https://orcid.org/0009-0004-5029-741X
Figures
Abstract
Studies on the fecal microbiome of wild animals reveal valuable information on the feeding habits of the host and the possible roles of bacteria in digestion. In this work we characterized the fecal microbiota of seven male and seven female Myotis velifer bats using the V3-V4 regions of the 16S rRNA gene. Fecal samples were collected at the El Salitre cave in Mexico. We obtained 81 amplicon sequence variants, identifying four phyla, 12 families and 14 genera for females and seven phyla, 21 families and 26 genera for males. The phylum Synergistota is reported for the first time in bats. The most abundant phyla were Pseudomonadota and Fusobacteriota. Male feces showed a greater taxonomic richness than those from females. This study revealed that the fecal microbiota of M. velifer had a unique and more diverse composition compared to the microbiota reported for other bats. We identified 24 families and two abundant genera Cetobacterium and Haematospirillum in both males and females. Cetobacterium may produce vitamin B12 that is not produced by animals and Haematospirillum, which has been reported as an emerging human pathogen, may produce non-volatile organic acids. These genera had not been previously reported in the bat microbiota.
Citation: Arellano-Hernández HD, Montes-Carreto LM, Guerrero JA, Martinez-Romero E (2024) The fecal microbiota of the mouse-eared bat (Myotis velifer) with new records of microbial taxa for bats. PLoS ONE 19(12): e0314847. https://doi.org/10.1371/journal.pone.0314847
Editor: Beza Ramasindrazana, Institut Pasteur de Madagascar, MADAGASCAR
Received: June 20, 2024; Accepted: November 17, 2024; Published: December 5, 2024
Copyright: © 2024 Arellano-Hernández et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Data Availability: The sequence data are available in the NCBI BioProject number: PRJNA1120100, with BioSamples: SAMN41675229 to SAMN41675242, and SRA: SRS21518157 to SRS21518170. https://www.ncbi.nlm.nih.gov/bioproject/PRJNA1120100.
Funding: This study was carried out with the financial support of Universidad Nacional Autónoma de México, PAPIIT UNAM IN206124 grant to Esperanza Martinez-Romero. Leslie M. Montes-Carreto received a CONAHCyT Mexico 2022(1) postdoctoral fellowship (CVU: 667266). The funders were not involved in the study design, data collection and analysis, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
Introduction
Typically, mammals establish symbiotic relationships with microbes that inhabit their skin, mucous membranes, and gastrointestinal tract, among others [1]. The microbiota is defined as a community of microorganisms residing within a host organ or tissue. The intestinal microbiota is species-rich and may have diverse capabilities to produce vitamins, aminoacids, short-chain fatty acids (SCFAs), enzymes, and neuroactive molecules such as serotonin [2–4]. It will, therefore, have effects on the host’s health [5], physiology [2, 6], sex and reproductive success [7, 8] and lifestyle [6, 9]. Bacterial communities are influenced by the same factors leading to highly dynamic interactions [10]. The gut microbiome encodes enzymes such as hydrolases necessary for the degradation of ingested food [3, 11–13]. For example, gut bacteria in insectivorous bats provide chitinases to break down chitin, the main component of insect exoskeletons [13]. In hematophagous bats, proteases facilitate protein digestion, absorption and metabolism [14]. Studies of wildlife microbiota have recently gained popularity due to their importance in host health, evolution, and ecology [2, 11, 15]. Microbial ecology has been revolutionized by the development of culture-independent techniques using shotgun sequencing [16] for metagenomics or mass sequencing of rRNA amplicons [17, 18]. These advancements enable researchers to assess the composition, diversity, structure, and functionality of microorganisms associated with their hosts [5, 11, 19]. One way to explore the intestinal microbiota noninvasively is by studying the fecal microbiota [20]. Though it has been argued that such samples are not representative of the gastrointestinal tract [21], fecal sampling has proven to be the most convenient non-invasive method, as it allows for repeated sampling of individuals over time, ensuring rapid collection and intact preservation [20, 22].
Bats, as the second most diverse and ecologically relevant group of mammals after rodents [23], harbor a variety of microorganisms that can be either beneficial or pathogenic [24–27]. In Mexico, 142 bat species have been identified, 100 of which are insectivorous [28–30]. Studies show that bats play important roles in ecosystems such as plant pollination, seed dispersal [31, 32] and control of insect populations by consuming large numbers of nocturnal insects, even consuming up to 150% of their body weight in one night [33]. Therefore, by consuming large quantities of insects, including crop pests and vectors of various diseases of medical importance [34, 35], bats provide economic, social and health benefits. They also act as natural fertilizers due to the nutrient richness of their guano (feces), which contains nitrogen, phosphorus, and microorganisms that can act as bioremediators, nematicides, and fungicides [36]. In this way, guano can be used as a fertilizer in crop fields or horticulture, potentially leading to economic savings by reducing the use of chemical pesticides and promoting sustainable agricultural practices [34].
Compared to the microbiota of non-flying mammals, especially herbivores [11, 37], bat microbiota exhibits less taxonomic diversity [38, 39]. The bat microbiota is dominated by the metabolically diverse bacterial phyla Pseudomonadota, Bacillota and Bacteroidota [24, 26, 38, 40]. At lower taxonomic levels, the composition of the gut microbiota of bats seems to be determined by the diet with specific bacteria linked to dietary specialization such as insectivory, frugivory, nectarivory, carnivory, or sanguivory diets [22, 24, 38, 41–44], suggesting a potential role for bacteria in nutritional and ecological differentiation [38]. When comparing different bat species, phytophagous (frugivorous and nectarivorous) species tend to harbor genera such as Weissella, Ureaplasma, Klebsiella, Enterobacter, Escherichia, Enterococcus, and Fructobacillus [24, 45], whereas insectivorous species are characterized by genera such as Plesiomonas, Enterococcus, Lactobacillus, Bacillus, Lactococcus, Paeniclostridium, Undibacterium, Serratia and Yersinia [21, 24, 26, 42, 46, 47]. In addition, differences in microbial composition between the sexes have been observed in nectarivorous [8] and frugivorous [48] species.
The mouse-eared bat (Myotis velifer) has a wide distribution on the American continent (from northern Kansas to Honduras and Guatemala). It is classified as a species of least concern according to red list of the IUCN [49]. It plays an important ecological role as a pest controller, consuming large quantities of nocturnal insects such as Coleoptera, Homoptera, Diptera, Lepidoptera, Hemiptera, among others [50–52]. The goal of this study was to describe the fecal microbiota of Myotis velifer, evaluate its richness and abundance, and compare the microbial diversity between females and males and between sexually active and inactive males.
