AFRICAN SWINE FEVER: EMERGING ISSUE AND UPDATES

AFRICAN SWINE FEVER: EMERGING ISSUE AND UPDATES

Dr Maotoshi Ozukum and Dr Ashish Soni

College of Veterinary Science & A H Mhow Indore 453446

1. INTRODUCTION

African swine fever (ASF) is a highly contagious and severe haemorrhagic disease of pigs that produces a wide range of clinical signs and lesions that can closely resemble those of classical swine fever. African swine fever virus (ASFV) has a case fatality rate of up to 100% in domestic pigs and wild boar. The virus is enzootic in many African countries, where it is maintained in the environment through a sylvatic cycle involving warthogs and Ornithodoros (soft) ticks (MSD Veterinary Manual, 2022).

It is indistinguishable in the field from classical swine fever because both are haemorrhagic diatheses, and it is just as contagious. It is responsible for a highly fatal disease in domesticated pigs. It is the greatest limitation to the development of the pig industry (Constable et al., 2016).

2. ETIOLOGY

African swine fever virus (ASFV) is the causative agent of ASF, the only species belonging to the genus Asfivirus of the family Asfarviridae. It is a large enveloped double-stranded (ds) DNA virus and is the only DNA arbovirus (arthropod-borne) known (Guinat et al., 2016).

The virus particle is organized as a complex multi-layer structure having an internal nucleoprotein core, 70-100 nm in diameter, surrounded by an internal lipid layer and an icosahedral capsid, 170-190 nm in diameter and an outer lipid envelope. The virus replicates in the cytoplasm and shares a similar genome organization with that of Poxvirus and Iridovirus (Das et al., 2021).

3. EPIDEMIOLOGY

3.1. Species Affected

Only pigs are affected; domestic pigs of all ages and breeds are highly susceptible, but the virus can be passed in tissue cultures of rabbits, goats, and embryonated hen eggs. The three African wild species (warthogs, giant forest hogs, and bush pigs) are resistant to infection, but European wild boar are susceptible (Constable et al., 2016).

3.2. Morbidity and Case Fatality

Most often the morbidity is 40% to 85%, and the mortality may be as high as 90% to 100% when a virulent virus is involved but maybe only 20% to 40% in less virulent outbreaks. (Constable et al., 2016).

3.4. Epidemiological Cycles of African Swine Fever

The epidemiology of ASF varies substantially between regions, countries and continents depending on the presence or absence of wild boars and arthropod vectors, and the type of pig production system (Stoain et al., 2019).

ASF epidemiology encompasses four independent epidemiologic cycles

• Sylvatic cycle
• Tick–pig cycle
• Wild-boar habitat cycle
• Domestic cycle

It involves soft ticks, particularly Ornithodoros spp., wild African pigs (mainly warthogs), domestic pigs, pig-derived products such as pork and habitat contaminated by carcasses of infected wild boar (Probst et al., 2019).

3.4.1. Ornithodoros soft ticks as vectors of ASFV

Vector-mediated transmission of ASFV occurs through the bites of some members of soft-bodied ticks in the family Argasidae, particularly the genus Ornithodoros (Costard et al., 2013).

To date, 8 Ornithodoros species have been demonstrated as competent vectors for ASFV (Dixon et al., 2019).

The Ornithodoros porcinus porcinus often referred to as O. moubata porcinus or O. moubata ticks serve as the biological vector and reservoir hosts for ASFV in both domestic and wild pigs in southern and eastern Africa, as well as in Madagascar (Ravaomanana et al., 2010).

Currently, no studies have shown evidence of ASF replication in the hard ticks, Ixodes ricinus and Dermacentor reticulatus in Europe, but the virus can survive for 6-8 weeks in the ticks. This fact makes them potential mechanical vectors of ASFV (Carvalho et al., 2014).

3.4.2. Sylvatic cycle

In endemic regions of Africa, ASFV is maintained in a sylvatic cycle among wild pigs, particularly warthogs and soft tick vectors of the Ornithodoros moubata complex without causing disease in the vertebrate host. The infected ticks transmit the virus to the young warthogs and the healthy ticks get infected by sucking the infected blood (Sánchez et al., 2012).

