Application of the Nagoya Protocol to veterinary pathogens: concerns for the control of foot-and-mouth disease.

The Nagoya Protocol is an international agreement adopted in 2010 (and entered into force in 2014) which governs access to genetic resources and the fair and equitable sharing of benefits from their utilisation. The agreement aims to prevent misappropriation of genetic resources and, through benefit sharing, create incentives for the conservation and sustainable use of biological diversity. While the equitable sharing of the benefits arising from the utilisation of genetic resources is a widely accepted concept, the way in which the provisions of the Nagoya Protocol are currently being implemented through national access and benefit-sharing legislation places significant logistical challenges on the control of transboundary livestock diseases such as foot-and-mouth disease (FMD). Delays to access FMD virus isolates from the field disrupt the production of new FMD vaccines and other tailored tools for research, surveillance and outbreak control. These concerns were raised within the FMD Reference Laboratory Network and were explored at a recent multistakeholder meeting hosted by the European Commission for the Control of FMD. The aim of this paper is to promote wider awareness of the Nagoya Protocol, and to highlight its impacts on the regular exchange and utilisation of biological materials collected from clinical cases which underpin FMD research activities, and work to develop new epidemiologically relevant vaccines and other diagnostic tools to control the disease.

A TaqMan-based RT-qPCR assay for serotyping of Southern African territories (SAT) 2 strains of Foot-and-Mouth disease virus (FMDV) in Southern Africa.

OBJECTIVE: Determining the serotype of circulating virus strains is important in implementing effective vaccination. In this study, Foot-and-Mouth Disease (FMD) Southern African territory 2 (SAT2) specific primers and TaqMan probe were designed towards rapid SAT2 detection and serotyping. The primers were tested by endpoint reverse transcription (RT) polymerase chain reaction (PCR) and quantitative PCR (RT-qPCR) using the vaccine strain SAT2035. The SAT2 serotype-specific RT-qPCR assay was compared with currently used ELISA and VP1 sequencing using Cohen's kappa statistics. RESULTS: The primers yielded amplicons of band size 190 bp during endpoint RT-PCR. When coupled with the probe, the primers reaction efficiency was determined to be 99% with an r2 value of 0.994. The results show that the SAT2 assay has comparable performance to VP1 sequencing (k = 1) and a moderate degree of agreement with ELISA (k = 0.571). The data shows that the newly designed assay could be considered for serotyping of SAT2 strains. However, for this assay to be complete there is a need to design effective SAT1 and SAT3 primers and probes that can be multiplexed to target other serotypes that co-circulate within relevant FMD endemic pools. For future implementation of the assay there is also a need to increase the number of field samples towards validation of the assay.

Heartwater: A situation report of the Southern district of Botswana from 2017 to 2019.

Heartwater is a tick-borne haemoparasitic disease that can limit agro-business expansion in Botswana. It poses a threat to national food security due to low animal production as well as livestock morbidity and mortality. This report gives a snapshot view of heartwater in the Southern district of Botswana. Ixodid ticks parasitizing livestock in four Southern sub-districts of Botswana were collected and identified using morphological and molecular methods. A wide distribution of Amblyomma hebraeum in all four Southern sub-districts was revealed. The annual number of heartwater cases across the Southern district of Botswana was determined from veterinary clinical case reports and confirmed through Giemsa-stained brain smears. A concerning gradual annual increase in heartwater cases was shown in the Moshupa sub-district - a hardveld terrain with rock outcrops where the vector thrives. Goats were affected most (55%) by heartwater followed by sheep (37%) and then cattle (8%). Farmers were interviewed on the management of the heartwater burden within their respective sub-districts and they reported that their animals were affected by heartwater despite 17 out of the 27 farmers interviewed attempting to control vectors through acaricide use. The presented heartwater situation warrants further investigation of the prevalence of heartwater and the effectiveness of existing disease control interventions in the disease-endemic Southern district of Botswana.

A novel multiplex qPCR­HRM assay for the simultaneous detection of four abortive zoonotic agents in cattle, sheep, and goats.

Abortifacient pathogens induce substantial economic losses in the livestock industry worldwide, and many of these pathogens are zoonotic, impacting human health. As Brucella spp., Coxiella burnetii, Leptospira spp., and Listeria monocytogenes cause abortion, rapid differential molecular diagnostic tests are needed to facilitate early and accurate detection of abortion to establish effective control measures. However, the available molecular methods are laborious, time-consuming, or costly. Therefore, we developed and validated a novel multiplex real-time polymerase chain reaction (qPCR) method based on high-resolution melting (HRM) curve analysis to simultaneously detect and differentiate four zoonotic abortifacient agents in cattle, goats, and sheep. Our HRM assay generated four well-separated melting peaks allowing the differentiation between the four zoonotic abortifacients. Out of 216 DNA samples tested, Brucella spp. was detected in 45 samples, Coxiella burnetii in 57 samples, Leptospira spp. in 12 samples, and Listeria monocytogenes in 19 samples, co-infection with Brucella spp. and Coxiella burnetii in 41 samples, and 42 samples were negative. This assay demonstrated good analytical sensitivity, specificity, and reproducibility. This is a valuable rapid, cost-saving, and reliable diagnostic tool for detecting individual and co-infections for zoonotic abortifacient agents in ruminants.