Materials and methods
Study site
The collection of the samples was at “El Salitre” cave, located in the municipality of Tlaltizapan in the state of Morelos, Mexico (18° 45’00” N and 99°11’24” W) at an altitude of 1,100 meters above sea level. It is surrounded by patches of deciduous forest alternating with secondary vegetation, croplands, and grasslands. The cave has an entrance ~1.8 m high and ~3.5 m wide and consists of three chambers with a total length of 225 m. The temperature inside the cave is 20–25°C and humidity is 79–99% [53].
Collection of individuals and feces
Access to the cave and the capture of bats was authorized by the Dirección General de Áreas Naturales Protegidas of Morelos, Mexico. We captured 14 Myotis velifer bats (seven males and seven females) with a 6 m long mist net placed at the cave entrance, between 10:00 p.m. and 11:30 p.m. on August 27, 2022, during the rainy season of the year. Each of the captured bats was placed individually in a clean, non-sterile blanket bag to ensure air circulation and prevent suffocation. The collection of feces began at 2:00 a.m., after having kept the bats in the individual bags for around two and a half hours. To avoid possible contamination between fecal, nitrile gloves were sanitized with 70% alcohol after handling each bat sample. Feces from each individual were stored in different sterile 1.5 ml Eppendorf tubes with DNA/RNA shield to preserve the DNA integrity at room temperature until arrival at the laboratory, where they were stored at -20°C until processing. In addition, standard morphological measurements of bats were taken for their taxonomic identification according to the Guide for the Identification of Mexican bats [54]: forearm length (electronic vernier in mm), body mass (with a 100 g spring balance), age category (juvenile or adult) estimated according to ossification of the wing bones (metacarpals and phalanges) and the condition in which they were found (males: scrotum, inguinal or abdominal testicles, females: lactating, pregnant or inactive). Finally, they were released at the entrance of the cave. We followed the protocols of the Animal Care and Use Committee of the American Society for Mammalogists [55] while handling bats.
DNA extraction from feces
DNA was extracted from 14 fecal samples using the Wizard Genomic DNA Purification kit (Promega, USA) following the manufacturer’s instructions. The negative control was nuclease-free water. From each of the stool samples collected in sterile 1.5 ml tubes, ~180 mg was placed in a new tube and macerated using a sterile pestle. Subsequently, the extracts were purified with the High Pure PCR Template Preparation Kit (Roche, Germany). DNA concentration was quantified using a NanoDrop spectrophotometer at wavelengths of 230, 260, and 280 nm and DNA was observed in 1.2% agarose gels.
Sequencing
The hypervariable regions (V3-V4) of the 16S rRNA molecular marker were amplified by PCR for each sample using specific primers 518 (5´-CCAGCAGCCGCGGTAATACG3´) F and 800 (5´TACCAGGGTATCTAATCC-3´) R [56]. The PCR reactions (50μL) contained 2μL of DNA, 25μL of Dream Taq PCR Master Mix (2X) (Thermo Scientific, USA), 20.5 μL of nuclease-free water, 0.25μL of bovine serum albumin (BSA), 1.25μL of dimethylsulfoxide (DMSO) and 0.5μL of each primer. Samples were amplified using a denaturation protocol at 94°C (5 min), followed by 30 cycles of 94°C (60 s), 53°C (60 s), and 72°C (60 s) and a final extension (72°C, 10 min). Finally, the amplified products were sequenced at Macrogen with Illumina NovaSeq 6000 paired-end 2x300 bp with reported adapters [56].
Bioinformatics analysis
The raw reads were processed using the FastQC v0.12.0 program to evaluate their quality. Once this process was completed, the Fastp v0.23.4 program [57] was used to clean the sequences, trim adapters, and filter by quality with a Quality Score ≥ Q28. Using the QIIME2-2023.9 tool [58], amplicon sequence variants (ASVs) were obtained using DADA2 [59]. Taxonomic assignment of ASVs was obtained using feature-classifier [60], classify-sklearn [40, 58] using full-length sequences from Greengenes2-2022.10 [18] as reference sequences. Samples with unclassified bacteria were not considered in the further analysis.
The relative abundance of microbial taxa was calculated by dividing the total number of sequences of each phylum, family, and genus, by the total number of sequences of all groups for each taxonomic level, to recognize the most abundant taxa from the of M. velifer microbiota [61].
Analysis of alpha and beta diversity
To ensure that the richness and diversity of the samples are representative of the fecal microbiota, we performed a coverage analysis of the different groups compared [11, 62].
Microbial richness and taxonomic diversity between males and females, and sexually active and inactive males, at the genus level were estimated using Hill numbers q (qD) [63], where q = 0 corresponds to species richness, q = 1 corresponds to the exponent of Shannon entropy (effective number of common elements), and q = 2 corresponds to the inverse of Simpson’s index (effective number of dominant elements [64, 65]. The qD diversity values, sample coverage, and their confidence intervals were obtained with the iNEXT package in R [66] using as criteria the maximum number of contigs of the samples (31,261 sequences) as the endpoint and a 95% confidence interval (CI) with 1,000 bootstraps to construct rarefaction curves. Variation in overall composition for phylogenetic diversity, beta- weighted diversity metric UniFrac [67] as well as principal coordinate analysis (PCoA) were estimated using diversity in QIIME2-2023.5 [58]. For taxonomic beta diversity, a similarity analysis (ANOSIM) was performed between groups (M. velifer males and females and sexually active and inactive males) based on the Bray-Curtis similarity metric and with 1000 permutations, using Past software v217b [68].
Results
The total number of bats collected was 14, of which seven were females and seven were males. The reproductive condition in which they were categorized was as follows: seven inactive females and four inactive males, and three active males with scrotal testicles. All individuals were adults.
A total of 1,162,492 raw reads were obtained from the 14 samples. After cleaning, 867,768 reads remained, preserving an average of 95% of the sequences per sample. After taxonomic assignment, 84 amplicon sequence variants (ASVs) with a length range of 423 bp were identified.
