Microbiome composition and presence of cultivable commensal groups of Southern Tamanduas (Tamandua tetradactyla) varies with captive conditions.

Southern Tamanduas (Tamandua tetradactyla) belong to the specialized placental myrmecophages. There is not much information about their intestinal microbiome. Moreover, due to their food specialization, it is difficult to create an adequate diet under breeding conditions. Therefore, we used 16S rDNA amplicon sequencing to analyze the fecal microbiome of captive Southern Tamanduas from four locations in the Czech Republic and evaluated the impact of the incoming diet and facility conditions on microbiome composition. Together with the microbiome analysis, we also quantified and identified cultivable commensals. The anteater fecal microbiome was dominated by the phyla Bacillota and Bacteroidota, while Pseudomonadota, Spirochaetota, and Actinobacteriota were less abundant. At the taxonomic family level, Lachnospiraceae, Prevotellaceae, Bacteroidaceae, Oscillospiraceae, Erysipelotrichaceae, Spirochaetaceae, Ruminococcaceae, Leuconostocaceae, and Streptococcaceae were mainly represented in the fecal microbiome of animals from all locations. Interestingly, Lactobacillaceae dominated in the location with a zoo-made diet. These animals also had significantly lower diversity of gut microbiome in comparison with animals from other locations fed mainly with a complete commercial diet. Moreover, captive conditions of analyzed anteater included other factors such as the enrichment of the diet with insect-based products, probiotic interventions, the presence of other animals in the exposure, which can potentially affect the composition of the microbiome and cultivable microbes. In total, 63 bacterial species from beneficial commensal to opportunistic pathogen were isolated and identified using MALDI-TOF MS in the set of more than one thousand selected isolates. Half of the detected species were present in the fecal microbiota of most animals, the rest varied across animals and locations.

Species and Strain Variability among Sarcina Isolates from Diverse Mammalian Hosts.

Sarcina spp. has been isolated from the gastrointestinal tracts of diverse mammalian hosts. Their presence is often associated with host health complications, as is evident from many previously published medical case reports. However, only a handful of studies have made proper identification. Most other identifications were solely based on typical Sarcina-like morphology without genotyping. Therefore, the aim of this work was culture detection and the taxonomic classification of Sarcina isolates originating from different mammalian hosts. Sarcina-like colonies were isolated and collected during cultivation analyses of animal fecal samples (n = 197) from primates, dogs, calves of domestic cattle, elephants, and rhinoceroses. The study was carried out on apparently healthy animals kept in zoos or by breeders in the Czech Republic and Slovakia. Selected isolates were identified and compared using 16S rRNA gene sequencing and multi-locus sequence analysis (MLSA; Iles, pheT, pyrG, rplB, rplC, and rpsC). The results indicate the taxonomic variability of Sarcina isolates. S. ventriculi appears to be a common gut microorganism in various captive primates. In contrast, a random occurrence was also recorded in dogs. However, dog isolate N13/4e could represent the next potential novel Sarcina taxonomic unit. Also, a potentially novel Sarcina species was found in elephants, with occurrences in all tested hosts. S. maxima isolates were detected rarely, only in rhinoceroses. Although Sarcina bacteria are often linked to lethal diseases, our results indicate that Sarcina spp. appear to be a common member of the gut microbiota and seem to be an opportunistic pathogen. Further characterization and pathogenic analyses are required.

The transition from the female-like great calls to male calls during ontogeny in southern yellow-cheeked gibbon males (Nomascus gabriellae).