Warthogs remain asymptomatic carriers of ASFV throughout their whole lives, but they cannot transmit the disease to other representatives of their species either horizontally or vertically, so the survival of the virus in the wild environment is dependent on ticks (Frant et al., 2017).

The ticks are able to retain the ASFV for long periods, transmitting the virus to wart hogs in the next season. Furthermore, the virus is maintained in the tick population through transtadial, transovarial and sexual transmission which allows the virus to persist even in the absence of viraemic hosts (Arias et al., 2018).

3.4.3. Tick-pig cycle

In the tick–pig cycle, the virus is mostly transmitted and maintained among domestic pigs, with the ticks serving as a reservoir allowing the virus to persist locally in the environment without involving the warthogs (Wilkinson, 1984).

3.4.4. Wild-boar habitat cycle

The virus can persist in wild boar carcasses and the surrounding environment for months, retaining the ability to infect other susceptible hosts. This cycle is thus, characterized by both direct transmission between wild boar and indirect transmission via the habitat (Probst et al., 2019).

3.4.5. Domestic cycle

On the other hand, the domestic cycle which was responsible for the vast majority of outbreaks of ASF globally, involves the transmission of the virus when the warthogs come in contact with the domestic pigs. It involves pigs of local breeds with or without tick involvement. Once the disease is introduced in the swine population, it can spread locally through the clothing of pig workers, shoes, equipment, agricultural vehicles, secretions and excretions of pigs, and direct contact between pigs or their meat (Oganesyan et al., 2013). This cycle does not involve the natural reservoirs (Ruiz-Gonzalvo et al., 1996).

3.4.6. Transmission and Contagiosity

The transmission of ASFV occurs by both direct and indirect contact with infected animals and their products, or via the environment and potential vectors. Direct contact between sick and healthy animals is one of the most obvious ways of virus transmission (Guinat et al., 2016).

Domestic pigs can become infected with ASFV via infectious body fluids by nasal, oral, subcutaneous, or ocular route. The pigs, once infected remain as carriers and shed the virus into the environment. Furthermore, the infected carcasses can continue to contribute to virus dissemination as the virus can persist in blood and tissues for prolonged periods (Oganesyan et al., 2013).

On the other hand, swill feeding, a common practice in the traditional pig production systems with free-ranging and backyard pigs globally plays an important role in the ASFV transmission to domestic pigs (Costard et al., 2009b).

The most possible source and a major cause of transmission across ASF-free countries are thought to be the import of ASFV-contaminated pork products. (Cortiñas et al., 2017).

There is no reliable evidence for the transmission of ASFV from sows to foetuses during pregnancy. Sexual transmission in pigs has also not been documented, but ASFV is shed in genital secretions and therefore the Terrestrial Animal Health Code provides guidelines to ensure that semen is free of ASF (OIE, 2019).

4. PATHOGENESIS

ASF is characterized by severe leukopenia, mostly associated with lymphopenia and a general state of immunodeficiency.

Monocytes and macrophages are the main target cells for ASFV. ASFV is a DNA virus, but the replication occurs within the cytoplasm and not in the nucleus. The virus replication induces necrosis in the infected cells and virions are released by budding, and can be observed free in the blood, lymph, and interstitial tissue (Constable et al., 2016). The incubation period of disease ranges from 3-19 days (Gogin et al., 2013).

The clinical signs vary according to the virulence of the ASFV strain, the route of exposure, the dose of the virus and the species of pig infected, normally wild boar are more resistant. The virulence of ASFV strains can be distinguished into highly virulent strains with 90-100% mortality, moderately virulent strains with 70-80% mortality in young and 20-40% mortality in adults, and low virulent strains with 10-30% mortality (Wilkinson, 1984)

5. CLINICAL FINDINGS

The disease occurs in acute, subacute and chronic forms. When it occurs as a new infection (epidemic), it is often acute, but it is subacute to chronic when endemic.

5.1. Acute form

In the acute form of the disease, the animals die in an acute state of shock characterized by disseminated intravascular coagulation with multiple haemorrhages in all tissues. The incubation period after contact exposure varies from 4 to 19 days depending on virus dose and the route of infection, but only 2 to 5 days in experimental infections.