The first detection of a serotype O foot-and-mouth disease virus in Namibia.

This report describes the molecular characterization of a serotype O foot-and-mouth disease virus (FMDV) recovered from a field outbreak in the Zambezi region, Namibia during July 2021. Sequence analysis demonstrates that this FMDV belongs to the O/EA-2 topotype sharing closest nucleotide identity (99.5%) to FMD viruses collected since 2018 in Zambia. This is the first detection of serotype O in Namibia, and together with the cases that have been recently detected in southern Zambia, represent the first time that this serotype has been detected in the Southern African FMD endemic pool since 2000, when a virus of Asian origin (O/ME-SA/PanAsia) caused an outbreak in South Africa. This incursion poses a new threat for the region and the potential onward spread of O/EA-2 will now need to be closely monitored since serotype O vaccines are not widely used in Namibia, nor in neighbouring countries.

A Five-Year Retrospective Study of Foot-and-Mouth Disease Outbreaks in Southern Africa, 2014 to 2018.

Foot-and-mouth disease (FMD) virus (FMDv), like other ribonucleic acid (RNA) genome viruses, has a tendency to mutate rapidly. As such, available vaccines may not confer enough cross-protection against incursion of new lineages and sublineages. This paper is a retrospective study to determine the topotypes/lineages that caused previous FMD outbreaks in 6 southern African countries and the efficacy of the current vaccines to protect cattle against them. A total of 453 bovine epithelial tissue samples from 33 FMD outbreaks that occurred in these countries from 2014 to 2018 were investigated for the presence of FMDv. The genetic diversity of the identified Southern African Type (SAT)-FMD viruses was determined by comparing sequences from outbreaks and historical prototype sequences. Of the 453 samples investigated, 176 were positive for four FMDv serotypes. Out of the 176 FMD positive cases there were 105 SAT2 samples, 32 SAT1 samples, 21 SAT3 samples, and 18 serotype O samples. Phylogenetic analysis grouped the SATs VP1 gene sequences into previously observed topotypes in southern Africa. SAT1 viruses were from topotypes I and III, SAT2 viruses belonged to topotypes I, II, III, and IV, and SAT3 viruses were of topotypes I and II. Vaccine matching studies on the field FMDv isolates produced r 1-values greater than or equal to 0.3 for the three SAT serotypes. This suggests that there is no significant antigenic difference between current SAT FMD vaccine strains and the circulating SAT serotypes. Therefore, the vaccines are still fit-purpose for the control FMD in the region. The study did not identify incursion of any new lineages/topotypes of FMD into the sampled southern African countries.

Foot-and-mouth disease in Southern Ghana: occurrence and molecular characterization of circulating viruses.

Foot-and-mouth disease (FMD) is considered to be endemic in Ghana. However, our knowledge of the local epidemiology of the disease is restricted by a lack of serological information and data for characterized viruses causing field outbreaks. In order to improve our understanding of the prevailing situation, this study was initiated to establish the FMD viruses (FMDV) circulating in the country. During 2016, sera (n = 93) and epithelia/oral swab (n = 20) samples were collected from cattle from four districts in Southern Ghana that experienced FMD outbreaks. Sera were analyzed using the PrioCHECK® FMDV non-structural protein (NSP) ELISA whereas the epithelia/oral swab samples were examined by virus isolation, antigen ELISA, reverse transcription polymerase chain reaction (RT-PCR), and sequencing of VP1 followed by phylogenetic analysis. Assay for antibodies against FMDV NSPs provided evidence of exposure to FMDV in 88.2% (82/93) of the sera tested. Serotypes O and A viruses were detected from clinical samples by RT-PCR and sequencing of VP1. Phylogenetic analysis of VP1 coding sequences revealed that the serotype O viruses belonged to the West Africa (WA) topotype and were most closely related to viruses from Niger and Benin, while the serotype A viruses clustered within genotype IV (G-IV) of the Africa topotype and were most closely related to viruses from Nigeria. This study provides useful information on FMDV serotypes and viral lineages that circulate in Ghana and West Africa that may aid in the formulation of effective FMD control strategies.

Complete Genome Sequences of Three African Foot-and-Mouth Disease Viruses from Clinical Samples Isolated in 2009 and 2010.

The complete genome sequences of three foot-and-mouth disease viruses (one virus of each serotype SAT1, SAT2 and O) were directly sequenced from RNA extracted from clinical bovine samples, demonstrating the feasibility of full-genome sequencing from strong positive samples taken from symptomatic animals.