It is well known that gibbons emit a pattern of vocalizations, which is specific for species and sex. A previous study showed, however, that immature southern yellow-cheeked gibbon (Nomascus gabriellae) males produce only female-like great calls from 2.3 to 5.3 years of age in co-singing interactions with their mothers. To date, nothing is known about how the vocal repertoire of a male changes from the female-like call (great call) to the male call (staccato notes and multi-modulation phrase) during vocal ontogeny. The goal of this study was to describe the transition from the female-like great call to the male call and the ontogeny of the male call. We predicted that the transition from the female-like great call to the male-specific call and the development of the male call is a normal part of the aging proces. If this is the case, the following phenomena will occur: (a) female vocalization should no longer be produced with the mature form of the multi-modulation phrase and (b) all stages of the male vocalization should occur gradually as the young male ages. Young males regularly emit both female-like great calls and male-specific calls between the ages of 5.6 to 7.1 years. Once the young males reached 7.1 years of age, they emitted male calls exclusively, and they continued to do so until the end of the observation period (at 8.11 years of age). It was confirmed that the young males emitted only female-like great calls during periods when they produced non-mature forms of a multi-modulation phrase (Fm0,1-none or one frequency modulation in second notes). Furhermore, the decrease in the number of female-like great calls was attributed to the development of the mature form of the multi-modulation phrase (Fm2-two or more frequency modulation in second notes), which developed with age. We also confirmed that the multi-modulation phrase developed gradually, while the development of the staccato notes occurred in leaps. A multi-modulation phrase developed as the initial part of the male-specific call. It was evolved from a simpler to a more complex form as the maximum frequency and age of the young males increased. Staccato notes subsequently developed in certain young males. Possible explanations for such vocal ontogeny in young males are discussed in this work.

The bifidobacterial distribution in the microbiome of captive primates reflects parvorder and feed specialization of the host.

Bifidobacteria, which commonly inhabit the primate gut, are beneficial contributors to host wellbeing. Anatomical differences and natural habitat allow an arrangement of primates into two main parvorders; New World monkeys (NWM) and Old World monkeys (OWM). The number of newly described bifidobacterial species is clearly elevated in NWM. This corresponds to our finding that bifidobacteria were the dominant group of cultivated gut anaerobes in NWM, while their numbers halved in OWM and were often replaced by Clostridiaceae with sarcina morphology. We examined an extended MALDI-TOF MS database as a potential identification tool for rapid screening of bifidobacterial distribution in captive primates. Bifidobacterial isolates of NWM were assigned mainly to species of primate origin, while OWM possessed typically multi-host bifidobacteria. Moreover, bifidobacterial counts reflected the feed specialization of captive primates decreasing from frugivore-insectivores, gummivore-insectivores, frugivore-folivores to frugivore-omnivores. Amplicon sequencing analysis supported this trend with regards to the inverse ratio of Actinobacteria and Firmicutes. In addition, a significantly higher diversity of the bacterial population in OWM was found. The evolution specialization of primates seems to be responsible for Bifidobacterium abundance and species occurrence. Balanced microbiota of captive primates could be supported by optimized prebiotic and probiotic stimulation based on the primate host.

Antibiotic susceptibility screening of primate-associated Clostridium ventriculi.

Clostridium ventriculi (syn. Sarcina ventriculi) is a Gram-positive opportunistic pathogen with sarcina morphology. In the case of gastrointestinal disorders, the treatment is often empirical. Due to the common occurrence in primates and the potential risk of dysbiosis; the antibiotic susceptibility screening of C. ventriculi strains isolated from guenon monkeys and crested gibbons to 58 antibiotics was performed to reduce potentially ineffective antibiotic use in case of disease. Isolates were found to be susceptible to the majority of the tested antibiotics, mainly to (fluoro)quinolones, macrolides, penicillins, and tetracyclines. The susceptibility profiles were similar despite the hosts. Tested strains showed also natural resistance to a few antibiotics on the genus level. Detected in vitro antibiotic efficiency is consistent with documented human treatment cases.

A first report of separation calls in southern yellow-cheeked gibbons (Nomascus gabriellae) in captivity.