• A high fever of 105°F appears abruptly and persists, without other apparent signs, for about 4 days. The fever then subsides, and the pigs show marked cyanotic blotching of the skin, depression, anorexia, huddling together, disinclination to move, weakness, and incoordination.
• Extreme congestion and discolouration of the hindquarters with difficulty in walking are early and characteristic signs.
• Purple discolouration of the skin may be present on the limbs, snout, abdomen, and ears.
• Abortion may occur in all stages of gestation about 5 to 8 days after the infection commences or after 1 to 2 days of fever.
• Death usually occurs within a day or two after the appearance of obvious signs of illness, and death is often preceded by convulsions (Constable et al., 2016).

5.2. Subacute form

It is characterized by thrombocytopenia, leukopenia, and numerous haemorrhagic lesions.

High fever and varying degrees of depression and lethargy are observed during the phase, but some pigs continue to eat; the case-fatality rate is usually less than 5%; the fever subsides in 2 to 3 weeks; and the pigs return to full feed and grow at a normal rate. Recovered pigs have no lesions suggestive of the disease but may be viraemic for several weeks. These persistently infected pigs would pass routine antemortem inspection at slaughter and potentially infectious offal and carcass trimming could be fed unknowingly to other pigs (Constable et al., 2016).

5.3. Chronic form

Cases are intermittently febrile, become emaciated, and develop soft oedematous swellings over limb joints and under the mandible (Constable et al., 2016).

The chronic form is generally caused by isolates of low virulence or is detected in resistant populations. It is characterized by a large variety of clinical signs which are mainly the result of secondary bacterial complications, the most significant being reproduction and articular alterations. Mortality is low, affecting between 2 and 10% of all the sick animals. (Sánchez et al., 2007).

6. NECROPSY FINDINGS

6.1. Gross Changes at Necropsy

The gross changes observed during necropsy bear a close resemblance to those found in hog cholera. However, in cases of acute African swine fever (ASF), the lesions tend to be more severe. The pathology varies based on the virulence of the virus but typically involves extensive haemorrhages and necrosis of lymphoid tissues in acute cases. In subacute and chronic cases, lesions may be minimal or even absent. The most pronounced lesions are commonly found in the spleen, heart, lymph nodes, and kidneys.

Specific Gross Findings:

1. Swollen and Haemorrhagic Lymph Nodes: Gastrohepatic and renal lymph nodes are often severely affected, to the point that they may resemble the spleen.
2. Kidney Lesions: Subcapsular petechiation of the kidneys is a common finding (Fig.4).
3. Cardiac and Serosal Ecchymoses: Ecchymoses(bruise-like discolourations) are observed on the cardiac surfaces and various serosae (Fig.9).

6.2. Histological Lesions

Histological examination provides more diagnostic insights. The virus causes the destruction of the mononuclear phagocyte system and subsequently infects various cell types, including megakaryocytes, tonsillar crypt cells, renal cells, hepatocytes, and endothelial cells. Postcapillary venules undergo hyalinization and endothelial swelling.

Destruction of monocytes/macrophages is evident in the lymph nodes, spleen, and bone marrow. Extensive destruction of hepatocytes occurs in the liver. Marked karyorrhexis of lymphocytes is observable in both normal lymphoid tissues and the infiltrating cell population within parenchymatous organs.

 

7. DIAGNOSIS

7.1. Clinical pathology

7.1.1. Haematology

As in hog cholera, there is a fall in the total leukocyte count to about 40% to 50% of normal by the fourth day of fever. In particular, there is the emergence of immature cells and atypical lymphocytes in the host blood following ASF infection, but the mechanisms are as yet unknown.

There is a pronounced lymphopenia and an increase in immature neutrophils. In chronic cases there is hypergammaglobulinemia.

Clotting times are increased from about 4 days post-infection. Thrombocytopenia is detectable from days 6 to 9. Serum concentrations of C-reactive proteins, serum amyloid A, and haptoglobin have been measured and all increased significantly in pigs inoculated with either ASF or CSF (Constable et al., 2016).

7.2. Detection of the Virus

Currently, the PCR is considered as the ‘gold standard’ test for early detection of the disease due to its superior sensitivity and specificity. Conventional and real-time PCR methods are used for the detection of the virus genome, and multiple primers have been developed to create a rapid diagnostic tool (Beltran et al., 2017).

