Synchronized shift of oral, faecal and urinary microbiotas in bats and natural infection dynamics during seasonal reproduction

Dietrich, Muriel; Kearney, Teresa; Seamark, Ernest C. J.; Paweska, Janusz T.; Markotter, Wanda

In this study, we investigated shifts of the microbiota in saliva, urine and faeces during seasonal bat reproduction, and its association to changes in both bacterial and viral shedding patterns. We used a comparative approach in two cave-dwelling bat species in South Africa: the insectivorous bat Miniopterus natalensis and the frugivorous bat Rousettus aegyptiacus. These species seasonally aggregate in huge numbers in maternity roosts [23, 24] , which make them good candidates for active transmission of infectious agents [12, 13] . Indeed, they harbour several infectious agents [25] [26] [27] , including some that have caused sporadic outbreaks in humans (i.e. Marburg virus) [19] . First, we used high-throughput Illumina sequencing to analyse the diversity and structure of bacterial community composition, and changes over the reproductive season. Then, we investigated the temporal dynamics of bacterial and viral shedding, and tested for any association between the observed changes in microbiota and infection dynamics. We focused on three infectious agents highly prevalent in bat populations, which are specifically excreted in three distinct body habitats: herpesviruses (HVs) in saliva, Leptospira bacteria in urine and adenoviruses (AdVs) in faeces. Although they may represent a risk for human health, transmission of these infectious agents from bats to humans has never been reported to date. Gatkop cave (S 24.61806; E 027.65223), and R. aegyptiacus at Matlapitsi (also known as Mahune cave) (S 24.11483; E 30.12110), 256 km from each other. These caves are maternity colonies where female bats seasonally aggregate to give birth and raise the newborns. Bats were captured using harp-traps placed at the entrance of the cave, and each individual was placed directly in a numbered cotton cloth bag. Bats were processed immediately on site using appropriate biosafety conditions, including Tyvek suits coupled with powered air-purifying respirators. Bats were morphologically identified using taxonomic keys [23, 24] and we recorded the sex, the age class (adults, or juveniles = born during the 2015-2016 reproductive season) as well as the reproductive condition. We classified males as reproductive if they exhibited testicular swelling (scrotal), which is indicative of spermatogenesis. We classified females that were pregnant, post-parturient, lactating or post-lactating as reproductive, and those that were neither pregnant nor lactating by the date on which we first observed evidence of parturition as nonreproductive. Pregnant females were identified by gentle palpation of their abdomen. Recently parturient females were identified on the basis of an enlarged, less constricted and yellow vulva. We identified lactating females with enlarged nipples without fur around them. Post-lactating status was determined on the basis of enlarged dry nipples with recovering fur around them. However, fur loss from around the nipple during lactation is less in M. natalensis than R. aegyptiacus.