The effects of social separation, including vocalization, have been studied for a very long time in non-human primates under laboratory conditions. As part of the long-term research on the vocal behaviour of Nomascus gibbons in zoos, this study provides the first record of calls of the southern yellow-cheeked gibbon (Nomascus gabriellae) in response to involuntary separation. Our study revealed that calls were also emitted by an infant (aged 1 year 8 months), and that the acoustic structure of the infant's calls was similar to that of older individuals' calls. Separation-induced calls seem to have a shorter developmental convergence than vocalizations with a stable pattern (which are specific for species and sex). The acoustic structure of the calls reported here comprised simple syllables, and differed from the sex- and species-specific vocal patterns of this species. Our findings demonstrate a novel paradigm in this genus, and provide evidence of the ability of gibbons to express distress when socially separated.

Five novel bifidobacterial species isolated from faeces of primates in two Czech zoos: Bifidobacterium erythrocebi sp. nov., Bifidobacterium moraviense sp. nov., Bifidobacterium oedipodis sp. nov., Bifidobacterium olomucense sp. nov. and Bifidobacterium panos sp. nov.

Five Bifidobacterium strains, VB23T, VB24T, VB25T, VB26T and VB31T, were isolated from chimpanzee (Pan troglodytes), cotton-top tamarin (Saguinus oedipus), Goeldi's marmoset (Callimico goeldii), moustached tamarin (Saguinus mystax) and patas monkey (Erythrocebus patas), respectively, which were kept in two Czech zoos. These strains were isolated from faecal samples and were Gram-positive, non-motile, non-sporulating, anaerobic and fructose-6-phosphate phosphoketolase-positive. Phylogenetic analyses based on 16S rRNA revealed close relatedness between VB23T and Bifidobacterium angulatum LMG 11039T (96.0 %), VB24T and Bifidobacterium pullorum subsp. pullorum DSM 20433T (96.1 %), VB25T and Bifidobacterium goeldii LMG 30939T (96.5 %), VB26T and Bifidobacterium imperatoris LMG 30297T (98.1 %), and VB31T and B. angulatum LMG 11039T (99.40 %). Internal transcribed spacer profiling revealed that VB23T, VB24T, VB25T, VB26T and VB31T had highest similarity to Bifidobacterium breve LMG 13208T (77.2 %), Bifidobacterium longum subsp. infantis ATCC 15697T (85.8 %), Bifidobacterium biavatii DSM 23969T (76.9 %), B. breve LMG 13208T (81.2 %) and B. angulatum LMG 11039T (88.2 %), respectively. Average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) analyses with their closest neighbours supported the independent phylogenetic positions of the strains with values between 86.3 and 94.3 % for ANI and 25.8 and 54.9 % for dDDH. These genomic and phylogenetic analyses suggested that the evaluated strains were novel Bifidobacterium species named Bifidobacterium erythrocebi sp. nov. (VB31T=DSM 109960T=CCUG 73843T), Bifidobacterium moraviense sp. nov. (VB25T=DSM 109958T=CCUG 73842T), Bifidobacterium oedipodis sp. nov. (VB24T=DSM 109957T=CCUG 73932T), Bifidobacterium olomucense sp. nov. (VB26T=DSM 109959T=CCUG 73845T) and Bifidobacterium panos sp. nov. (VB23T=DSM 109963T=CCUG 73840T).

Fur color change and hormonal development in captive females of northern white-cheeked (Nomascus leucogenys) and buff-cheeked (Nomascus gabriellae) gibbons.