The nucleotide and deduced amino acid sequences and phylogenetic analysis of p72, p54 and CVR region of B602L genes have clearly characterized the Mizoram, India strain in genotype II.

For simultaneous and differential detection of other swine pathogens such as CSF virus multiplex PCR techniques have also been developed.

Multiplex PCR refers to the use of polymerase chain reaction (PCR) to amplify multiple DNA sequences simultaneously. This technique involves the use of multiple primer sets within a single PCR mixture to produce amplicons of different sizes, which can be differentiated and visualized using gel electrophoresis or other detection methods. Furthermore, other alternative molecular tests, isothermal assays have been described to be useful in field conditions. This assay targets the p72 gene of ASFV, the 5′ region of CSFV, and the ORF6 gene of PRRSV. The mPCR assay has been evaluated for its ability to detect multiple infections in swine simultaneously, using four pairs of primers to detect these five viruses (Hu et al., 2015).

Other techniques such as direct immunofluorescence (DIF), and the hemadsorption (HA) technique, are used in reference laboratories. DIF and HA should be used with other techniques because they both can produce false negatives. The antigen can be detected by the fluorescent antibody technique in the tonsil and submandibular lymph node within 24 to 48 hours of infection and elsewhere once generalization has occurred. The indirect fluorescence antibody and direct fluorescence tests are commonly carried out on pooled visceral fluid samples (Constable et al., 2016).

7.3. Serological Tests

Appropriate samples for laboratory testing are whole blood, serum and tissues, mainly spleen, lymph nodes, bone marrow, lung, tonsil and kidney.

The ELISA is highly sensitive and specific and can be automated for screening large numbers of sera. It has been developed for a variety of ASF proteins, such as p73 or p30. More than 90% of infected pigs can be detected by the demonstration of specific antibodies against the virus (Constable et al., 2016).

For antigen detection, a haemadsorption (HAD) test, fluorescent antibody test (FAT) and Sandwich ELISA can be used.

Serological detection is crucial for the detection of ASFV infection. Anti-ASF antibodies can be detected with – indirect ELISA and other alternative serological tests such as indirect immunoperoxidase test (IPT), indirect fluorescent antibody test (IFAT), immunoblotting (IBT) or counter immunoelectrophoresis (CIE) (Gaudreault et al., 2020).

ELISA followed by immunoblotting is OIE recommended standard serological test often used for international trade purposes. (OIE, 2019).

7.4. Currently used diagnostics

INgezim ASFV/CSFV CROM Ab 25 tests

INgezim® ASFV-CSFV CROM Ab is an immunochromatographic test which uses proteins of African swine fever virus (ASFV) and Classical swine fever virus (CSFV) to detect specific antibodies to these viruses  (Gold Standard Diagnostics, 2022).

8. DIFFERENTIAL DIAGNOSIS

The disease under consideration can be easily mistaken for hog cholera, necessitating a thorough and meticulous examination for accurate differentiation. Several clinical and laboratory factors play a crucial role in distinguishing the two diseases.

9. TREATMENT

There is no treatment for ASF.

10. PREVENTION AND CONTROL

African swine fever is devastating to the swine industry and currently, there is no commercially available vaccine to control the disease (Gaudreault and Richt, 2019).

• Establishment of control zones
• Establishment of quarantine facilities
• Prohibition of scavenging pig production systems
• Enhanced biosecurity in backyard and small-scale farms
• Control on interstate movement
• Ban/control illegal import of contaminated pork and pork products
• Improve ASF surveillance
• Quick elimination of infected animals and proper disposal
• Disinfection of infected premises
• Investigations on the prevalence of soft ticks

11. VACCINE (Future perspectives/ challenges in vaccine development)

Vaccine development for ASFV is ongoing and challenging due to the range of genetic and antigenic variability as well as the myriad of strategies utilized by the virus to evade the host’s immune response. Further work is essential to develop a vaccine that is both safe and provides a high degree of protection across virulent ASFV strains (Alvarez et al., 2019).

The genome of ASFV is very large, ranging from 170 to 193 kb and encoding 150 to 167 genes with virulence and immune-related functions which is one of the reasons why ASF vaccines are difficult to commerce. Prior studies have noted the importance of ASFV growing stably and rapidly in vitro. Thus, in-depth research on ASF genomics and the function of different genes, including those related to the immune response, is vital to developing an ASF vaccine.

