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Previously, we isolated a novel lactic acid bacteria species (Lactobacillus nasalidis) from the fresh forestomach contents of a captive proboscis monkey (Nasalis larvatus) in a Japanese zoo. In this study, we isolated two strains of L. nasalidis from the freeze-dried forestomach contents of a wild proboscis monkey inhabiting a riverine forest in Malaysia. The samples had been stored for more than six years. Phenotypic analysis showed that strains isolated from the wild individual had more diverse sugar utilization and lower salt tolerance than strains previously isolated from the captive counterpart. These phenotypic differences are most likely induced by feeding conditions; wild individuals consume a wide variety of natural food, unlike their zoo-raised counterparts that consume formula feed with sodium sufficiency. Since 16s rRNA sequences of L. nasalidis were detected in the previously created 16S rRNA libraries of wild, provisioned, and captive proboscis monkeys in Malaysia and Japan, L. nasalidis may be an essential bacterium of the foregut microbial community of the proboscis monkey. The currently established method for the isolation of gut bacteria from freeze-dried samples under storage will be applicable to many already-stored precious samples. [ABSTRACT FROM AUTHOR]

Simple Summary: The Tsushima leopard cat is an endangered wild felid that lives solely on Tsushima Island, Nagasaki, Japan. Approximately, only 100 Tsushima leopard cats can be found in the wild, and there are concerns that the population will further reduce due to habitat degradation and traffic accidents. In 1994, Japan's Ministry of the Environment (MOE) developed a conservation and breeding project for the Tsushima leopard cat. The MOE is working with the Japanese Association of Zoo and Aquariums for ex situ conservation of this species. However, considering genetic diversity, it is difficult to conduct captive breeding programs using only natural breeding; hence, assisted reproductive technologies are required. This study aimed to breed Tsushima leopard cats using artificial insemination (AI) by depositing sperms into the oviducts. Ovulation was artificially induced in two females, laparoscopically inseminated with fresh sperms into the oviducts. The pregnancies were monitored via fecal levels of progestogens and radiography. One female had spontaneous delivery of a female kitten 66 days post-AI. This is the first successful case of AI in a Tsushima leopard cat. The Tsushima leopard cat (Prionailurus bengalensis euptilurus) is an endangered wild felid that lives solely on Tsushima Island, Nagasaki, Japan. Japan's Ministry of the Environment is working with the Japanese Association of Zoo and Aquariums for ex situ conservation of this species. However, considering genetic diversity, it is difficult to conduct captive breeding programs by natural breeding alone; hence, assisted reproductive technologies are required. This study aimed to breed Tsushima leopard cats, which otherwise cannot be paired, using laparoscopic oviductal artificial insemination (AI). Female Tsushima leopard cats (female 1, aged 7 years; female 2, aged 6 years) were treated with 200 IU equine chorionic gonadotropin, followed by administration of 1000 IU porcine luteinizing hormone (pLH) after 96 h to induce ovulation. Laparoscopic AI was performed 32 h post-pLH administration. Females 1 and 2 were inseminated in the oviduct with 2.4 × 10⁶ and 3.3 × 10⁶ motile spermatozoa, respectively, collected from two males. Pregnancy was confirmed by radiography 45 and 51 days post-AI in females 1 and 2, respectively; one fetus was found in female 2. Moreover, female 2 had spontaneous delivery of a female kitten 66 days post-AI. This is the first successful case of AI in a Tsushima leopard cat. [ABSTRACT FROM AUTHOR]