Myotis velifer fecal microbiota
We obtained a total of seven phyla, 24 families (S1 Fig) and 31 genera in the fecal microbiota of the vespertilionid M. velifer. In both male and female samples, bacterial genera showed 100% sample coverage for q = 0, q = 1 and q = 2 demonstrating that diversity is complete for each sample (Fig 1A and 1B). Then, the comparisons of the diversity were carried out directly according to their confidence intervals [62].
(A) Rarefaction curves based on sample coverage by sex. (B) Rarefaction curves based on sample coverage by active and inactive males.
Some of the most dominant genera were Cetobacterium (33%), Haematospirillum (16%), Paraclostridium (7%), Ammoniphilus (6%), Escherichia (5%), Dysgonomonas (4%), Entomobacter (3%), Caccocola (3%), Aeromonas (3%), Providencia (3%), Enterocloster (2%), Lactococcus (2%), Clostridium (2%), Saezia (1%), Morganella (1%), Plesiomonas (1%), Orbus (1%), Adiutrix (1%) and Vagococcus (1%) (Fig 2 and S2 Fig).
Percentage of relative abundance of the genera from domain Bacteria in the fecal microbiota of M. velifer. The genera are organized by relative abundance, with the most abundant at the top and the least abundant at the bottom.
Fecal microbiota by sex
In the fecal microbiota from females, 14 genera were found, the dominant ones were Cetobacterium (28%), Haematospirillum (25%), Paraclostridium (16%) and Dysgonomonas (9%) (Fig 3A). In males, 26 genera were found, nine of which are shared with the females. Among the most abundant were Cetobacterium (37%), Haematospirillum (10%), Ammoniphilus (9%) and Escherichia (8%) (Fig 3A).
(A) Relative abundances of the bacterial genera present in the fecal microbiota from M. velifer males and females. (B) Relative abundances of bacterial genera present in the fecal microbiota of reproductively active and inactive M. velifer males. The genera are organized by relative abundance, with the most abundant at the top and the least abundant at the bottom.
In the fecal microbiota of the reproductively active males, a total of seven phyla were found, with Pseudomonadota (39%), Bacillota (21%) and Synergistota (21%) being dominant. We identified 17 families, and the most abundant ones were Synergistaceae (21%), Acetobacteraceae (17%) and Lachnospiraceae (14%) as well as 19 genera, Caccocola (21%), Entomobacter (17%), Enterocloster (14%) and Saezia (8%) as the most abundant (Fig 3B). As for sexually inactive males, a total of three phyla were found, with Fusobacteriota (51%), Pseudomonadota (35%), Bacillota (14%) as the dominant ones. Nine families were identified, among the most abundant were Fusobacteriaceae (51%), Enterobacteriaceae (14%) and Paenibacillaceae (13%); a richness of 11 bacterial genera was observed, of which four are shared with the active ones. The most abundant genera for these individuals were Cetobacterium, Ammoniphilus, Haematospirillum and Escherichia (51%, 13%, 13%, 12%, respectively) (Fig 3B and S3 Fig).
Alpha diversity analysis
The richness of males (q0 = 40) was twice as high as that of females (q0 = 21), while the equitability (q1 female = 11.24, q1 male = 13.44) and dominance (q2 female = 8.76, q2 male = 7.76) of species was similar in both sexes (Fig 4A).
(A) Rarefaction curves based on the number of sequences per sex. (B) Rarefaction curves based on the number of sequences per sexual condition in males (active and inactive).
The richness of sexually active males (q0 = 19) was twice that of sexually inactive males (q0 = 11), while the equitability (q1 active = 11.28, q1 inactive = 4.69) and dominance (q2 active = 8.69, q2 inactive = 3.25) of sexually active males was three times that of sexually inactive males (Fig 4B).
Beta diversity analysis
Principal coordinate analysis (PCoA) of phylogenetic beta diversity using weighted Unifrac distance as a quantitative measure of dissimilarity, revealed no differences between the fecal microbiota of males and females of the M. velifer (Fig 5A). Additionally, PCoA of phylogenetic beta diversity indicated no differences in the fecal microbiota of sexually active versus inactive males (Fig 5B).
(A) PCoA of the beta diversity present in the fecal microbiota from M. velifer males and females, using weighted unifrac distance. (B) PCoA of the beta diversity present in the fecal microbiota from M. velifer sexually active and inactive males, using weighted unifrac distance.
The analysis of similarity between groups (ANOSIM) indicated no significant differences in beta diversity between male and female groups (r = 0.08267, p = 0.7543; Fig 6A). Similarly, ANOSIM analysis revealed no significant differences in beta diversity between sexually active and inactive males (r = 0.2778, p = 0.2052; Fig 6B).
(A) Similarity analysis between males (blue) and females (pink) (ANOSIM). (B) Similarity analysis between sexually active (blue) and inactive (red) males (ANOSIM).
Discussion
There are several studies on the fecal microbiota of wild insectivorous bats that may be summarized as follows: (i) the most studied bat genera have been Myotis, Rhinolophus and Hipposideros, (ii) the bacterial phyla that dominate are Pseudomonadota and Bacillota, (iii) the main bacterial genera reported have been Pleisomonas, Enterococcus, Lactococcus and Lactobacillus, and iv) most of the feces collected for these studies belong to bats distributed in China [21, 24, 26, 40, 46, 47]. However, to our knowledge, this is the first investigation to characterize the fecal microbiota of M. velifer, an insectivorous bat distributed mainly in Mexico. The mouse-eared bat establishes symbiotic relationships with communities of microorganisms (such as bacteria, archaea and fungi) [3] that make up its gut microbiota.
Here we report that the fecal microbiota of the bat M. velifer is dominated by the phyla Pseudomonadota, Fusobacteriota, Bacillota and Synergistota. This finding is consistent with previous studies of the intestinal and fecal microbiota found for other bats. For example, in the gut microbiota of phytophagous (nectarivorous and frugivorous) and insectivorous bats from southern China, the microbiota was dominated by Pseudomonadota, Bacillota and Bacteroidota [24, 26]. In lesser horseshoe bats Pseudomonadota dominates, followed by Bacillota [47]. The fecal bacterial microbiota of the insectivorous bat Mops condylurus had Bacillota and Pseudomonadota [40] and in two insectivorous bats (Rhinolophus sinicus and Myotis altarium) in three different sampling sources (small intestine, large intestine and feces) the most dominant phylum was Pseudomonadota, while Fusobacteriota was the least dominant [21]. Unlike the results obtained from the research of Wu and collaborators [21], in this study we found that the phylum Fusobacteriota was the second dominant phylum with 23% of the fecal microbiota of M. velifer.