Due to the quick demise of sensitive animals when exposed to virulent strains, previous investigations have been unable to establish a connection between viruses and hosts.

11.1. Current breakthrough in vaccine development

Vietnam is the first country to successfully develop and produce 2 vaccines against ASF. The evaluation of the Navet-ASFVAC vaccine, produced by Navetco National Veterinary Joint Stock Company (Navetco), and AVAC ASF LIVE produced by AVAC is underway (Pig Progress, 2023).

AVAC ASF LIVE vaccine

The AVAC ASF LIVE vaccine is a freeze-dried, attenuated vaccine. The vaccine virus is cultivated on the DMAC cell line, which was developed by AVAC Vietnam., JSC. The vaccine is intended for pigs aged 4 weeks and older, with a recommended single-dose administration. It provides a protective immunity period of no less than 5 months.

Each dose contains at least 10^3,5HAD₅₀ of attenuated African Swine Fever (ASF) virus MGF strain and stabilizers. The vaccine is administered to pigs 4 weeks of age and older for active immunity to prevent ASF. At 2-4 weeks of vaccination, the vaccinated pigs are protected against ASF infection with an immunity duration of at least 5 months (AVAC, 2023)

NAVET-ASFVAC Vaccine

It was developed in the year 2022. ASFVAC vaccine involves the use of an attenuated live-virus vaccine. It involves a two-injection doses of the vaccine. The first dose is given to piglets at 8–10 weeks of age and second dose given after 21-30 days after the first dose.

The Vietnamese Dabaco Group has also announced the successful development of its Dacovac-ASF2 vaccine against African Swine Fever (ASF). The company is the third one to do so. Dabaco Group conducted trials with 200 pigs and found the vaccine efficacy rate to be 80% to 100%, according to Vietnam Agriculture. The vaccine is an attenuated freeze-dried vaccine developed from the ASFv-G-I177L/LVR strain.

12. CONCLUSION

African swine fever (ASF) stands as a formidable threat to global pig husbandry, being a highly contagious and severe viral disease affecting both domestic and wild pigs. Recent reports have indicated outbreaks in Assam and Arunachal Pradesh in India, raising concerns for the entire pig farming sector due to the disease’s elevated morbidity and mortality rates. Currently, there are no specific treatments or protective vaccines available against ASF, making prevention a critical focus.

The virus exhibits a robust transmission capacity through various means, including both animate and inanimate objects, with ticks serving as both biological and mechanical carriers. Given these challenges, it is imperative for governments to take decisive precautions to safeguard pig populations and support vulnerable pig farmers, thereby sustaining the rural economy.

Drawing lessons from the past, particularly the entry and outbreak of Porcine Reproductive and Respiratory Syndrome (PRRS) in India in 2011, can offer valuable insights for effective containment and control strategies against ASF. Implementing stringent measures, such as restricting the movement of animals, handlers, vehicles, and even items like feeds and processed pork products from affected areas, is crucial. This necessitates the enforcement of robust rules and regulations to minimize the risk of further spread and mitigate the economic impact on the pig farming community.

Biosecurity may be the most crucial method to resist the spread of ASF until an effective vaccine is developed.

REFERENCES

Adkin, A., Coburn, H., England, T., Hall, S., Hartnett, E., Marooney, C. and Seaman, M. (2004). Risk assessment for the illegal import of contaminated meat and meat products into Great Britain and the subsequent exposure of GB livestock (IIRA): foot and mouth disease (FMD), classical swine fever (CSF), African swine fever (ASF), swine vesicular disease (SVD). New Haw: Veterinary Laboratories Agency.

Alvarez, J., Bakker, D. and Bezos, J. (2019). Epidemiology and control of notifiable animal diseases. Frontiers in Veterinary Science, 6: 43.

Arias, M., Jurado, C., Gallardo, C., Fernández-Pinero, J. and Sánchez-Vizcaíno, J.M. (2018). Gaps in African swine fever: Analysis and priorities. Transboundary and Emerging Diseases, 65(1): 235‐247.

