In vitro fertilization program in white rhinoceros.

In brief: To save endangered rhinoceros species, assisted reproductive technologies are warranted. We here report in vitro blastocyst generation of the Near-Threatened Southern white rhinoceros and, for the first time, also of the technically Extinct Northern white rhinoceros. Abstract: The Anthropocene is marked by a dramatic biodiversity decline, particularly affecting the family Rhinocerotidae. Three of five extant species are listed as Critically Endangered (Sumatran, Javan, black rhinoceros), one as Vulnerable (Indian rhinoceros), and only one white rhino (WR) subspecies, the Southern white rhinoceros (SWR), after more than a century of successful protection is currently classified as Near Threatened by the IUCN, while numbers again are declining. Conversely, in 2008, the SWR's northern counterpart and second WR subspecies, the Northern white rhinoceros (NWR), was declared extinct in the wild. Safeguarding these vanishing keystone species urgently requires new reproductive strategies. We here assess one such strategy, the novel in vitro fertilization program in SWR and - for the first-time NWR - regarding health effects, donor-related, and procedural factors. Over the past 8 years, we performed 65 procedures in 22 white rhinoceros females (20 SWR and 2 NWR) comprising hormonal ovarian stimulation, ovum pick-up (OPU), in vitro oocyte maturation, fertilization, embryo culture, and blastocyst cryopreservation, at an efficiency of 1.0 ± 1.3 blastocysts per OPU, generating 22 NWR, 19 SWR and 10 SWR/NWR hybrid blastocysts for the future generation of live offspring.

Ethical Analysis of the Application of Assisted Reproduction Technologies in Biodiversity Conservation and the Case of White Rhinoceros (Ceratotherium simum) Ovum Pick-Up Procedures.

Originally applied on domestic and lab animals, assisted reproduction technologies (ARTs) have also found application in conservation breeding programs, where they can make the genetic management of populations more efficient, and increase the number of individuals per generation. However, their application in wildlife conservation opens up new ethical scenarios that have not yet been fully explored. This study presents a frame for the ethical analysis of the application of ART procedures in conservation based on the Ethical Matrix (EM), and discusses a specific case study-ovum pick-up (OPU) procedures performed in the current conservation efforts for the northern white rhinoceros (Ceratotherium simum cottoni)-providing a template for the assessment of ART procedures in projects involving other endangered species.

Peste des Petits Ruminants Virus Infection at the Wildlife-Livestock Interface in the Greater Serengeti Ecosystem, 2015-2019.

Peste des petits ruminants (PPR) is a viral disease of goats and sheep that occurs in Africa, the Middle East and Asia with a severe impact on livelihoods and livestock trade. Many wild artiodactyls are susceptible to PPR virus (PPRV) infection, and some outbreaks have threatened endangered wild populations. The role of wild species in PPRV epidemiology is unclear, which is a knowledge gap for the Global Strategy for the Control and Eradication of PPR. These studies aimed to investigate PPRV infection in wild artiodactyls in the Greater Serengeti and Amboseli ecosystems of Kenya and Tanzania. Out of 132 animals purposively sampled in 2015-2016, 19.7% were PPRV seropositive by ID Screen PPR competition enzyme-linked immunosorbent assay (cELISA; IDvet, France) from the following species: African buffalo, wildebeest, topi, kongoni, Grant's gazelle, impala, Thomson's gazelle, warthog and gerenuk, while waterbuck and lesser kudu were seronegative. In 2018-2019, a cross-sectional survey of randomly selected African buffalo and Grant's gazelle herds was conducted. The weighted estimate of PPRV seroprevalence was 12.0% out of 191 African buffalo and 1.1% out of 139 Grant's gazelles. All ocular and nasal swabs and faeces were negative by PPRV real-time reverse transcription-polymerase chain reaction (RT-qPCR). Investigations of a PPR-like disease in sheep and goats confirmed PPRV circulation in the area by rapid detection test and/or RT-qPCR. These results demonstrated serological evidence of PPRV infection in wild artiodactyl species at the wildlife-livestock interface in this ecosystem where PPRV is endemic in domestic small ruminants. Exposure to PPRV could be via spillover from infected small ruminants or from transmission between wild animals, while the relatively low seroprevalence suggests that sustained transmission is unlikely. Further studies of other major wild artiodactyls in this ecosystem are required, such as impala, Thomson's gazelle and wildebeest.

An Ethical Assessment Tool (ETHAS) to Evaluate the Application of Assisted Reproductive Technologies in Mammals' Conservation: The Case of the Northern White Rhinoceros (Ceratotherium simum cottoni).

Assisted reproductive technologies (ARTs) can make a difference in biodiversity conservation. Their application, however, can create risks and raise ethical issues that need addressing. Unfortunately, there is a lack of attention to the topic in the scientific literature and, to our knowledge, there is no tool for the ethical assessment of ARTs in the context of conservation that has been described. This paper reports the first applications of the Ethical Assessment Tool (ETHAS) to trans-rectal ovum pick-up (OPU) and in vitro fertilization (IVF) procedures used in a northern white rhinoceros (Ceratotherium simum cottoni) conservation project. The ETHAS consists of two checklists, the Ethical Evaluation Sheet and the Ethical Risk Assessment, and is specifically customized for each ART procedure. It provides an integrated, multilevel and standardized self-assessment of the procedure under scrutiny, generating an ethical acceptability ranking (totally, partially, not acceptable) and a risk rank (low, medium, high), and, hence, allows for implementing measures to address or manage issues beforehand. The application of the ETHAS to the procedures performed on the northern white rhinoceros was effective in ensuring a high standard of procedures, contributing to the acceptability and improved communication among the project's partners. In turn, the tool itself was also refined through an iterative consultation process between experts and stakeholders.