This study is the first report of the Synergistota phylum in bat microbiota. This phylum, which comprised 5% of the total fecal microbiota of M. velifer, has been described only in the intestinal tract of pigs [69], termites [70] and in the human mouth [71]. This group of bacteria is characterized by anaerobic degraders of amino acids [72–74].
Regarding the composition at the family level, 24 families were identified, with Fusobacteriaceae, Rhodospirillaceae and Enterobacteriaceae being the most abundant. Fusobacteriaceae and Enterobacteriaceae are characterized by being potentially opportunistic pathogens and are reported in different proportions in the oral, gastrointestinal [21, 75] and fecal microbiota of bats [24]. On the other hand, Rhodospirillaceae has not been reported in bats, and this group of bacteria has a wide variety of habitats, ranging from plant tissues, soil contaminated with oil to aquatic environments such as oceans and stagnant water [76].
Our results highlight the dominance of the genus Cetobacterium in the microbiota of M. velifer. It has also been reported as a dominant genus in the fecal microbiota of freshwater fish, where they function as producers of vitamin B12, which is distributed throughout the intestinal tract [77, 78]. Therefore, it can be speculated that they perform the same function in the intestinal tract of M. velifer, since this vitamin is not synthesized by animals or plants, but by some bacteria and archaea [79]. It is worth mentioning that this genus has been isolated from feces of marine mammals [80] and humans [81]. Future studies should focus on the role of Cetobacterium in bats and the implications it has for their ecology.
The second dominant genus from the M. velifer fecal microbiota is Haematospirillum, isolated for the first time in human blood [82] and later found in the blood of an insectivorous bird [83]. It is described as a possible emerging infectious pathogen for humans [82, 84, 85]. Our finding constitutes the first molecular evidence of this species in the fecal microbiota of bats. Haematospirillum may produce non-volatile organic acids and may be involved in degrading organic substrates containing sulfur [82, 86, 87].
Regarding reproductively active and inactive males, differences in composition and relative abundances were observed as described before in the fecal microbiota of different reproductive stages of the nectivorous bat Leptonycteris yerbabuenae [8]. Other bacterial genera in the fecal microbiota of active and inactive individuals were Caccocola which was identified through metagenomic analyzes in chicken feces [88], Entomobacter, isolated from the intestine of the Madagascar hissing cockroach (Gromphadorhina portentosa) [89], Enterocloster bacteria described as opportunistic pathogens and commonly found in the intestine of humans [90] and Saezia, belonging to the order Burkholderiales [91]. These bacterial genera are newly described here for bats in the Americas.
From males, we observed seven phyla, that included 21 of the 24 total families and 26 of the 31 total genera, that is, 84% of the richness described for the fecal microbiota of the bat M. velifer. On the other hand, in females, four of seven phyla, 12 of 24 families and 14 of 31 genera were identified, which is equivalent to 45% of the total richness characterized. It should be noted that Pseudomonadota was the most abundant phylum consistent with the results of Banskar [45]. Fusobacteriaceae and Cetobacterium were the dominant family and genus for Myotis velifer, however, between sexes, the relative abundance values change.
Alpha diversity analyses revealed significant differences between sexes and among male reproductive stages (active and inactive). Previous analyses of the microbiota of phyllostomid bats, comparing sexes and reproductive conditions within sexes, have generally shown that both reproductive males and females harbor the most diverse bacterial communities [8, 48]. However, in our study, reproductively active males exhibited a more diverse fecal microbiota compared to both females and inactive males. According to Riopelle [48], this finding underscores the urgent need for further studies on bacteria not shared by male and female bats to clarify sex differences.
In contrast to the alpha diversity analyses, the beta diversity analyses (UNIFRAC, PCoA and ANOSIM) indicated no significant differences between the sexes or between sexually active and inactive males. This lack of beta diversity differences may suggest that, although variation exists in the abundance and distribution of bacterial genera across groups, they share a fecal microbiota with a similar composition. This could indicate the presence of a stable bacterial community that does not differ structurally at the genus level, likely due to shared factors such as diet, environment, and lifestyle [92].
Conclusions
The detailed study on the fecal microbiota of the Myotis velifer bat using massive amplicon sequencing techniques revealed a diverse and distinctive composition compared to the microbiota from other previously studied bats. The dominance of Pseudomonadota, Fusobacteriota, Bacillota and Synergistota, together with the identification of 33 families and the prevalence of genera such as Cetobacterium and Haematospirillum revealed the uniqueness of the microbiota of this insectivore which has an extensive geographical distribution.
Although some similarities were observed with previous studies in bats, such as the prevalence of Pseudomonadota, this study highlights notable differences, such as the significant presence of Fusobacteriota. On the other hand, the identification of differences in the composition of the microbiota between males and females of M. velifer is notable, despite the absence of significant differences in beta diversity analyses.
Supporting information
S1 Fig. Bacterial families from the fecal microbiota of Myotis velifer per sex.
Percentage of relative abundance of the families from domain Bacteria in the fecal microbiota of M. velifer, with the most abundant at the top and the least abundant at the bottom.
https://doi.org/10.1371/journal.pone.0314847.s001
(TIF)
S2 Fig. Bacterial genera per individual found in the fecal microbiota of Myotis velifer.
Percentage of relative abundance of the genera from domain Bacteria in the fecal microbiota of M. velifer, with the most abundant at the top and the least abundant at the bottom.
https://doi.org/10.1371/journal.pone.0314847.s002
(TIF)
S3 Fig. Sexually active and inactive males (per individual).
Percentage of relative abundance of the genera from domain Bacteria in active and inactive males, with the most abundant at the top and the least abundant at the bottom.
https://doi.org/10.1371/journal.pone.0314847.s003
(TIF)
Acknowledgments
We thank to Luis Gerardo Ávila Torresagatón, Daniela Michel Carreño Ochoa, Alma Itzel Reyna Salazar and Emmanuel C. Paniagua Domínguez at FCB-UAEM for their help in the field work, as well as Osiris Gaona Pineda of IE-UNAM for her helpful comments on the manuscript and Michael F. Dunn of CCG-UNAM for critical reading of the manuscript. We thank CCG-UNAM for giving us access to its computer cluster and Julio Cesar Martínez-Romero for technical support.
