An entry risk assessment of African horse sickness virus into the controlled area of South Africa through the legal movement of equids.

South Africa is endemic for African horse sickness (AHS), an important health and trade-sensitive disease of equids. The country is zoned with movement control measures facilitating an AHS-free controlled area in the south-west. Our objective was to quantitatively establish the risk of entry of AHS virus into the AHS controlled area through the legal movement of horses. Outcomes were subcategorised to evaluate movement pathway, temporal, and spatial differences in risk. A 'no-control' scenario allowed for evaluation of the impact of control measures. Using 2019 movement and AHS case data, and country-wide census data, a stochastic model was developed establishing local municipality level entry risk of AHSV at monthly intervals. These were aggregated to annual probability of entry. Sensitivity analysis evaluated model variables on their impact on the conditional means of the probability of entry. The median monthly probability of entry of AHSV into the controlled area of South Africa ranged from 0.75% (June) to 5.73% (February), with the annual median probability of entry estimated at 20.21% (95% CI: 15.89%-28.89%). The annual risk of AHSV entry compared well with the annual probability of introduction of AHS into the controlled area, which is ~10% based on the last 20 years of outbreak data. Direct non-quarantine movements made up most movements and accounted for most of the risk of entry. Spatial analysis showed that, even though reported case totals were zero throughout 2019 in the Western Cape, horses originating from this province still pose a risk that should not be ignored. Control measures decrease risk by a factor of 2.8 on an annual basis. Not only do the outcomes of this study inform domestic control, they can also be used for scientifically justified trade decision making, since in-country movement control forms a key component of export protocols.

Post-outbreak African horse sickness surveillance: A scenario tree evaluation in South Africa's controlled area.

An African horse sickness (AHS) outbreak occurred in March and April 2016 in the controlled area of South Africa. This extended an existing trade suspension of live equids from South Africa to the European Union. In the post-outbreak period ongoing passive and active surveillance, the latter in the form of monthly sentinel surveillance and a stand-alone freedom from disease survey in March 2017, took place. We describe a stochastic scenario tree analysis of these surveillance components for 24 months, starting July 2016, in three distinct geographic areas of the controlled area. Given that AHS was not detected, the probability of being free from AHS was between 98.3% and 99.8% assuming that, if it were present, it would have a prevalence of at least one infected animal in 1% of herds. This high level of freedom probability had been attained in all three areas within the first 9 months of the 2-year period. The primary driver of surveillance outcomes was the passive surveillance component. Active surveillance components contributed minimally (<0.2%) to the final probability of freedom. Sensitivity analysis showed that the probability of infected horses showing clinical signs was an important parameter influencing the system surveillance sensitivity. The monthly probability of disease introduction needed to be increased to 20% and greater to decrease the overall probability of freedom to below 90%. Current global standards require a 2-year post-incursion period of AHS freedom before re-evaluation of free zone status. Our findings show that the length of this period could be decreased if adequately sensitive surveillance is performed. In order to comply with international standards, active surveillance will remain a component of AHS surveillance in South Africa. Passive surveillance, however, can provide substantial evidence supporting AHS freedom status declarations, and further investment in this surveillance activity would be beneficial.

Establishing post-outbreak freedom from African horse sickness virus in South Africa's surveillance zone.

An African horse sickness (AHS) outbreak occurred in South Africa's AHS controlled area in autumn 2016. A freedom from disease survey was performed to establish the likelihood of ongoing circulation of the associated virus during the same period the following year. A single-stage surveillance strategy was employed with a population-level design prevalence of 1% to establish a survey population sensitivity of 95% (probability that one or more positive horses would be detected if AHS was present at a prevalence greater than or equal to the design prevalence). In March 2017, a total of 262 randomly selected horses from 51 herds were sampled from the 2016 outbreak containment zone. Three within-herd and herd-level design prevalence scenarios were used in evaluating the post-survey probability of freedom. Depending on the underlying design prevalence scenarios, effectively ranging between 0.8% and 6.4%, and the use of informed or uninformed priors, the probability of freedom derived from this surveillance ranged between 73.1% and 99.9% (uninformed prior) and between 96.6% and 100% (informed prior). Based on the results, the authors conclude that it is unlikely that the 2016 AHS virus was still circulating in the autumn of 2017 in the 2016 outbreak containment zone. The ability to perform freedom from disease surveys, and also to include risk-based methods, in the AHS controlled area of South Africa is influenced by the changing underlying population at risk and the high level of vaccination coverage in the horse population. Ongoing census post-outbreak must be undertaken to maintain a valid sampling frame for future surveillance activity. The seasonality of AHS, the restricted AHS vaccination period and the inability to easily differentiate infected from vaccinated animals by laboratory testing impact the ability to perform a freedom from disease survey for AHS in the 12 months following an outbreak in the controlled area.