AVAC (2023). PRODUCT: AVAC ASF LIVE. Retrieved from AVAC: https://www.avac.com.vn/en/products-for-pigs/avac-asf-live/

Bastos, A.D., Penrith, M.L., Macome, F., Pinto, F. and Thomson, G.R. (2004). Co-circulation of two genetically distinct viruses in an outbreak of African swine fever in Mozambique: No evidence for individual coinfection. Veterinary Microbiology, 103(3-4): 169-182.

Bellini, S., Rutili, D. and Guberti, V. (2016). Preventive measures aimed at minimizing the risk of African swine fever virus spread in pig farming systems. Acta Veterinaria Scandinavica, 58(1): 82.

Beltran-Alcrudo, D., Gallardo, M.A.A.C., Kramer, S.A., Penrith, M.L., Kamata, A. and Wiersma, L. (2017). African Swine Fever: Detection and Diagnosis. FAO Animal Production and Health Manual, Rome, Italy, pp 88.

Blome, S., Gabriel, C. and Beer, M. (2014). Modern adjuvants do not enhance the efficacy of an inactivated African swine fever virus vaccine preparation. Vaccine, 32: 3879-3882.

Boshoff, C.I., Bastos, A.D., Gerber, L.J. and Vosloo, W. (2007). Genetic characterisation of African swine fever viruses from outbreaks in southern Africa (1973-1999). Veterinary Microbiology, 121(1-2): 45-55.

Cappai, S., Sanna, G., Loi, F., Coccollone, A., Marrocu, E., Oggiano, A. and Bandino, E. (2018). African swine fever detection on field with antigen rapid kit test. Journal of Animal Science and Research, 2(3): 1-8.

Chapman, D.A., Tcherepanov, V., Upton, C. and Dixon, L.K. (2008). Comparison of the genome sequences of non-pathogenic and pathogenic African swine fever virus isolates. Journal of General Virology, 89(2): 397-408.

Constable, P.D., Hinchcliff, K.W., Done, S.H. and Grünberg, W. (2016). Veterinary Medicine: A Textbook of the Diseases of Cattle, Horses, Sheep, Pigs, and Goats. ELSEVIER.

Cortiñas Abrahantes, J., Gogin, A., Richardson, J. and Gervelmeyer, A. (2017). Epidemiological analyses on African swine fever in the Baltic countries and Poland. European Food Safety Authority Journal, 15(3): e04732.

Costard, S., Wieland, B., De Glanville, W., Jori, F., Rowlands, R., Vosloo, W. and Dixon, L.K. (2009b). African swine fever: how can global spread be prevented? Philosophical Transactions of the Royal Society B: Biological Sciences, 364(1530): 2683-2696.

Costard, S, Mur L, Lubroth J, Sanchez-Vizcaino J, Pfeiffer D. (2013). Epidemiology of African swine fever virus. Virus Research;173(1):191-197.

Das, S., Deka, P., Deka, P., Kalita, K., Ansari, T., Hazarika, R. and Barman, N.N. (2021). African swine fever: Etiology, epidemiology, control strategies and progress toward vaccine development: A comprehensive review. Journal of Entomology and Zoology Studies, 9(1): 919-929.

Davies, K., Goatley, L.C., Guinat, C., Netherton, C.L., Gubbins, S., Dixon, L.K. and Reis, A.L. (2017). Survival of African swine fever virus in excretions from pigs experimentally infected with the Georgia 2007/1 isolate. Transboundary and Emerging Diseases, 64(2): 425-431.

de Carvalho Ferreira, H.C., Zúquete, S.T., Wijnveld, M., Weesendorp, E., Jongejan, F., Stegeman, A. and Loeffen, W.L. (2014). No evidence of African swine fever virus replication in hard ticks. Ticks and Tick-Borne Diseases, 5(5): 582-589.

Department of Animal Husbandry and Dairying (2020). Government of India. Ministry of Fisheries, Animal Husbandry and Dairying online http://dahd.nic.in/sites/default/filess/NAP%20on%20Pig%20.pdf

Estrada-Peña, A., Ayllón, N. and De La Fuente, J. (2012). Impact of climate trends on tick-borne pathogen transmission. Frontiers in Physiology, 3: 64.