References
- 1. Muegge BD, Kuczynski J, Knights D, Clemente JC, Gonzalez A, Fontana L, et al. Diet drives convergence in gut microbiome functions across mammalian phylogeny and within humans. Science. 2011;332(6032):970–4. pmid:21596990; PubMed Central PMCID: PMC3303602.
- 2. Amato KR. Co-evolution in context: The importance of studying gut microbiomes in wild animals. Microbiome Science and Medicine. 2013;1(1).
- 3. Backhed F, Ley RE, Sonnenburg JL, Peterson DA, Gordon JI. Host-bacterial mutualism in the human intestine. Science. 2005;307(5717):1915–20. pmid:15790844.
- 4. Zhou M, Fan Y, Xu L, Yu Z, Wang S, Xu H, et al. Microbiome and tryptophan metabolomics analysis in adolescent depression: roles of the gut microbiota in the regulation of tryptophan-derived neurotransmitters and behaviors in human and mice. Microbiome. 2023;11(1):145. Epub 20230630. pmid:37386523; PubMed Central PMCID: PMC10311725.
- 5. Ley RE, Hamady M, Lozupone C, Turnbaugh PJ, Ramey RR, Bircher JS, et al. Evolution of mammals and their gut microbes. Science. 2008;320(5883):1647–51. pmid:18497261; PubMed Central PMCID: PMC2649005.
- 6. Sommer F, Backhed F. The gut microbiota—masters of host development and physiology. Nat Rev Microbiol. 2013;11(4):227–38. Epub 20130225. pmid:23435359.
- 7. Edwards SM, Cunningham SA, Dunlop AL, Corwin EJ. The Maternal Gut Microbiome During Pregnancy. MCN Am J Matern Child Nurs. 2017;42(6):310–7. pmid:28787280; PubMed Central PMCID: PMC5648614.
- 8. Gaona O, Gomez-Acata ES, Cerqueda-Garcia D, Neri-Barrios CX, Falcon LI. Fecal microbiota of different reproductive stages of the central population of the lesser-long nosed bat, Leptonycteris yerbabuenae. PLoS One. 2019;14(7):e0219982. Epub 2019/07/19. pmid:31318946; PubMed Central PMCID: PMC6639036.
- 9. Mosca A, Leclerc M, Hugot JP. Gut Microbiota Diversity and Human Diseases: Should We Reintroduce Key Predators in Our Ecosystem? Front Microbiol. 2016;7:455. Epub 20160331. pmid:27065999; PubMed Central PMCID: PMC4815357.
- 10. Degnan PH, Pusey AE, Lonsdorf EV, Goodall J, Wroblewski EE, Wilson ML, et al. Factors associated with the diversification of the gut microbial communities within chimpanzees from Gombe National Park. Proc Natl Acad Sci U S A. 2012;109(32):13034–9. Epub 20120723. pmid:22826227; PubMed Central PMCID: PMC3420156.
- 11. Montes-Carreto LM, Aguirre-Noyola JL, Solís-García IA, Ortega J, Martinez-Romero E, Guerrero JA. Diverse methanogens, bacteria and tannase genes in the feces of the endangered volcano rabbit (Romerolagus diazi). PeerJ. 2021;9. pmid:34458021
- 12. Scott KP, Gratz SW, Sheridan PO, Flint HJ, Duncan SH. The influence of diet on the gut microbiota. Pharmacol Res. 2013;69(1):52–60. Epub 20121109. pmid:23147033.
- 13. Whitaker JO, Dannelly HK, Prentice DA. Chitinase in Insectivorous Bats. J Mammal. 2004;85(1):15–8.
- 14. Muller HE, Pinus M, Schmidt U. Aeromonas hydrophila as a normal intestinal bacterium of the vampire bat (Desmodus rotundus). Zentralbl Veterinarmed B. 1980;27(5):419–24. pmid:7053178.
- 15. Ley RE, Lozupone CA, Hamady M, Knight R, Gordon JI. Worlds within worlds: evolution of the vertebrate gut microbiota. Nat Rev Microbiol. 2008;6(10):776–88. pmid:18794915; PubMed Central PMCID: PMC2664199.
- 16. Franzosa EA, Morgan XC, Segata N, Waldron L, Reyes J, Earl AM, et al. Relating the metatranscriptome and metagenome of the human gut. Proc Natl Acad Sci U S A. 2014;111(22):E2329–38. Epub 20140519. pmid:24843156; PubMed Central PMCID: PMC4050606.
- 17. McDonald D, Hyde E, Debelius JW, Morton JT, Gonzalez A, Ackermann G, et al. American Gut: an Open Platform for Citizen Science Microbiome Research. mSystems. 2018;3(3). Epub 20180515. pmid:29795809; PubMed Central PMCID: PMC5954204.
- 18. McDonald D, Jiang Y, Balaban M, Cantrell K, Zhu Q, Gonzalez A, et al. Greengenes2 enables a shared data universe for microbiome studies. Biorxiv. 2023.
- 19. Martinez-Romero E, Aguirre-Noyola JL, Bustamante-Brito R, Gonzalez-Roman P, Hernandez-Oaxaca D, Higareda-Alvear V, et al. We and herbivores eat endophytes. Microb Biotechnol. 2021;14:1282–129. Epub 2020/12/16. pmid:33320440.
- 20. Ingala MR, Simmons NB, Wultsch C, Krampis K, Speer KA, Perkins SL. Comparing Microbiome Sampling Methods in a Wild Mammal: Fecal and Intestinal Samples Record Different Signals of Host Ecology, Evolution. Front Microbiol. 2018;9:803. Epub 2018/05/17. pmid:29765359; PubMed Central PMCID: PMC5938605.
- 21. Wu H, Xing Y, Sun H, Mao X. Gut microbial diversity in two insectivorous bats: Insights into the effect of different sampling sources. Microbiologyopen. 2019;8(4):e00670. Epub 2018/07/05. pmid:29971963; PubMed Central PMCID: PMC6530527.
- 22. Carrillo-Araujo M, Tas N, Alcantara-Hernandez RJ, Gaona O, Schondube JE, Medellin RA, et al. Phyllostomid bat microbiome composition is associated to host phylogeny and feeding strategies. Front Microbiol. 2015;6:447. Epub 20150519. pmid:26042099; PubMed Central PMCID: PMC4437186.
- 23.
Simmons NB, Cirranello AL. Bats of the World: A Taxonomic and Geographic Database: Zenodo; 2024.
