americana an excellent invertebrate model organism with a long history in neuroscience and physiological research. Current genetic resources available for P. americana highlight its large, complex genome and richly diverse transcriptional capabilities, but fall short of producing a complete, chromosome-level genome. Here, we present a high-quality de novo genome assembly of a laboratory-raised adult female P. americana using a combination of high fidelity PacBio long reads and Hi-C sequencing. The final 3.23 Gb genome was assembled with chromosomal resolution into 17 scaffolds, consistent with previous karyotype analysis, and has a scaffold N50 of 188.1 Mb and genome BUSCO score of 99.7%. This assembly includes a chromosome that was missing from the previous reference genome for this species. Protein prediction and annotation were performed via the NCBI Eukaryotic Genome Annotation Pipeline, which identified 16,780 protein-coding genes and generated an annotation BUSCO score of 97.8%. Ortholog comparisons with available Blattodea assemblies highlight the expanded chemosensory and immune capabilities of P. americana compared to termite relatives. This genome assembly is a valuable tool for facilitating future research on the biology and evolution of this remarkable insect.

The gut microbiome plays an active role in host health, producing gut metabolites that influence host digestive and immune function while also mediating microbial crosstalk. Dietary fiber is a major source of important fermentation byproducts that are generally implicated in gut community stability and host wellbeing, but dissecting microbe-specific contributions to polysaccharide metabolism in the context of a complex gut community is challenging with conventional model organisms. Using the American cockroach (Periplaneta americana) as a model omnivore, we use chemically-defined synthetic diets to identify how complex gut microbial communities respond to two of the most abundant plant polysaccharides, xylan and cellulose. To do so, we fed cockroaches synthetic diets containing one of these fibers or a mix of both in differing ratios. Through both 16S rRNA gene profiling and RNA-seq, we show that mixed fibers enrich for organisms characteristic of the source fibers as well as additional organisms only enriched in mixed-fiber diets. Through an organism-centric pangenome approach, we identify the impact of these fibers on gut microbiome activity. We found that gut communities responded strongly to xylan, with Bacteroidota belonging to Bacteroides, Dysgonomonas, and Parabacteroides producing xylan-active CAZymes at high levels. Multiple groups of Bacillota also responded strongly to a xylan diet, but appeared to act as cross-feeding secondary degraders, producing primarily xylosidases and transcripts associated with xylose utilization. In contrast, cellulose diets were associated with higher transcriptional activity among Fibrobacterota, which are typically a minor component of the cockroach gut microbiome but were the primary producers of CAZymes associated with cellulose and cellobiose degradation. These experiments provide new insight into gut microbial metabolism of these complex plant polysaccharides. Further, they highlight the utility of the cockroach model and synthetic diets to answer fundamental questions about gut microbial responses to different polysaccharides alone and in combination.