Fernández‐Pinero, J., Gallardo, C., Elizalde, M., Robles, A., Gómez, C., Bishop, R. and Arias, M. (2013). Molecular diagnosis of African swine fever by a new real‐time PCR using universal probe library. Transboundary and Emerging Diseases, 60(1): 48-58.

Food and Agricultural Organization (2020). Manual on Livestock Disease Surveillance and Information Systems online http://www.fao.org/3/Y0510E/Y0510E06.htm

Food and Agriculture Organization of the United Nations (2010). Good Practices for Biosecurity in the Pig Sector—Issues and Options in Developing and Transition Countries. Food and Agricultural Organization, Roma, Italy, Paper No.169.

Frant, M., Woźniakowski, G. and Pejsak, Z. (2017). African swine fever (ASF) and ticks. No risk of tick-mediated ASF spread in Poland and Baltic states.  Journal of Veterinary Research, 61(4): 375.

Gallardo, C., Nieto, R., Soler, A., Pelayo, V., Fernández-Pinero, J., Markowska-Daniel, I. and Arias, M. (2015a). Assessment of African swine fever diagnostic techniques as a response to the epidemic outbreaks in Eastern European Union countries: How to improve surveillance and control programs. Journal of Clinical Microbiology, 53(8): 2555-2565.

Gaudreault, N.N. and Richt, J.A. (2019). Subunit vaccine approaches for African swine fever virus. Vaccines, 7(2): 56.

Gaudreault, N.N., Madden, D.W., Wilson, W.C., Trujillo, J.D. and Richt, J.A. (2020). African swine fever virus: an emerging DNA arbovirus. Frontiers in Veterinary Science, 7: 215.

Gervasi, V., Marcon, A., Bellini, S. and Guberti, V. (2019). Evaluation of the efficiency of active and passive surveillance in the detection of African swine fever in wild boar. Veterinary Sciences, 7(1): 5.

Gogin, A., Gerasimov, V., Malogolovkin, A. and Kolbasov, D. (2013). African swine fever in the North Caucasus region and the Russian Federation in years 2007–2012. Virus Research, 173(1): 198-203.

Gold Standard Diagnostics (2022). INgezim ASFV/CSFV CROM Ab 25 tests online https://www.goldstandarddiagnostics.com/ingezim-asfv-csfv-crom-ab-25 tests.html#: ~:text=INgezim%C2%AE%20ASFV%2DCSFV%20CROM, specific%20antibodies%20to%20these%20viruses.

Goswami, P. and Borkataki, S. (2017). Pen Side Diagnosis of Infectious disease-A Current Status. International Journal of Advanced Research and Review, 2: 44-55.

Guberti V., Khomenko S., Masiulis M. and Kerba S. (2018). Handbook on African Swine Fever in Wild Boar and Biosecurity during Hunting, Global Framework for the Progressive Control of Transboundary Animal Diseases, Rome, Italy, pp 111.

Guinat, C., Gogin, A., Blome, S., Keil, G., Pollin, R., Pfeiffer, D.U. and Dixon, L. (2016). Transmission routes of African swine fever virus to domestic pigs: current knowledge and future research directions. Veterinary Record, 178(11): 262-267.

Hu, L., Lin, X.Y., Yang, Z.X., Yao, X.P., Li, G.L., Peng, S.Z. and Wang, Y. (2015). A multiplex PCR for simultaneous detection of classical swine fever virus, African swine fever virus, highly pathogenic porcine reproductive and respiratory syndrome virus, porcine reproductive and respiratory syndrome virus and pseudorabies in swines. Polish Journal of Veterinary Sciences, 18(4): 715-723

Manzano-Román, R., Díaz-Martín, V., de la Fuente, J. and Pérez-Sánchez, R. (2012). Soft ticks as pathogen vectors: distribution, surveillance and control. Parasitology, 7: 125-162.

Mebus, C., Arias, M., Pineda, J.M., Tapiador, J., House, C. and Sanchez-Vizcaino, J.M. (1997). Survival of several porcine viruses in different Spanish dry-cured meat products. Food Chemistry, 59(4): 555-559.

MSD Veterinary Manual (2022). African Swine Fever onhttps://www.msdvetmanual.com/generalized-conditions/african-swine-fever/african-swine-fever

Nix, R.J., Gallardo, C., Hutchings, G., Blanco, E. and Dixon, L.K. (2006). Molecular epidemiology of African swine fever virus studied by analysis of four variable genome regions. Archives of Virology, 151: 2475-2494.