- 24. Li J, Li L, Jiang H, Yuan L, Zhang L, Ma JE, et al. Fecal Bacteriome and Mycobiome in Bats with Diverse Diets in South China. Curr Microbiol. 2018;75(10):1352–61. Epub 20180619. pmid:29922970.
- 25. Muhldorfer K. Bats and bacterial pathogens: a review. Zoonoses Public Health. 2013;60(1):93–103. Epub 20120802. pmid:22862791.
- 26. Sun DL, Gao YZ, Ge XY, Shi ZL, Zhou NY. Special Features of Bat Microbiota Differ From Those of Terrestrial Mammals. Front Microbiol. 2020;11:1040. Epub 20200603. pmid:32582057; PubMed Central PMCID: PMC7284282.
- 27. Wang LF, Walker PJ, Poon LL. Mass extinctions, biodiversity and mitochondrial function: are bats ’special’ as reservoirs for emerging viruses? Curr Opin Virol. 2011;1(6):649–57. Epub 20111109. pmid:22440923; PubMed Central PMCID: PMC7102786.
- 28.
Ceballos G, Oliva G. Los mamiferos silvestres de Mexico. 1 ed: Fondo de Cultura Económica.; 2005. 1000 p.
- 29. Coates R, Ramírez-Lucho I, González-Christen A. Una lista actualizada de los murciélagos de la región de Los Tuxtlas, Veracruz. Revista Mexicana de Biodiversidad. 2017;88(2):349–57.
- 30. Medellín R, Gaona O. Hechos en casa: Investigaciones de Frontera: Los murciélagos, los animales más calumniados y maItratados en México y en el mundo. Oikos. 2010;1:11–3.
- 31. Irving AT, Ahn M, Goh G, Anderson DE, Wang LF. Lessons from the host defences of bats, a unique viral reservoir. Nature. 2021;589(7842):363–70. Epub 20210120. pmid:33473223.
- 32.
Jones KE. Chiroptera (Bats). In: Wiley J, Sons, editors. eLS Evolution & Diversity of Life2005.
- 33.
Hutson AM, Mickleburgh SP, Racey PA. Microchiropteran bats: global status survey and conservation action plan. Gland, Switzerland and Cambridge, UK2001. 258 p.
- 34. Gandara G, Sandoval A, Cienfuegos C. Valoración económica de los servicios ecológicos que prestan los murciélagos “Tadarida brasiliensis” como controladores de plagas en el norte de México. 2006.
- 35. Russo D, Salinas-Ramos VB, Cistrone L, Smeraldo S, Bosso L, Ancillotto L. Do We Need to Use Bats as Bioindicators? Biology (Basel). 2021;10(8). Epub 20210721. pmid:34439926; PubMed Central PMCID: PMC8389320.
- 36. Zarate Martinez DG, Serrato Diaz A, Lopez-Wilchis. Importancia ecologica de los murcielagos. ContactoS. 2012;85:19–27.
- 37. de Jonge N, Carlsen B, Christensen MH, Pertoldi C, Nielsen JL. The Gut Microbiome of 54 Mammalian Species. Front Microbiol. 2022;13:886252. Epub 20220616. pmid:35783446; PubMed Central PMCID: PMC9246093.
- 38.
Ingala MR. Microbiomes of bats. In: Russo D, Fenton B, editors. A Natural History of Bat Foraging Evolution, Physiology, Ecology, Behavior, and Conservation: Academic Press; 2024. p. 217–32.
- 39. Song SJ, Sanders JG, Delsuc F, Metcalf J, Amato K, Taylor MW, et al. Comparative Analyses of Vertebrate Gut Microbiomes Reveal Convergence between Birds and Bats. mBio. 2020;11(1). Epub 20200107. pmid:31911491; PubMed Central PMCID: PMC6946802.
- 40. Edenborough KM, Mu A, Muhldorfer K, Lechner J, Lander A, Bokelmann M, et al. Microbiomes in the insectivorous bat species Mops condylurus rapidly converge in captivity. PLoS One. 2020;15(3):e0223629. Epub 20200320. pmid:32196505; PubMed Central PMCID: PMC7083271.
- 41. Aizpurua O, Nyholm L, Morris E, Chaverri G, Herrera Montalvo LG, Flores-Martinez JJ, et al. The role of the gut microbiota in the dietary niche expansion of fishing bats. Anim Microbiome. 2021;3(1):76. Epub 20211028. pmid:34711286; PubMed Central PMCID: PMC8555116.
- 42. Federici L, Masulli M, De Laurenzi V, Allocati N. An overview of bats microbiota and its implication in transmissible diseases. Front Microbiol. 2022;13:1012189. Epub 20221020. pmid:36338090; PubMed Central PMCID: PMC9631491.
- 43. Ingala MR, Becker DJ, Bak Holm J, Kristiansen K, Simmons NB. Habitat fragmentation is associated with dietary shifts and microbiota variability in common vampire bats. Ecol Evol. 2019;9(11):6508–23. Epub 20190509. pmid:31236240; PubMed Central PMCID: PMC6580296.
- 44. Ingala MR, Simmons NB, Dunbar M, Wultsch C, Krampis K, Perkins SL. You are more than what you eat: potentially adaptive enrichment of microbiome functions across bat dietary niches. Anim Microbiome. 2021;3(1):82. Epub 20211214. pmid:34906258; PubMed Central PMCID: PMC8672517.
- 45. Banskar S, Mourya DT, Shouche YS. Bacterial diversity indicates dietary overlap among bats of different feeding habits. Microbiol Res. 2016;182:99–108. Epub 20151021. pmid:26686618.
- 46. Dai W, Leng H, Li J, Li A, Li Z, Zhu Y, et al. The role of host traits and geography in shaping the gut microbiome of insectivorous bats. mSphere. 2024;9(4):e0008724. Epub 20240321. pmid:38509042; PubMed Central PMCID: PMC11036801.
- 47. Selvin J, Lanong S, Syiem D, De Mandal S, Kayang H, Kumar NS, et al. Culture-dependent and metagenomic analysis of lesser horseshoe bats’ gut microbiome revealing unique bacterial diversity and signatures of potential human pathogens. Microb Pathog. 2019;137:103675. Epub 20190829. pmid:31473248; PubMed Central PMCID: PMC7127535.
