Prevalence and Pathology of suspected cases of SRLV infections in Farmed goats

Introduction

Lentiviruses, belonging to the Retroviridae family, are enveloped, single-stranded RNA viruses with a global distribution (Peterhans et al., 2004) [20] . They infect a variety of hosts, including goats, horses, sheep, apes, and cats (Gufler et al., 2007) [9] . Among them, Small Ruminant Lentiviruses (SRLVs) comprise a group of retroviruses that cause chronic, multisystemic, and progressively debilitating lifelong infections in sheep and goats. The major diseases associated with SRLVs include Caprine Arthritis Encephalitis Virus (CAEV), Maedi-Visna Virus (MVV), and Ovine Progressive Pneumonia Virus (OPPV). Clinically, Maedi is characterized by progressive interstitial pneumonia, while Visna presents as meningioleucoencephalitis. CAE primarily causes polysynovitis/arthritis in adult goats and progressive paresis or leucoencephalomyelitis in kids. Pathological and epidemiological studies can be conducted to establish correlations with lentiviral infections in goats. These infections are known to cause immunodeficiency and chronic inflammatory changes, and are unique in their ability to infect both dividing and nondividing cells (Blacklaws, 2012; Zinc et al., 1987) [1,29] . SRLVs infect circulating monocytes and replicate extensively in tissue macrophages, leading to chronic inflammation in organs such as the lungs, mammary glands, central nervous system, and joints. Consequently, histopathological evaluation plays a critical role in the diagnosis of SRLV-associated diseases. In India, the overall seropositivity of SRLVs ranges from 9.53% to 19.58% (Dar, 2017; Mishra, 2018; Kumar, 2019) [3,16,11] , while PCR positivity is reported between 6.84% and 10.41% (Dar, 2017; Mishra, 2018; Kumar, 2019) [3, 17 11] . Also, seropositivity toward CAEV found with prevalence rates of 4.5% (Didugu et al., 2016) [4] , 3.33% (Waseem et al., 2015) [28] , and 6.96% (Singh et al., 2018) [24] . Currently, there are no known treatments or commercially available vaccines for CAEV. Therefore, controlling the spread and reducing the prevalence of SRLVs remains one of the most effective strategies for disease management. PCR is highly specific for the detection of CAEV proviral DNA in infected animals, with no cross-reactivity observed with other ruminant retroviruses (Clavijo and Thorsen, 1996) [2] . Many underreporting cases due to their subclinical nature calls for the

Materials and Methods

The study was conducted at the Department of Veterinary 1. DNA was extracted using the QIAGEN kit, and PCR products were visualized via 2% agarose gel electrophoresis under UV illumination. Cytological analysis of synovial fluid was performed for differential cell counts.

Results and Discussion

Prevalence of SRLV infection in goats Overall Prevalence

The overall prevalence of suspected Small Ruminant Lentivirus (SRLV) infection in goats across Mumbai was 3.14% (110/3500). Among these, chronic respiratory distress was the most commonly observed clinical sign, present in 78.18% (86/110) of affected goats, followed by mastitis (4.55%; 5/110), arthritis (2.73%; 3/110), and mixed-type infections (14.55%; 16/110) involving more than one clinical manifestation. The respiratory signs were further categorized into chronic cough, progressive weight loss, nasal discharge, and dyspnoea. Details of clinical signs showed by goats suspected of SRLV infection are summarized in Table 2.

During the study period, 41 out of 3500 goats (1.17%) died.

Postmortem examination revealed lung lesions as the most frequent pathological finding, observed in 31 goats (0.88%).

Combined lesions involving the lung and udder were found in 3 goats (0.08%), while 2 goats (0.05%) showed concurrent lung, joint, and udder lesions. Additionally, 1 goat (0.02%) exhibited joint lesions alone, and 4 goats (0.11%) had udder lesions without other organ involvement.