A field investigation of an African horse sickness outbreak in the controlled area of South Africa in 2016.

An outbreak of African horse sickness (AHS) caused by AHS virus type 1 occurred within the South African AHS surveillance zone during April and May 2016. The index case was detected by a private veterinarian through passive surveillance. There were 21 cases in total, which is relatively low compared to case totals during prior AHS outbreaks in the same region (and of the same AHS virus type) in 2004, 2011 and 2014. The affected proportion of horses on affected properties was 0.07 (95% CI 0.04, 0.11). Weather conditions were conducive to high midge activity immediately prior to the outbreak but midge numbers decreased rapidly with the advent of winter. The outbreak was localized, with 18 of the 21 cases occurring within 8 km of the index property and the three remaining cases on two properties within 21 km of the index property, with direction of spread consistent with wind-borne dispersion of infected midges. Control measures included implementation of a containment zone with movement restrictions on equids. The outbreak was attributed to a reversion to virulence of a live attenuated vaccine used extensively in South Africa. Outbreaks in the AHS control zones have a major detrimental impact on the direct export of horses from South Africa, notably to the European Union.

West Nile Virus Lineage 2 in Horses and Other Animals with Neurologic Disease, South Africa, 2008-2015.

During 2008-2015 in South Africa, we conducted West Nile virus surveillance in 1,407 animals with neurologic disease and identified mostly lineage 2 cases in horses (7.4%, 79/1,069), livestock (1.5%, 2/132), and wildlife (0.5%, 1/206); 35% were fatal. Geographic correlation of horse cases with seropositive veterinarians suggests disease in horses can predict risk in humans.

African Horse Sickness Caused by Genome Reassortment and Reversion to Virulence of Live, Attenuated Vaccine Viruses, South Africa, 2004-2014.

African horse sickness (AHS) is a hemorrhagic viral fever of horses. It is the only equine disease for which the World Organization for Animal Health has introduced specific guidelines for member countries seeking official recognition of disease-free status. Since 1997, South Africa has maintained an AHS controlled area; however, sporadic outbreaks of AHS have occurred in this area. We compared the whole genome sequences of 39 AHS viruses (AHSVs) from field AHS cases to determine the source of 3 such outbreaks. Our analysis confirmed that individual outbreaks were caused by virulent revertants of AHSV type 1 live, attenuated vaccine (LAV) and reassortants with genome segments derived from AHSV types 1, 3, and 4 from a LAV used in South Africa. These findings show that despite effective protection of vaccinated horses, polyvalent LAV may, paradoxically, place susceptible horses at risk for AHS.

Quantitative Risk Assessment for African Horse Sickness in Live Horses Exported from South Africa.