- 48. Riopelle JC, Shamsaddini A, Holbrook MG, Bohrnsen E, Zhang Y, Lovaglio J, et al. Sex differences and individual variability in the captive Jamaican fruit bat (Artibeus jamaicensis) intestinal microbiome and metabolome. Sci Rep. 2024;14(1):3381. Epub 20240209. pmid:38336916; PubMed Central PMCID: PMC10858165.
- 49. Solari S. IUCN Red List of Threatened Species: Myotis velifer. IUCN Red List Threat Species 2018.
- 50. Fitzgerald KV, Ammerman LK, Ortega J. Cave Myotis (Myotis velifer) consume diverse prey items and provide important ecosystem services. J Mammal. 2024.
- 51. Kunz TH. Feeding Ecology of a Temperate Insectivorous Bat (Myotis Velifer). Ecology. 1974;55(4):693–711.
- 52. Marquardt SR, Choate JR. Influence of Thermal Environment on Food Habits of Female Cave Myotis (Myotis velifer). The Southwestern Naturalist. 2009;54(2):166–75.
- 53.
Ávila-Torresagatón LG, Fuentes-Vargas L. La cueva El Salitre como refugio de vida silvestre. In: Gómez Hernández CV, NCK C., López Higareda , Cruz Medina J, Melgarejo ED, editors. La biodiversidad en Morelos Estudio de Estado 2. III. Ciudad de México: Comisión Nacional para el Conocimiento y Uso de la Biodiversidad; 2020. p. 280–4.
- 54.
Medellín R, Arita H, Sanchez O. Identificación de los murciélagos de México. Clave de campo. Segunda Edición ed: Instituto de Ecología. Universidad Nacional Autónoma de México.; 2008.
- 55. Sikes RS, Animal C, Use Committee of the American Society of M. 2016 Guidelines of the American Society of Mammalogists for the use of wild mammals in research and education. J Mammal. 2016;97(3):663–88. Epub 2016/06/09. pmid:29692469; PubMed Central PMCID: PMC5909806.
- 56. Klindworth A, Pruesse E, Schweer T, Peplies J, Quast C, Horn M, et al. Evaluation of general 16S ribosomal RNA gene PCR primers for classical and next-generation sequencing-based diversity studies. Nucleic Acids Res. 2013;41(1):e1. Epub 20120828. pmid:22933715; PubMed Central PMCID: PMC3592464.
- 57. Chen S, Zhou Y, Chen Y, Gu J. fastp: an ultra-fast all-in-one FASTQ preprocessor. Bioinformatics. 2018;34(17):i884–i90. Epub 2018/11/14. pmid:30423086; PubMed Central PMCID: PMC6129281.
- 58. Bolyen E, Rideout JR, Dillon MR, Bokulich NA, Abnet CC, Al-Ghalith GA, et al. Reproducible, interactive, scalable and extensible microbiome data science using QIIME 2. Nat Biotechnol. 2019;37(8):852–7. pmid:31341288; PubMed Central PMCID: PMC7015180.
- 59. Callahan BJ, McMurdie PJ, Rosen MJ, Han AW, Johnson AJ, Holmes SP. DADA2: High-resolution sample inference from Illumina amplicon data. Nat Methods. 2016;13(7):581–3. Epub 20160523. pmid:27214047; PubMed Central PMCID: PMC4927377.
- 60. Bokulich NA, Kaehler BD, Rideout JR, Dillon M, Bolyen E, Knight R, et al. Optimizing taxonomic classification of marker-gene amplicon sequences with QIIME 2’s q2-feature-classifier plugin. Microbiome. 2018;6(1):90. Epub 20180517. pmid:29773078; PubMed Central PMCID: PMC5956843.
- 61. Neu AT, Allen EE, Roy K. Defining and quantifying the core microbiome: Challenges and prospects. Proc Natl Acad Sci U S A. 2021;118(51). pmid:34862327; PubMed Central PMCID: PMC8713806.
- 62. Chao A, Jost L. Coverage-based rarefaction and extrapolation: standardizing samples by completeness rather than size. Ecology. 2012;93(12):2533–47. Epub 2013/02/26. pmid:23431585.
- 63. Chiu CH, Chao A. Estimating and comparing microbial diversity in the presence of sequencing errors. PeerJ. 2016;4:e1634. Epub 2016/02/09. pmid:26855872; PubMed Central PMCID: PMC4741086.
- 64. Ma ZS, Li L. Measuring metagenome diversity and similarity with Hill numbers. Mol Ecol Resour. 2018;18(6):1339–55. Epub 2018/07/10. pmid:29985552.
- 65. Ma ZS, Li L, Gotelli NJ. Diversity-disease relationships and shared species analyses for human microbiome-associated diseases. ISME J. 2019;13(8):1911–9. Epub 2019/03/22. pmid:30894688; PubMed Central PMCID: PMC6775969.
- 66. Hsieh TC, Ma KH, Chao A, McInerny G. iNEXT: an R package for rarefaction and extrapolation of species diversity (Hill numbers). Methods Ecol Evol. 2016;7(12):1451–6.
- 67. Lozupone CA, Hamady M, Kelley ST, Knight R. Quantitative and qualitative beta diversity measures lead to different insights into factors that structure microbial communities. Appl Environ Microbiol. 2007;73(5):1576–85. Epub 20070112. pmid:17220268; PubMed Central PMCID: PMC1828774.
- 68. Hammer O, Harper DAT, Ryan PD. PAST: Paleontological statistics software package for education and data analysis Palaeontologia Electronica 2001;4(1):9pp.
- 69. Leser TD, Amenuvor JZ, Jensen TK, Lindecrona RH, Boye M, Moller K. Culture-independent analysis of gut bacteria: the pig gastrointestinal tract microbiota revisited. Appl Environ Microbiol. 2002;68(2):673–90. pmid:11823207; PubMed Central PMCID: PMC126712.
- 70. Hongoh Y, Ohkuma M, Kudo T. Molecular analysis of bacterial microbiota in the gut of the termite Reticulitermes speratus (Isoptera; Rhinotermitidae). FEMS Microbiol Ecol. 2003;44(2):231–42. pmid:19719640.
- 71. Wade WG. The oral microbiome in health and disease. Pharmacol Res. 2013;69(1):137–43. Epub 20121128. pmid:23201354.
- 72. Godon JJ, Moriniere J, Moletta M, Gaillac M, Bru V, Delgenes JP. Rarity associated with specific ecological niches in the bacterial world: the ’Synergistes’ example. Environ Microbiol. 2005;7(2):213–24. pmid:15658988.
