First Detection of Influenza D Virus Infection in Cattle and Pigs in the Republic of Korea.

Influenza D virus (IDV) belongs to the Orthomyxoviridae family, which also include the influenza A, B and C virus genera. IDV was first detected and isolated in 2011 in the United States from pigs with respiratory illness. IDV circulates in mammals, including pigs, cattle, camelids, horses and small ruminants. Despite the broad host range, cattle are thought to be the natural reservoir of IDV. This virus plays a role as a causative agent of the bovine respiratory disease complex (BRDC). IDV has been identified in North America, Europe, Asia and Africa. However, there has been no information on the presence of IDV in the Republic of Korea (ROK). In this study, we investigated the presence of viral RNA and seroprevalence to IDV among cattle and pigs in the ROK in 2022. Viral RNA was surveyed by the collection and testing of 999 cattle and 2391 pig nasal swabs and lung tissues using a real-time RT-PCR assay. IDV seroprevalence was investigated by testing 742 cattle and 1627 pig sera using a hemagglutination inhibition (HI) assay. The viral RNA positive rate was 1.4% in cattle, but no viral RNA was detected in pigs. Phylogenetic analysis of the hemagglutinin-esterase-fusion (HEF) gene was further conducted for a selection of samples. All sequences belonged to the D/Yamagata/2019 lineage. The seropositivity rates were 54.7% in cattle and 1.4% in pigs. The geometric mean of the antibody titer (GMT) was 68.3 in cattle and 48.5 in pigs. This is the first report on the detection of viral RNA and antibodies to IDV in the ROK.

Data-driven risk assessment of the incursion of African swine fever virus via pig products brought illegally into South Korea by travelers based on the temporal relationship between outbreaks in China.

Since 2018, Asian countries have been affected by the African swine fever (ASF) virus, with major socioeconomic consequences. Moreover, the number of people traveling in Asian countries has been increasing, leading to an inevitable increase in the risk of ASF spread through livestock products carried by travelers. China and South Korea have close geo-economic ties and numerous international travelers. After the ASF outbreak in China in 2018, many illegally imported pig products (IIPPs) that were confiscated from travelers from China at the port of entry in South Korea tested positive for ASF. The detection of ASF virus (ASFV)-positive IIPPs highlights the need to further assess the risk of incursion by travelers and review the existing prevention strategies. Here, we investigated the temporal relationship between ASF outbreaks in China and the detection of ASFV-positive IIPPs in randomly confiscated samples from all ports of entry, such as flights and ships to South Korea, from 2018 to 2019 using a cross-correlation analysis. Based on the significantly correlated temporal lags between the bivariate time-series data, a risk assessment model, using the Bayesian framework, was built to estimate the distribution of the parameters for the risk assessment model and the monthly probability of ASF being introduced via IIPPs from China to South Korea. ASF outbreaks in China were significantly associated with the detection of ASFV-positive IIPPs in South Korea 5 months later. Hence, the monthly probability of ASFV-infected pig products imported from China via a traveler to South Korea was estimated to be 2.00 × 10-5, corresponding to a 0.98 mean monthly probability of at least one ASF-infected pig product arriving at ports of entry via travelers, from 2018 to 2019. To our knowledge, this study is the first attempt to estimate the risk of ASF introduction via pig products carried by international travelers to all ports from neighboring countries in the Asian region using commonly exchanged observed data. The data presented in this study can be used to refine the intervention strategies to combat the spread of transboundary animal diseases.

Emergence of a novel intergenic region (IGR) IV variant of african swine fever virus genotype II in domestic pigs in Vietnam.

African swine fever virus (ASFV) causes African swine fever (ASF), a deadly disease affecting both domestic pigs and wild boars. ASF has become endemic in Vietnam since its first appearance in early 2019. Our previous molecular surveillance studies revealed that all the ASFV strains circulating in Vietnam belong to p72 genotype II, p54 genotype II, CD2v serogroup 8, and CVR of B602L gene variant type I. However, the genetic analysis based on the tandem repeat sequences located between I73R and I329L genes revealed three different intergenic region (IGR) variants; I, II, and III. In this study, using ASFV field isolates collected from September 24th to December 27th, 2021, we report, for the first time, novel IGR IV variants circulating in the Vietnamese pig population.