African horse sickness (AHS) is a severe, often fatal, arbovirus infection of horses, transmitted by Culicoides spp. midges. AHS occurs in most of sub-Saharan Africa and is a significant impediment to export of live horses from infected countries, such as South Africa. A stochastic risk model was developed to estimate the probability of exporting an undetected AHS-infected horse through a vector protected pre-export quarantine facility, in accordance with OIE recommendations for trade from an infected country. The model also allows for additional risk management measures, including multiple PCR tests prior to and during pre-export quarantine and optionally during post-arrival quarantine, as well as for comparison of risk associated with exports from a demonstrated low-risk area for AHS and an area where AHS is endemic. If 1 million horses were exported from the low-risk area with no post-arrival quarantine we estimate the median number of infected horses to be 5.4 (95% prediction interval 0.5 to 41). This equates to an annual probability of 0.0016 (95% PI: 0.00015 to 0.012) assuming 300 horses exported per year. An additional PCR test while in vector-protected post-arrival quarantine reduced these probabilities by approximately 12-fold. Probabilities for horses exported from an area where AHS is endemic were approximately 15 to 17 times higher than for horses exported from the low-risk area under comparable scenarios. The probability of undetected AHS infection in horses exported from an infected country can be minimised by appropriate risk management measures. The final choice of risk management measures depends on the level of risk acceptable to the importing country.

Complete Genome Sequences of Four African Horse Sickness Virus Strains from a Commercial Tetravalent Live Attenuated Vaccine.

This is a report of the complete genome sequences of plaque-selected isolates of each of the four virus strains included in a South African commercial tetravalent African horse sickness attenuated live virus vaccine.

Complete Genome Sequences of the Three African Horse Sickness Virus Strains from a Commercial Trivalent Live Attenuated Vaccine.

This is a report of the complete genome sequences of plaque-selected isolates of each of the three virus strains included in a South African commercial trivalent African horse sickness attenuated live virus vaccine.

Development of three triplex real-time reverse transcription PCR assays for the qualitative molecular typing of the nine serotypes of African horse sickness virus.

Blood samples collected as part of routine diagnostic investigations from South African horses with clinical signs suggestive of African horse sickness (AHS) were subjected to analysis with an AHS virus (AHSV) group specific reverse transcription quantitative polymerase chain reaction (AHSV RT-qPCR) assay and virus isolation (VI) with subsequent serotyping by plaque inhibition (PI) assays using AHSV serotype-specific antisera. Blood samples that tested positive by AHSV RT-qPCR were then selected for analysis using AHSV type specific RT-qPCR (AHSV TS RT-qPCR) assays. The TS RT-qPCR assays were evaluated using both historic stocks of the South African reference strains of each of the 9 AHSV serotypes, as well as recently derived stocks of these same viruses. Of the 503 horse blood samples tested, 156 were positive by both AHSV RT-qPCR and VI assays, whereas 135 samples that were VI negative were positive by AHSV RT-qPCR assay. The virus isolates made from the various blood samples included all 9 AHSV serotypes, and there was 100% agreement between the results of conventional serotyping of individual virus isolates by PI assay and AHSV TS RT-qPCR typing results. Results of the current study confirm that the AHSV TS RT-qPCR assays for the identification of individual AHSV serotypes are applicable and practicable and therefore are potentially highly useful and appropriate for virus typing in AHS outbreak situations in endemic or sporadic incursion areas, which can be crucial in determining appropriate and timely vaccination and control strategies.

Diagnostic accuracy of a duplex real-time reverse transcription quantitative PCR assay for detection of African horse sickness virus.

Blood samples collected from 503 suspect cases of African horse sickness (AHS) and another 503 from uninfected, unvaccinated South African horses, as well as 98 samples from horses from an AHS free country, were tested with an AHS virus (AHSV) specific duplex real-time reverse transcription quantitative PCR (RT-qPCR) assay and virus isolation (VI). The diagnostic sensitivity and specificity of this AHSV RT-qPCR assay and VI were estimated using a 2-test 2-population Bayesian latent class model which made no assumptions about the true infection status of the tested animals and allowed for the possibility of conditional dependence (correlation) in test results. Median diagnostic sensitivity and specificity of the AHSV RT-qPCR were 97.8% and 99.9%, respectively. Median diagnostic specificity of virus isolation was >99% whereas the estimated diagnostic sensitivity was 44.2%. The AHSV RT-qPCR assay provides for rapid, high-throughput analysis of samples, and is both analytically and diagnostically sensitive and specific. This assay is potentially highly useful for demonstrating freedom or infection of horses with AHSV, thus it is appropriate that its reproducibility be evaluated in other laboratories as a global standard for detection of AHSV.