4. DISCUSSION

4.1 Analysis of gut microbiota composition in different states of Catharsius molossus

Gut microbiota, as the ”inner ecosystem” of the gut, plays a crucial role in insects’ digestive systems that rely on limited nutrients or hard-to-digest food sources(Tafesh-Edwards & Eleftherianos, 2023; Wang et al., 2020). We observed stable and enduring microbial community structures in the host, referred to as ”dominant microbial taxa.” The presence of such microbial communities is vital for maintaining ecological balance and host health(Chen et al., 2021). Studies have demonstrated that gut microbiota in insects likePachysoma MacLeay from arid coastal regions of southwestern Africa and flightless Thorectes lusitanicus contain dominant taxa such as Bacteroidetes, Firmicutes, Proteobacteria, and Actinobacteria(Franzini et al., 2016; Hernández et al., 2015). In our study, the predominant phyla in the gut of Catharsius molossus in both hungry and refeeding states were Proteobacteria, Firmicutes, Actinobacteria, and Bacteroidetes. Notably, Bacteroidetes, known as obligate or strict anaerobes, are challenging to culture in vitro, suggesting potential similarities in the dominant microbial taxa among different beetle species and their significant roles in host growth and development. After refeeding, the proportion of Firmicutes and Bacteroidetes increased, while Proteobacteria decreased. The consistent proportion of Actinobacteria may relate to the fiber-rich diet ofCatharsius molossus , as Firmicutes possesses genes involved in fermenting dietary fiber and interacts with intestinal mucosa, contributing to maintaining host equilibrium (Sun et al., 2022). Bacteroidetes, known for polysaccharide degradation and aiding dietary fiber energy release(Pereira et al., 2021), exhibited significant differences between hungry and refeeding states, indicating its potential role in polysaccharide metabolism that warrants further experimental validation (Vera-Ponce de León et al., 2020). Regarding the variation in Proteobacteria, previous studies have suggested that an increase in its proportion could lead to an elevated risk of diseases, serving as an indicator of microbial community disruption, and potentially even as a diagnostic marker for diseases (Rizzatti et al., 2017; Shin et al., 2015). Importantly, within the context of our study, we have observed a relatively higher abundance of Proteobacteria within the gut microbiota of Catharsius molossus during periods of dietary deprivation compared to the refeeding state. This observation provides suggestive insights that starvation could potentially induce an unfavorable physiological state, potentially contributing to the instability of the host’s gut microbiota ecology. Additionally, the relatively small proportion of Dysgonomonas in our study differs from Vagococcus and Romboutsia , which were dominant genera according to Suarez-Moo et al. (Suárez-Moo et al., 2020).Dysgonomonas has been associated with nitrogen fixation capabilities (Bar-Shmuel et al., 2020; Skrzypczak & Przybylski, 2022), and its higher abundance during starvation suggests a potential role in nitrogen fixation to sustain host survival. However, more research is needed to explore whether Dysgonomonas assimilates nitrogen into the beetle or its direct environment. In summary, hunger and refeeding affect gut microbiota diversity and variability in Catharsius molossus . Yet, the impact of changes in gut microbial abundance on host physiological functions and their mechanisms remains to be investigated. Our study provides valuable insights into the adaptability of gut microbiota in beetles under unique environments and their ecological functions.

4.2 Predictive analysis of Catharsius molossus gut microbiota functions

Utilizing PICRUSt, we conducted functional predictions, revealing a higher abundance of genes related to primary metabolic pathways and genetic information processing at the primary pathway level. At the tertiary pathway level, the gut microbiota of hungry Catharsius molossus showed higher gene abundances in pathways related to ansamycin biosynthesis, vancomycin group biosynthesis, and D-glutamine and D-glutamate metabolism, while refeeding state displayed elevated gene abundances in pathways related to ansamycin biosynthesis, valine, leucine, and isoleucine biosynthesis, and D-glutamine and D-glutamate metabolism. Ansaamycins are essential antibiotics, particularly against Mycobacterium tuberculosis(Skrzypczak & Przybylski, 2022). This suggests that Catharsius molossus may utilize its gut microbiota’s metabolic functions to interact with the environment, assisting in defense against pathogen and parasite infections, which may contribute to its unique adaptation to a diet of feces and living in such an environment. However, it’s important to note that while our functional predictions offer valuable clues, they are based on predictions and don’t fully represent the actual functions of gut microbiota. To delve into the precise roles of these gene functions inCatharsius molossus’ environmental adaptability, integration of multi-omics data is essential. Such comprehensive research would enhance our understanding of the physiological functions, interdependencies, and coordination mechanisms of gut microbiota, uncovering their complex functions and potential applications. This approach could facilitate further resource development and utilization in the context of gut microbiota.