Genetic Characterization of African Swine Fever Virus from Pig Farms in South Korea during Outbreaks in 2019-2021.

In South Korea, a total of 21 African swine fever (ASF) infected farms were confirmed during 2019-2021. ASF viruses (ASFVs) were isolated from the blood and spleen samples of the 21 affected farms and their genetic characteristics were analyzed. Phylogenetic analysis indicated that the 21 Korean ASFV strains belonged to p72 genotype II and serogroup 8. All isolates were of the intergenic region (IGR) II variant with 10 tandem repeat sequences between I73R and I329L and the central variable region (CVR) 1 variant of the B602L gene. There were no IGR variations between the A179L and A137R and between the MGF 505 9R and10R nor mutations in the O174L, K145R, MGF 505-5R, CP204L, and Bt/Sj regions. The genes of the 21 ASFV strains were identical to those of Georgia 2007/1 and Chinese and Vietnamese strains (Pig/HLJ/2018, China/2018/AnhuiXCGQ, and ASFV_NgheAn_2019); however, X69R of the J268L region of the 18th isolate (Korea/Pig/Goseong/2021) had three nucleotide (CTA) insertions at the 209th position, which led to the addition of one tyrosine (Y) at the C-terminal. This suggests that there are variations among ASFVs circulating in South Korea and the 18th ASFV-infected farm was due to a variant different from those of the other 20 pig farms.

Comparison of the Virulence of Korean African Swine Fever Isolates from Pig Farms during 2019-2021.

African swine fever (ASF) was first reported in South Korea in September 2019, and as of 31 December 2021, a total of 21 cases in domestic pig farms and 1875 ASFV-infected wild boars have been confirmed in the country. With the continued circulation of ASF in wild boars, and subsequent outbreaks in domestic pigs, concerns were raised about the possible changes in virulence occurring among African swine fever viruses (ASFV) circulating in South Korea. In this study, four Korean ASFV strains isolated from domestic pig farms at different time points between 2019 and 2021 were chosen, and used to experimentally infect domestic pigs by intramuscular inoculation to compare their virulence. All challenged pigs died at 4-9 days post-inoculation, with many showing clinical symptoms of fever, depression, loss of appetite, and recumbency. Gross lesions observed at necropsy included enlargement and hemorrhage of the lymph nodes and hydropericardium. The study showed that all four Korean ASFV isolates caused acute forms of illness, which supports the view that virulence among the circulating ASFV isolates in South Korea remained unchanged and highly virulent during this period.

Detection of African Swine Fever at an Abattoir in South Korea, 2020.

In October 2020, a suspect case of African swine fever (ASF) was detected at an abattoir located in the north-central border region of South Korea. The farm of origin was traced and confirmed positive for ASF. This recurrence was following a period of absence of outbreaks in domestic pigs after the first incursion in 2019, during which a total of 14 domestic pig farms were confirmed between September and October 2019. In 2020, a total of two farms were confirmed, and the molecular characterization of key regions of the genome showed that the two isolates from 2020 were identical with the previous isolates from South Korea in 2019. The continued spread and circulation of ASF in the wild boar population represents an increased risk of spill-over outbreaks in domestic pigs, and, therefore, additional control measures should be implemented for farms in these regions, including a heightened level of surveillance. This was the case for the index farm, which was required to send pigs only to the designated abattoir at which the suspect case was quickly detected. The improvement of biosecurity in pig farms, particularly at the wild boar-domestic pig interface, will be key to the successful control of ASF in the region.

Multiple variants of African swine fever virus circulating in Vietnam.