Hematobiochemical Examination

Hematobiochemical findings revealed no alterations in the blood samples of suspected cases when compared with normal healthy goats. Mild rise in neutrophils and low lymphocyte count was observed in certain cases. The variation in WBC pattern suggested changes in immunity due to infections. Difference in leukocytes average and patterns, indicates growth-related physiological changes, and may also reflect some changes in immunity as a consequence of viral infection (Lipecka et al., 2010) [14] . In the present study, results of biochemical examination are aligned with the study conducted by Garba et al., (2019) [7] , who observed a significant increase in the levels of ALT, total protein, and globulin among the mastitis-positive goats. [12] , as miscellaneous lesions showed the highest frequency while cyst lesions had the lowest prevalence.

Gross Examination

Gross examination of lung tissues revealed abnormalities in 35 out of 41 samples (85.36%), based on visual inspection, palpation, and assessment of size, location, texture, consistency, and color. Lung lesions were categorized into bronchopneumonia, interstitial pneumonia, cysts, nodules, consolidation, ovine pulmonary adenocarcinoma (OPA), and miscellaneous lesions (including emphysema, atelectasis, congestion, and haemorrhages). Among the pulmonary lesions, miscellaneous lesions were the most prevalent, observed in 17.07% of cases, whereas cystic lesions were the least common, identified in only two lungs (4.8%). These findings are consistent with observations made by Lee et al., (2017) [12] , who reported a similar classification pattern, with miscellaneous lesions being the most frequent and cysts the least prevalent. Affected joints were found to be swollen due to excessive synovial fluid or purulent material, indicative of septic arthritis likely caused by bacterial infection. Additionally, 7 of the 41 diseased goats (17.07%) exhibited signs of mastitis.

Histopathological Evaluation

Histopathological evaluation of 35 abnormal lung tissues revealed bronchopneumonia as the most common lesion (17/35 cases), followed by miscellaneous lesions (10/35), interstitial pneumonia (6/35), and ovine pulmonary adenocarcinoma (OPA; 2/35). These findings are consistent with Javed (2017) and Singh et al., (2017), who also recorded bronchopneumonia as the predominant pathology. Histopathological lesions in lung tissues of goats suspected for SRLV infection are detailed in Table 3. Fibrinous bronchopneumonia was characterized by inflammatory exudates, mainly neutrophils and macrophages within bronchioles and adjacent alveolar spaces. Suppurative bronchopneumonia showed similar exudation along with pulmonary oedema, indicative of acute inflammation. These features align with reports by Singh et al., (2017) [23] , Mahile et al., (2022) [15] , and Kokate (2022) [10] . Interstitial pneumonia exhibited thickened inter-alveolar and interlobular septa (Fig. 2), congestion, oedema, neutrophilic infiltration, and capillary engorgement which are consistent with the findings of El-Mashad et al., (2020) [5] and Valecha (2020). [27] Additional findings included mononuclear cell infiltration (Fig. 1), fibroblast proliferation with collagen deposition, hyperplasia of type II pneumocytes, bronchusassociated lymphoid tissue (BALT), and smooth muscle proliferation. Similar observations were reported by Georgsson et al., (1971), Dousti et al., (2020), Mahile et al., (2022) [15] , and Kokate (2022) [10] . In contrast, Madhav et al., (2015) noted lymphoid nodules and multinucleated giant cells. Other lung lesions included congestion (7.31%), congestion with oedema (4.8%), congestion with haemorrhage (2.43%), pulmonary oedema (7.31%), and emphysema with atelectasis (2.43%), aligning with Valecha (2022) [27] . Histopathological examination of joints showed oedematous and congested synovial membranes, hypertrophied or necrotic villi, and mild inflammatory infiltration, indicative of synovitis. Among seven mammary gland samples examined, four showed atrophy and fibrosis, two had mastitis (one with neutrophilic and one with lymphocytic infiltration), and one appeared normal.

