Diet Determined by Next Generation Sequencing Reveals Pest Consumption and Opportunistic Foraging by Bats in Macadamia Orchards in South Africa

Recent studies have documented the economically significant impact of bats as predators of agricultural pest insects. We used Next Generation Sequencing (NGS) of the cytochrome oxidase I gene to elucidate the diet of six species of bats based on faecal pellets collected from individuals and roosts in macadamia orchards at Levubu, Limpopo Province, South Africa. For five of these species, we compared the molecular data with published results from microscopic analysis of faecal pellets, culled parts and stomach contents. We provide the first description of the molecular diet of the large African molossid bat, Mops midas. Expectations from skull morphology and a single limited study of stomach contents were that this species should be a beetle-specialist. However, NGS revealed that the diet of M. midas contained a much higher prevalence and diversity of lepidopteran (81 taxa from 17 families) compared to coleopteran (two taxa) prey. While this result is predicted by the allotonic frequency hypothesis for a bat species with low echolocation frequency, it could also be explained by unequal PCR amplification, a constraint of amplicon sequencing. Apart from the above-mentioned species where our sample was probably unbiased (24 pellets from multiple roosts and occasions), sample sizes of the other five species were very low and therefore potentially biased (1-6 pellets). Nevertheless, these samples revealed for each bat species surprisingly many prey taxa spanning several insect orders, indicating that individual bats were capable of consuming a wide diversity of prey during one or two nights of foraging. Contrary to expectations, bats of all foraging groups (clutter, clutteredge and open-air) fed opportunistically on mostly-flightless cockroaches (Order Blattodea). About one third of all faecal pellets tested from five species of bats of all foraging groups contained DNA from the significant macadamia pest species, Nezara viridula (Order Heteroptera), indicating the value of intact bat communities in the biological control of pest stink bugs in macadamia orchards. Contrary to the general expectations of the allotonic frequency hypothesis, all six bat species studied fed predominantly on tympanate versus non-tympanate species of moths (57-75% of lepidopteran prey taxa), even those 'non-allotonic' bat species having intermediate echolocation peak frequencies that encompass the frequency sensitivity of hearing (tympanate) moths.