African swine fever (ASF) is a contagious and deadly viral disease affecting swine of all ages. ASF was first reported in Vietnam in February 2019, and it is now considered endemic in Vietnam. In this study, 122 ASF-positive samples collected from domestic pigs in 28 different provinces of northern, central, and southern Vietnam during outbreaks in 2019-2021 were genetically characterized. The findings confirmed that all ASF virus (ASFV) strains circulating in Vietnam belonged to p72 genotype II, p54 genotype II, CD2v serogroup 8, and CVR gene variant type I. However, further analysis based on the tandem repeat sequences located between I73R and I329L genes revealed that there were three different variants of ASFV, IGR I, II, and III, circulating in the domestic pig population in Vietnam. The IGR II variants were the most prevalent (117/122 strains) and were detected in pigs in all of the provinces tested, followed by IGR III (4/122 strains) and IGR I (1/122 strains). This study confirms for the first time the presence of IGR III variants in Vietnam.

Development and Application of a Multiplex Real-Time Polymerase Chain Reaction Assay for the Simultaneous Detection of Bacterial Aetiologic Agents Associated With Equine Venereal Diseases.

Venereal diseases caused by bacteria are important to the equine industry due to economic losses caused by decline of conception rate in breeding horses. Therefore, identification of infected animals as well as the implementation of appropriate managerial procedures based on accurate diagnosis is critical. In this study, two types of multiplex real-time polymerase chain reaction with high sensitivity and specificity were developed for the simultaneous detection and differentiation of five commonly associated bacterial pathogens of venereal diseases in horses, consisting of Taylorella equigenitalis, Taylorella asinigenitalis, Pseudomonas aeruginosa, Klebsiella pneumoniae and Streptococcus zooepidemicus. The assay was applied to samples collected as part of the surveillance of T.equigenitalis infection in South Korea. Swab samples collected from horses in 2015 were tested. T. equigenitalis and K. pneumoniae was detected in 21 (21.0%) and two (2.0%) samples, respectively. No samples were positive for T. asinigenitalis, P. aeruginosa, and S. zooepidemicus. Application of this assay to an existing surveillance program has allowed for an enhanced surveillance for a wider range of venereal diseases of equine to be implemented in South Korea.

Surveillance of Rift Valley Fever Virus in Mosquito Vectors of the Republic of Korea.

Rift Valley fever (RVF) is an acute mosquito-borne viral zoonotic disease that affects mainly domestic ruminants and humans. RVF virus (RVFV) was first identified in Kenya in 1931 and was reported to be endemic in Africa but has recently spread to the Arabian Peninsula. With increasing climate change and globalization of trade in animals and animal products, there is great concern that the disease will spread worldwide to regions such as Europe, Asia, and the Americas. Although RVFV has not been reported in the Republic of Korea (ROK), the possibility of RVFV introduction is increasing because transmissible mosquito vectors are present and direct flights to Africa were added in 2012. For these reasons, we conducted a surveillance study to detect RVFV in mosquito vectors collected around the airport and harbor from 2012 to 2013. A total of 36,734 mosquitoes were collected and tested by real-time RT-PCR. A total of 1837 mosquito pools were used, and all were confirmed to be negative. This is the first report in the ROK concerning RVFV surveillance in mosquito vectors, and continuous surveillance will be conducted for the early warning of RVFV introduction.

Canine Sialolithiasis: Two Case Reports with Breed, Gender, and Age Distribution of 29 Cases (1964-2010).

This study was conducted to investigate the clinical data of two cases of canine sialolithiasis and to analyze 29 cases identified in the Veterinary Medical Database by year of admission, breed, gender, and age. Medical records from the University of Missouri Veterinary Medical Teaching Hospital were reviewed and two dogs diagnosed with sialolithiasis (calcium oxalate) were identified between 1990 and 2010. The two dogs had cervical or pharyngeal sialocele and were successfully treated by sialolith removal and concurrent sialoadenectomy. Signalments of dogs with sialolithiasis between 1964 and 2010 were collected from the Veterinary Medical Database and evaluated. Several breeds of dogs were represented and the 10 to <15 yr old age group was shown to have significant association with sialolithiasis. Sialolithiasis is a rare veterinary condition. In this study, older dogs were at higher risk. In dogs, concurrent sialocele was common and good outcome could be expected after surgical removal of sialoliths with concurrent sialoadenectomy.