- 73. Jumas-Bilak E, Carlier JP, Jean-Pierre H, Citron D, Bernard K, Damay A, et al. Jonquetella anthropi gen. nov., sp. nov., the first member of the candidate phylum ’Synergistetes’ isolated from man. Int J Syst Evol Microbiol. 2007;57(Pt 12):2743–8. pmid:18048718.
- 74. Pradel N, Fardeau ML, Bunk B, Sproer C, Boedeker C, Wolf J, et al. Aminithiophilus ramosus gen. nov., sp. nov., a sulphur-reducing bacterium isolated from a pyrite-forming enrichment culture, and taxonomic revision of the family Synergistaceae. Int J Syst Evol Microbiol. 2023;73(2). pmid:36749697.
- 75. Daniel DS, Ng YK, Chua EL, Arumugam Y, Wong WL, Kumaran JV. Isolation and identification of gastrointestinal microbiota from the short-nosed fruit bat Cynopterus brachyotis brachyotis. Microbiol Res. 2013;168(8):485–96. Epub 20130522. pmid:23706760.
- 76.
Baldani J, Videira SS, dos Santos Teixeira KR, Massena Reis V, Martinez de Oliveira AL, Schwab S, et al. The Family Rhodospirillaceae. In: Rosenberg E, DeLong EF, Lory S, Stackebrandt E, Thompson F, editors. The Prokaryotes: Alphaproteobacteria and Betaproteobacteria2014.
- 77. Ramirez C, Coronado J, Silva A, Romero J. Cetobacterium Is a Major Component of the Microbiome of Giant Amazonian Fish (Arapaima gigas) in Ecuador. Animals (Basel). 2018;8(11). Epub 20181024. pmid:30352962; PubMed Central PMCID: PMC6262583.
- 78. Tsuchiya C, Sakata T, Sugita H. Novel ecological niche of Cetobacterium somerae, an anaerobic bacterium in the intestinal tracts of freshwater fish. Lett Appl Microbiol. 2008;46(1):43–8. Epub 20071016. pmid:17944860.
- 79. Watanabe F, Bito T. Vitamin B(12) sources and microbial interaction. Exp Biol Med (Maywood). 2018;243(2):148–58. Epub 20171207. pmid:29216732; PubMed Central PMCID: PMC5788147.
- 80. Foster G, Ross HM, Naylor RD, Collins MD, Ramos CP, Fernandez Garayzabal F, et al. Cetobacterium ceti gen. nov., sp. nov., a new gram-negative obligate anaerobe from sea mammals. Lett Appl Microbiol. 1995;21(3):202–6. pmid:7576509.
- 81. Finegold SM, Vaisanen ML, Molitoris DR, Tomzynski TJ, Song Y, Liu C, et al. Cetobacterium somerae sp. nov. from human feces and emended description of the genus Cetobacterium. Syst Appl Microbiol. 2003;26(2):177–81. pmid:12866843.
- 82. Humrighouse BW, Emery BD, Kelly AJ, Metcalfe MG, Mbizo J, McQuiston JR. Haematospirillum jordaniae gen. nov., sp. nov., isolated from human blood samples. Antonie Van Leeuwenhoek. 2016;109(4):493–500. Epub 20160208. pmid:26857139.
- 83. Hornok S, Agh N, Takacs N, Kontschan J, Hofmann-Lehmann R. Haematospirillum and insect Wolbachia DNA in avian blood. Antonie Van Leeuwenhoek. 2018;111(3):479–83. Epub 20171023. pmid:29063344.
- 84. Hovan G, Hollinger A. Clinical Isolation and Identification of Haematospirillum jordaniae. Emerg Infect Dis. 2018;24(10):1955–6. pmid:30226180; PubMed Central PMCID: PMC6154156.
- 85. Pal E, Strumbelj I, Kisek TC, Kolenc M, Pirs M, Rus KR, et al. Haematospirillum jordaniae Cellulitis and Bacteremia. Emerg Infect Dis. 2022;28(10):2116–9. pmid:36148990; PubMed Central PMCID: PMC9514349.
- 86. Blachier F, Davila AM, Mimoun S, Benetti PH, Atanasiu C, Andriamihaja M, et al. Luminal sulfide and large intestine mucosa: friend or foe? Amino Acids. 2010;39(2):335–47. Epub 20091218. pmid:20020161.
- 87. Gao R, Liu H, Li Y, Liu H, Zhou Y, Yuan L. Correlation between dominant bacterial community and non-volatile organic compounds during the fermentation of shrimp sauces. Food Science and Human Wellness. 2023;12(1):233–41.
- 88. Gilroy R, Ravi A, Getino M, Pursley I, Horton DL, Alikhan NF, et al. Extensive microbial diversity within the chicken gut microbiome revealed by metagenomics and culture. PeerJ. 2021;9:e10941. Epub 20210406. pmid:33868800; PubMed Central PMCID: PMC8035907.
- 89. Guzman J, Sombolestani AS, Poehlein A, Daniel R, Cleenwerck I, Vandamme P, et al. Entomobacter blattae gen. nov., sp. nov., a new member of the Acetobacteraceae isolated from the gut of the cockroach Gromphadorhina portentosa. Int J Syst Evol Microbiol. 2019;71(3). Epub 20210202. pmid:33528344.
- 90. Haas KN, Blanchard JL. Reclassification of the Clostridium clostridioforme and Clostridium sphenoides clades as Enterocloster gen. nov. and Lacrimispora gen. nov., including reclassification of 15 taxa. Int J Syst Evol Microbiol. 2020;70(1):23–34. pmid:31782700.
- 91. Medina-Pascual MJ, Monzon S, Villalon P, Cuesta I, Gonzalez-Romo F, Valdezate S. Saezia sanguinis gen. nov., sp. nov., a Betaproteobacteria member of order Burkholderiales, isolated from human blood. Int J Syst Evol Microbiol. 2020;70(3):2016–25. pmid:32003711.
- 92. Lobato-Bailon L, Garcia-Ulloa M, Santos A, Guixe D, Camprodon J, Florensa-Rius X, et al. The fecal bacterial microbiome of the Kuhl’s pipistrelle bat (Pipistrellus kuhlii) reflects landscape anthropogenic pressure. Anim Microbiome. 2023;5(1):7. Epub 20230204. pmid:36739423; PubMed Central PMCID: PMC9898988.