O’Donnell, V., Holinka, L.G., Krug, P.W., Gladue, D.P., Carlson, J., Sanford, B. and Borca, M. V. (2015). African swine fever virus Georgia 2007 with a deletion of virulence-associated gene 9GL (B119L), when administered at low doses, leads to virus attenuation in swine and induces an effective protection against homologous challenge. Journal of Virology, 89(16): 8556-8566.

Office International des Epizooties (2019). African Swine Fever (Infection with African Swine Fever Virus). Terrestrial Manual, 1-3.

Office International des Epizooties (2022). Killing of Animals for Disease Control Purposes online: https://www.oie.int/fileadmin/Home/eng/Health_standards/tahc/current/chapitre_aw_killing.pdf

Oganesyan, A.S., Petrova, O.N., Korennoy, F.I., Bardina, N.S., Gogin, A. E. and Dudnikov, S.A. (2013). African swine fever in the Russian Federation: spatio-temporal analysis and epidemiological overview. Virus Research, 173(1): 204-211.

Penrith, M.L. and Vosloo, W. (2009). Review of African swine fever: transmission, spread and control. Journal of the South African Veterinary Association, 80(2): 58-62.

Petrov, A., Forth, J. H., Zani, L., Beer, M. and Blome, S. (2018). No evidence for long‐term carrier status of pigs after African swine fever virus infection. Transboundary and Emerging Diseases, 65(5): 1318-1328.

Pig Progress (2023). Dabaco Group succeeds in developing ASF vaccine online https://www.pigprogress.net/health-nutrition/health/vietnamese-dabaco-group-succeeds-in-developing-asf-vaccine/

Plowright, W. and Parker, J. (1968). The stability of African swine fever virus with particular reference to heat and pH inactivation. Archiv fur die Gesamte Virusforschung, 21: 383-402.

Probst, C., Gethmann, J., Amler, S., Globig, A., Knoll, B. and Conraths, F.J. (2019). The potential role of scavengers in spreading African swine fever among wild boar. Scientific Reports, 9(1): 11450.

Rajkhowa, T.K., Kiran, J., Hauhnar, L., Zodinpui, D., Paul, A. and Sagolsem, S. (2022). Molecular detection and characterization of African swine fever virus from field outbreaks in domestic pigs, Mizoram, India. Transboundary and Emerging Diseases, 69(4): e1028-e1036.

Ravaomanana, J., Michaud, V., Jori, F., Andriatsimahavandy, A., Roger, F., Albina, E. and Vial, L. (2010). First detection of African Swine Fever Virus in Ornithodoros porcinus in Madagascar and new insights into tick distribution and taxonomy. Parasites and Vectors, 3(1): 1-9.

Ruiz-Gonzalvo, F., Rodriguez, F. and Escribano, J.M. (1996). Functional and immunological properties of the baculovirus-expressed hemagglutinin of African swine fever virus. Virology, 218(1): 285-289.

Sanchez-Vizcaino, J.M., Mur, L.  and Arias, M. (2007). African Swine Fever online http://apps.sanidadanimal.info/cursos/asf/caps/cap1.html

Sanchez-Vizcaino, J.M. (2010). Compendium of Technical Items Presented to the OIE World Assembly of Delegates and to OIE Regional Commissions. World Organization for Animal Health, 129-168.

Sanchez-Vizcaino, J.M., Mur, L. and Arias, M. (2012). African swine fever. In: Straw B., D’Allaire S., Mengeling W. and Taylor D. (ed.). Diseases of Swine, 10th Edn., Lowa State University, USA, pp 396-404.

Times Now (2022). African swine fever attacks MP killing 2200 pigs in 13 days online https://www.timesnownews.com/mirror-now/in-focus/african-swine-fever-attacks-mp-killing-2200-pigs-in-13-days-article-93744272

Wikipedia (2020). Swine Diseases: African swine fever virus online https://en.wikipedia.org/wiki/African_swine_fever_virus#cite_note-71

Wilkinson, P.J. (1984). The persistence of African swine fever in Africa and the Mediterranean. Preventive Veterinary Medicine, 2(1-4): 71-82.