Detection and differentiation of Schmallenberg, Akabane and Aino viruses by one-step multiplex reverse-transcriptase quantitative PCR assay.

BACKGROUND: Schmallenberg virus (SBV), Akabane virus (AKAV) and Aino virus (AINV) are members of the Simbu serogroup within the genus Orthobunyavirus, family Bunyaviridae, which can cause reproductive disorders including abortion, stillbirth and congenital malformation in ruminants. Because, the clinical signs are similar, confirmatory diagnosis requires viral detection to differentiate infection between these three viruses. METHODS: In this study, a one-step multiplex reverse-transcriptase quantitative PCR (one-step mRT-qPCR) was developed for the simultaneous detection and differentiation of SBV, AKAV and AINV. RESULTS: The detection limit of the one-step mRT-qPCR for SBV, AKAV and AINV were 2.4 copies (10 (0.6) TCID 50/ml), 96.2 copies (10 (1.5) TCID 50/ml) and 52.3 copies (10 (1.2) TCID 50/ml), respectively. Various field samples such as bovine serum, bovine whole blood, bovine brain, goat serum and Culicoides were analyzed using the one-step mRT-qPCR and compared with previously published RT-qPCRs. The test results of the field samples were identical for the one-step mRT-qPCR and RT-qPCRs, which showed all samples to be negative for SBV, AKAV and AINV, except for one bovine brain sample (1/123) that was positive for AKAV. CONCLUSION: The one-step mRT-qPCR allows for the simultaneous detection of three viral pathogens (SBV, AKAV and AINV) that cause reproductive failure.

Species Diversity and Seasonal Distribution of Culicoides spp. (Diptera: Ceratopogonidae) in Jeju-do, Republic of Korea.

Biting midges belonging to the genus Culicoides (Diptera: Ceratopogonidae) were collected by Mosquito Magnet(®) and black light traps at 5 sites on Jeju-do, Republic of Korea (Korea), from May-November 2013 to determine species diversity and seasonal distribution. A total of 4,267 specimens were collected, of which 99.9% were female. The most common species was Culicoides tainanus (91.8%), followed by C. lungchiensis (7.2%) and C. punctatus (0.6%), while the remaining 4 species accounted for <0.5% of all Culicoides spp. that were collected. High numbers of C. tainanus were collected in May, followed by decreasing numbers through August, and then increasing numbers through November when surveillance was terminated. Peak numbers of C. lungchiensis were collected during September, with low numbers collected from May-August and October-November. The presence of C. lungchiensis in Korea was confirmed by morphological and molecular analyses.

Serological surveillance studies confirm the Rift Valley fever virus free status in South Korea.

Rift Valley fever is a mosquito-borne zoonotic disease of domestic ruminants. This disease causes abortions in pregnant animals, and it has a high mortality rate in newborn animals. Recently, a Rift Valley fever virus (RVFV) outbreak in the Arabian Peninsula increased its potential spread to new regions worldwide. In non-endemic or disease-free countries, early detection and surveillance are important for preventing the introduction of RVFV. In this study, a serological surveillance was conducted to detect antibodies against RVFV. A total of 2382 serum samples from goats and cattle were randomly collected from nine areas in South Korea from 2011 to 2013. These samples were tested for antibodies against RVFV, using commercial ELISA kits. None of the goats and cattle were positive for antibodies against RVFV. This finding suggests that this disease is not present in South Korea, and furthermore presents the evidence of the RVFV-free status of this country.

Japanese encephalitis virus in culicine mosquitoes (Diptera: culicidae) of the republic of Korea, 2008-2010.