Cytological Examination

Cytological analysis was performed on synovial fluid from 11 goats with arthritis outlined in Table 4. In 4 cases, the fluid appeared clear and colourless with minimal neutrophil presence, suggestive of myositis or periarticular trauma. In 3 goats, the synovial fluid was pale yellow and watery, a feature consistent with chronic hemorrhage and bilirubin pigment formation, as reported by Waseem et al., (2015) [28] and Moraes et al., (2010) [18] . The largest volume (6 mL) was collected from a hock joint and was serosanguineous with low viscosity, aligning with findings from Singh et al., (1997) [22] and Tyagi and Singh (1993) [26] , who noted increased fluid volume in infectious arthritis. Microscopic examination revealed a predominance of neutrophils (91%) and mononuclear cells (9%). Differential cell counts showed that 7 out of 11 samples had elevated neutrophil levels, while the remaining exhibited mononuclear cell predominance.

Molecular detection by PCR

Molecular detection using PCR revealed that none of the samples tested positive for Maedi-Visna virus (MVV) or Caprine Arthritis Encephalitis Virus (CAEV). This negative result is most likely due to a low concentration of viral genetic material, which may have limited the PCR's ability to detect CAEV. As noted by Ramirez et al., (2009), the sensitivity of the primers and the selection of the target region within the viral genome can significantly influence detection outcomes. The high genetic variability of small ruminant lentiviruses (SRLVs) further complicates PCRbased diagnosis, reducing the assay's effectiveness, as highlighted by Peterhans et al., (2004) [20] . In the present study, Ovine Pulmonary Adenocarcinoma (OPA) showed a prevalence rate of 1.41%, which aligns with findings reported by Sonawane et al., (2016) [25] . PCR analysis of blood and tissue samples revealed no positive results in blood; however, out of 31 lung tissue samples, 2 tested positive for the U3 LTR gene (Fig. 3), indicating a lung tissue prevalence of 6.45%. These findings confirm the presence of Jaagsiekte sheep retrovirus (JSRV) in goat populations in and around the Mumbai region. Furthermore, Mycoplasma was detected with an overall prevalence of 9.92% through PCR in the same geographic area. Results of PCR amplified products of Mycoplasma are featured in Fig. 4. These results are consistent with the findings of Lin et al., (2022) [13] , who reported a Mycoplasma prevalence ranging from 8.20% to 24.59% in dairy goats between 2017 and 2020.

Conclusion

Clinical signs indicative of small ruminant lentivirus (SRLV) infection was detected in 3.14% of goats, accompanied by a mortality rate of 1.17%. Hematobiochemical evaluations of suspected cases were inconclusive. In contrast, gross and histopathological assessments of lung tissues revealed a 9.75% prevalence of maedi-like interstitial pneumonia. Molecular analysis of SRLV-negative samples detected Jaagsiekte sheep retrovirus (JSRV) in 1.41% and Mycoplasma species in 9.92% of cases, underscoring the necessity for differential diagnosis. These results emphasize the critical role of combining histopathology with molecular techniques for precise identification of respiratory diseases in goats.