Molecular survey of Coxiella burnetii in wildlife and ticks at wildlife-livestock interfaces in Kenya.

Coxiella burnetii is the causative agent of Q fever, a zoonotic disease of public health importance. The role of wildlife and their ticks in the epidemiology of C. burnetii in Kenya is unknown. This study analysed the occurrence and prevalence of the pathogen in wildlife and their ticks at two unique wildlife-livestock interfaces of Laikipia and Maasai Mara National Reserve (MMNR) with the aim to determine the potential risk of transmission to livestock and humans. Blood from 79 and 73 animals in Laikipia and MMNR, respectively, and 756 and 95 ixodid ticks in each of the areas, respectively, was analysed. Ticks were pooled before analyses into 137 and 29 samples in Laikipia and MMNR, respectively, of one to eight non-engorged ticks according to species and animal host. Real-time PCR amplifying the repetitive insertion element IS1111a of the transposase gene was used to detect C. burnetii DNA. Although none of the animals and ticks from MMNR tested positive, ticks from Laikipia had an overall pooled prevalence of 2.92% resulting in a maximum-likelihood estimate of prevalence of 0.54%, 95% CI 0.17-1.24. Ticks positive for C. burnetii DNA belonged to the genus Rhipicephalus at a pooled prevalence of 2.96% (maximum-likelihood estimate of prevalence of 0.54%, 95% CI 0.17-1.26). These ticks were Rhipicephalus appendiculatus, R. pulchellus and R. evertsi at pooled prevalence of 3.77, 3.03 and 2.04%, respectively. The presence of C. burnetii in ticks suggests circulation of the pathogen in Laikipia and demonstrates they may play a potential role in the epidemiology of Q fever in this ecosystem. The findings warrant further studies to understand the presence of C. burnetii in domestic animals and their ticks within both study areas.

Molecular surveillance of spotted fever group rickettsioses in wildlife and detection of Rickettsia sibirica in a Topi (Damaliscus lunatus ssp. jimela) in Kenya.

Spotted fever group rickettsioses are a group of tick-borne zoonotic diseases caused by intracellular bacteria of the genus Rickettsia. The diseases are widely reported amongst international travellers returning from most sub-Saharan Africa with fever, yet their importance in local populations largely remains unknown. Although this has started to change and recently there have been increasing reports of the diseases in livestock, ticks and humans in Kenya, they have not been investigated in wildlife. We examined the presence, prevalence and species of Rickettsia present in wildlife in two regions of Kenya with a unique human-wildlife-livestock interface. For this purpose, 79 wild animals in Laikipia County and 73 in Maasai Mara National Reserve were sampled. DNA extracted from blood was tested using the polymerase chain reaction (PCR) to amplify the intergenic spacer rpmE-tRNAfMet and the citrate synthase-encoding gene gltA. Rickettsial DNA was detected in 2 of the 79 (2.5%) animals in Laikipia and 4 of the 73 (5.5%) in Maasai Mara. The PCR-positive amplicons of the gltA gene were sequenced to determine the detected Rickettsia species. This revealed Rickettsia sibirica in a Topi (Damaliscus lunatus ssp. jimela). This is the first report of spotted fever group rickettsioses in wildlife and the first to report R. sibirica in Kenya. The finding demonstrates the potential role of wild animals in the circulation of the diseases.

Knowledge, Attitudes and Practices Towards Spotted Fever Group Rickettsioses and Q Fever in Laikipia and Maasai Mara, Kenya.

Many factors contribute to misdiagnosis and underreporting of infectious zoonotic diseases in most sub-Saharan Africa including limited diagnostic capacity and poor knowledge. We assessed the knowledge, practices and attitudes towards spotted fever group rickettsioses (SFGR) and Q fever amongst local residents in Laikipia and Maasai Mara in Kenya. A semi-structured questionnaire was administered to a total of 101 respondents including 51 pastoralists, 17 human health providers, 28 wildlife sector personnel and 5 veterinarians. The pastoralists expressed no knowledge about SFGR and Q fever. About 26.7% of the wildlife sector personnel in Laikipia expressed some knowledge about SFGR and none in Maasai Mara. None of these respondents had knowledge about Q fever. About 45.5 and 33.3% of the health providers in Laikipia and Maasai Mara respectively expressed knowledge about SFGR and 9.1% in Laikipia expressed good knowledge on Q fever and none in Maasai Mara. The diseases are not considered amongst potential causes of febrile illnesses in most medical facilities except in one facility in Laikipia. Majority of pastoralists practiced at least one predisposing activity for transmission of the diseases including consumption of raw milk, attending to parturition and sharing living accommodations with livestock. Education efforts to update knowledge on medical personnel and One-Health collaborations should be undertaken for more effective mitigation of zoonotic disease threats. The local communities should be sensitized through a multidisciplinary approach to avoid practices that can predispose them to the diseases.