A total of 150,805 culicine female mosquitoes were captured by Mosquito Magnet, black light, and New Jersey light traps, and at resting collections in the Republic of Korea from 2008 to 2010 as part of the U.S. Forces Korea malaria and Japanese surveillance programs. Mosquitoes were identified and culicine mosquitoes placed in pools of up to 30 mosquitoes each, by species and date of collection, and screened for flaviviruses using a reverse transcription-polymerase chain reaction assay. A total of 98/6,845 (1.4%) pools were positive by reverse transcription-polymerase chain reaction for Japanese encephalitis virus (JEV). A total of 92/2,031 (4.5%) pools of Culex tritaeniorhynchus were positive for JEV and accounted for 93.9% (92/98) of all JEV positive pools. A total of 4/804 (0.5%) and 2/175 (1.1%) pools of C. pipiens and C. bitaeniorhynchus, respectively, were positive for JEV. The JEV maximum likelihood estimations (estimated number of viral RNA positive mosquitoes per 1,000) for C. tritaeniorhynchus, C. bitaeniorhynchus, and C. pipiens were 1.71, 1.02, and 0.36 respectively. JEV is a severe health threat for local populations and of significant concern for nonimmune (unvaccinated) U.S. soldiers, civilians, and family members deployed to the Republic of Korea.

First report of Bluetongue virus isolation in the Republic of Korea and analysis of the complete coding sequence of the segment 2 gene.

This study investigated the possible presence of the Bluetongue virus (BTV) in the Republic of Korea (ROK). Cell cultures were used to test blood samples collected from abattoirs throughout the country. Testing identified a single BTV isolate, which was characterized as BTV serotype 1 based on a nucleotide sequence analysis of the segment 2 gene. This report therefore indicates that BTV serotype 1 is present in the ROK. The potential importance of BTV in the ROK has been overlooked because cattle are mostly unaffected by the virus and because sheep, the most severely infected hosts, are uncommon in the ROK. However, as recent BTV serotype 8 outbreaks in Europe have demonstrated, certain BTV strains have the potential to cause severe disease in cattle. Additionally, with climate change continuously expanding the regions in which Culicoides vectors are able to survive, there is an increased need to study BTV in the Far East and ROK. To better prepare for future outbreaks of BTV, a sustained and effective level of surveillance for BTV in livestock will need to be established.

First collection records of Culiseta (Culicella) nipponica in the Republic of Korea.

The 1st confirmed record of Culiseta nipponica for the Republic of Korea was made at Osan Air Base and Tongilchon, Gyeonggi Province, and at the Incheon International Airport, Incheon metropolitan area, using carbon dioxide-baited light and Mosquito Magnet traps. Larval surveys near adult collection sites during June and August 2013 were negative for Culiseta nipponica.

Seasonal abundance of Culicoides (Diptera: Ceratopogonidae) collected by mosquito Magnet® in Northern Gyeonggi-do (Province), Korea.

Biting midges (Culicoides: Ceratopogonidae) were collected by Mosquito Magnet® traps at the Neutral Nations Supervisory Commission (NNSC) camp and Daeseongdong village inside the demilitarized zone (DMZ) and near the military demarcation line (MDL) separating North and South Korea and at Warrior Base (US Army training site) and Tongilchon 3 km south of the DMZ in northern Gyeonggi Province, Republic of Korea (ROK), from May-October 2010-2012, to determine their seasonal distributions. A total of 18,647 Culicoides females (18,399; 98.7%) and males (248; 1.3%) comprising 16 species were collected. Overall, the most commonly collected species was Culicoides nipponensis (42.9%), followed by C. erairai (29.2%), C. punctatus (20.3%), C. arakawae (3.3%), C. pallidulus (1.8%), and C. circumscriptus (1.4%), while the remaining 10 species accounted for only 1.1% of all Culicoides spp. collected. The seasonal distribution of C. nipponensis was bimodal, with high numbers collected during May-June and again during September. C. erairai was more frequently collected during June-July, followed by sharply decreased populations from August-October. C. punctatus was collected in low numbers from May-September with high numbers collected during October. C. erairai was predominantly collected from the NNSC camp (85.1% of all C. erairai collected) located adjacent to the MDL at Panmunjeom in the northernmost part of Gyeonggi-do (Province), while other sites yielded low numbers of specimens.