Gibbons of the genus Nomascus exhibit strong sexual dichromatism in fur color. Change of fur color in sub-adult wild Nomascus females is associated with the onset of puberty and the time of their dispersal. The variability in fur change may be influenced by social factors. In this study, we determined whether in captive females of crested gibbons begin reproductive maturity prior to dispersing and with association to their fur color. We collected 287 fecal extracts to analyze pregnandiol -3- glucuronide and 17ß estradiol profiles of 4 sub-adult females (Nomascus leucogenys and Nomascus gabriellae) and 183 samples from their mothers, using enzyme immunoassays. The sub-adult females were monitored from 4 years of age. Their hormone profiles showed the onset of ovulatory cycling between 4.6 and 5.8 years. Based on the information about the estrogen influence to the secondary sex characteristic (fur color of female) the positive link between estrogen concentration and age of the sub-adult females was found. However, the amount of the estrogen can apparently be influenced by the presence of mother. If the mother was presented, the level of estrogen was higher than if the mother was missing. Our findings suggest that the probability of changing to beige fur color by the sub-adult females increased with increased age and if they were without mother. This initial study presents the maternal influence as a possible social factor affecting the fur color change of female offspring.

The traits of the great calls in the juvenile and adolescent gibbon males Nomascus gabriellae.

Knowledge about vocal ontogeny and vocal plasticity during ontogeny in primate species is central to understanding the evolution of human speech. Vocalizations in gibbons (Hominoidea) are very interesting and contain complex species- and sex-specific patterns. However, ontogeny of gibbon songs is little studied. Here, we document regular production and ontogenetic changes of female-specific "great call" in 4 immature (2 juvenile-c.a. 3 years old; and 2 adolescent-c.a. 5 years old) males of southern yellow-cheeked gibbon (N. gabriellae) over nine months. None of the males produced fully developed adult-like "great call" and little ontogenetic changes to "great calls" occurred. "Great calls" of sons were shorter, started higher and ended lower than those of their mothers. Regular production of twitter part of great call likely appears around 4th year as it was observed in adolescent but not in juvenile males.

Production of a female-specific great call in an immature male gibbon, the Nomascus genus.

Production of species- and sex-specific vocalisation is a trait characteristic of all gibbons. However, the mechanisms of gibbon vocalisation ontogeny and the manner of developing sex-specific vocal patterns are not yet known. We recorded vocal behaviour of a juvenile male northern white-cheeked gibbon (Nomascus leucogenys). The male produced female-specific great calls. Out of 163 great-call sequences recorded during six sessions, the juvenile son joined in 157 cases with its own great call. In addition, we observed that the juvenile male improved the structure of great calls during the study by including the twitter part. Our results indicate an interesting pattern of vocal ontogeny in Nomascus and suggest a certain degree of plasticity in the vocal signalling in gibbons.

Preliminary insights into the impact of dietary starch on the ciliate, Neobalantidium coli, in captive chimpanzees.

Infections caused by the intestinal ciliate Neobalantidium coli are asymptomatic in most hosts. In humans and captive African great apes clinical infections occasionally occur, manifested mainly by dysentery; however, factors responsible for development of clinical balantidiasis have not been fully clarified. We studied the effect of dietary starch on the intensities of infection by N. coli in two groups of captive chimpanzees. Adult chimpanzees infected by N. coli from the Hodonín Zoo and from the Brno Zoo, Czech Republic, were fed with a high starch diet (HSD) (average 14.7% of starch) for 14 days, followed by a five-day transition period and subsequently with a period of low starch diet (LoSD) (average 0.1% of starch) for another 14 days. We collected fecal samples during the last seven days of HSD and LoSD and fixed them in 10% formalin. We quantified trophozoites of N. coli using the FLOTAC method. The numbers of N. coli trophozoites were higher during the HSD (mean ± SD: 49.0 ± 134.7) than during the LoSD (3.5 ± 6.8). A generalized linear mixed-effects model revealed significantly lower numbers of the N. coli trophozoites in the feces during the LoSD period in comparison to the HSD period (treatment contrast LoSD vs. HSD: 2.7 ± 0.06 (SE), z = 47.7; p<<0.001). We conclude that our data provide a first indication that starch-rich diet might be responsible for high intensities of infection of N. coli in captive individuals and might predispose them for clinically manifested balantidiasis. We discuss the potential nutritional modifications to host diets that can be implemented in part to control N. coli infections.