Control of foot-and-mouth disease during 2010-2011 epidemic, South Korea.

An outbreak of foot-and-mouth disease caused by serotype O virus occurred in cattle and pigs in South Korea during November 2010-April 2011. The highest rates of case and virus detection were observed 44 days after the first case was detected. Detection rates declined rapidly after culling and completion of a national vaccination program.

Molecular detection and genotyping of Japanese encephalitis virus in mosquitoes during a 2010 outbreak in the Republic of Korea.

Japanese encephalitis virus (JEV), a mosquito-borne zoonotic pathogen, is one of the major causes of viral encephalitis. To reduce the impact of Japanese encephalitis among children in the Republic of Korea (ROK), the government established a mandatory vaccination program in 1967. Through the efforts of this program only 0-7 (mean 2.1) cases of Japanese encephalitis were reported annually in the ROK during the period of 1984-2009. However, in 2010 there was an outbreak of 26 confirmed cases of Japanese encephalitis, including 7 deaths. This represented a >12-fold increase in the number of confirmed cases of Japanese encephalitis in the ROK as compared to the mean number reported over the last 26 years and a 3.7-fold increase over the highest annual number of cases during this same period (7 cases). Surveillance of adult mosquitoes was conducted during the 2010 outbreak of Japanese encephalitis in the ROK. A total of 6,328 culicine mosquitoes belonging to 12 species from 5 genera were collected at 6 survey sites from June through October 2010 and assayed by reverse-transcription polymerase chain reaction (RT-PCR) for the presence of JEV. A total of 34/371 pooled samples tested positive for JEV (29/121 Culex tritaeniorhynchus, 4/64 Cx. pipiens, and 1/26 Cx. bitaeniorhynchus) as confirmed by sequencing of the pre-membrane and envelope protein coding genes. The maximum likelihood estimates of JEV positive individuals per 1,000 culicine vectors for Cx. tritaeniorhynchus, Cx. pipiens, and Cx. bitaeniorhynchus were 11.8, 5.6, and 2.8, respectively. Sequences of the JEV pre-membrane and envelope protein coding genes amplified from the culicine mosquitoes by RT-PCR were compared with those of JEV genotypes I-V. Phylogenetic analyses support the detection of a single genotype (I) among samples collected from the ROK in 2010.

Livestock agroterrorism: the deliberate introduction of a highly infectious animal pathogen.

Agroterrorism refers to attacks with any of a variety of biological or chemical agents against commercial crops or livestock populations, either as targets in their own right or as vehicles to attack humans. An agroterrorism incident would generally involve bioterrorism, and potential agents include pathogens such as viruses, bacteria, or fungi. Within the context of agroterrorism, livestock agroterrorism is described as the intentional introduction of an animal-borne infectious disease with the goal of spreading fear, producing economic losses, and/or threatening social stability. Causing human illness or human casualties is another potential goal of livestock agroterrorism. Livestock agroterrorism is considered to be attractive to terrorists because biological agents that affect livestock or poultry are more readily available and more difficult to monitor than are agents that infect humans. In addition, a terrorist attack on animal husbandry may have huge economic consequences with no human casualties. Therefore, a biological attack that targets the animal husbandry sector should be regarded as both a "high-consequence" event and a grave national security risk. This review addresses the use of biological weapons that may be used to target livestock or poultry rather than agricultural inputs or equipment. It first defines livestock agroterrorism. Then, the common priority disease agents that may be used to target livestock or poultry in an agroterrorist attack and that are attractive to terrorists are outlined.