References

1. Blacklaws BA. Small ruminant lentiviruses: immunopathogenesis of visna-maedi and caprine arthritis and encephalitis virus. Comparative Immunology Microbiology and Infectious Diseases. 2012;35(3):259-269. doi:10.1016/j.cimid.2011.12.003
2. Clavijo A, Thorsen J. Application of polymerase chain reaction for the diagnosis of caprine arthritisencephalitis. Small Ruminant Research. 1996;22(1):6977.
3. Dar JA. Monitoring and pathology of viral pneumonias with special reference to retroviral infections in sheep and goats. ICAR-IVRI Deemed University, Izatnagar, Bareilly, India; 2017.
4. Didugu H, Sagi S, Reddy CEN, Kishore KN, Reddy MV, Vishnu PG. First report of Maedi-Visna and caprine arthritis-encephalitis viruses in Krishna district, Andhra Pradesh, India. Journal of Animal Research. 2016;6(1):177-179.
5. El-Mashad ABI, Moustafa SA, Amin A, Samy EM. Pathological studies on lung affections in sheep and goat at Kalubia Governorate. Benha Veterinary Medical Journal. 2020;38(1):17-23.
6. Emikpe BO, Jarikre TA, Eyarefe OD. Retrospective study of disease incidence and type of pneumonia in Nigerian small ruminants in Ibadan, Nigeria. African Journal of Biomedical Research. 2013;16(2):107-113.
7. Garba B, Habibullah SA, Saidu B, Suleiman N. Effect of mastitis on some hematological and biochemical parameters of Red Sokoto goats. Veterinary World. 2019;12:572-577.
8. Ghanem YM, El-Khodery SA, Saad AA, Elragaby SA, Abdelkader AH, Heybe A. Prevalence and risk factors of caprine arthritis encephalitis virus infection in Northern Somalia. Small Ruminant Research. 2009;85(2-3):142-148.
9. Gufler H, Baumgartner W. Overview of herd and CAEV status in dwarf goats in South Tyrol, Italy. Veterinary Quarterly. 2007;29(2):68-70.
10. Kokate. Pathoepidemiological studies on pulmonary adenomatosis in slaughtered sheep. Maharashtra Animal and Fishery Sciences University, Nagpur; 2022.
11. Kumar K. Immunopathological study of retroviral diseases in sheep and goats with their etiological characterization. MVSc Thesis. ICAR-IVRI Deemed University, Izatnagar, India; 2019.
12. Lee AM, Wolfe A, Cassidy JP, McV Messam LL, Moriarty JP, O'Neill R, et al. First confirmation by PCR of Jaagsiekte sheep retrovirus in Ireland and prevalence of ovine pulmonary adenocarcinoma in adult sheep at slaughter. Irish Veterinary Journal. 2017;70:1-12.
13. Lin H, Toung OP, Weng HX, Hsu P, Chang MH, Lai J. The prevalence and species of Mycoplasma identified from dairy goats in Taiwan. Research in Veterinary Science. 2022;151:184-188.
14. Lipecka CZ, Olech M, Gruszeek TM, Junkuszew A, Kuzmak J. Haematological and biochemical parameters blood of lambs born to maedi-visna virus-infected and uninfected ewes. Bulletin of the Veterinary Institute in Pulawy. 2010;54:135-139.
15. Mahile V, Gadhave PD, Kadam DP, Meshram PV, Ingole SD, Pawalkar DA, et al. Prevalence and histopathological investigation of pneumonia in slaughtered goats. The Pharma Innovation Journal. 2022;SP-11(7):2030-2033.
16. Mishra S. Pathology of ovine pulmonary adenomatosis with reference to tumour biomarkers and molecular characterization of JSRV in sheep and goats. ICARIVRI Deemed University, Izatnagar, India; 2018.
17. Mishra S, Kumar P, George N, Singh R, Singh V, Singh R. Survey of lung affections in sheep and goats: a slaughterhouse study. Journal of Entomology and Zoology Studies. 2018;6(4):118-120.
18. Moraes MFD, Neto PJD, Souza MI, Guimarães JA, Cattelan JW, Mendonça CL, et al. Physicochemical and cytological characteristics of synovial fluid in the carpal joint of mixed-breed goats. Ciencia Animal Brasileira. 2010;11(1):212-217.
19. Muheet, Malik HU, Parray OR, Bhat RA, Yatoo MI. Seasonal and periodic rhythmicity of respiratory infections in small ruminants. Biological Rhythm Research. 2020;51(7):1066-1074.
20. Peterhans E, Greenland T, Badiola J, Harkiss G, Bertoni G, Amorena B, et al. Routes of transmission and consequences of small ruminant lentiviruses infection and eradication schemes. Veterinary Research. 2004;35(3):257-274.
21. Ramírez H, San Román B, Glaria I, Reina R, Hernández MM, De Andrés X, et al. Antibody-based diagnosis of small ruminant lentivirus infection in seminal fluid. Theriogenology. 2009;72(8):1085-1096.
22. Singh DK, Paliwal OP, Dubey SC. Caprine arthritis encephalitis in Indian goats. 1997.
23. Singh R, Kumar P, Sahoo M, Bind RB, Kumar MA, Das T, et al. Spontaneously occurring lung lesions in sheep and goats. Journal of Entomology and Zoology Studies. 2017.
24. Singh R, Singh R, Kumari S. Seroprevalence study of small ruminant lentivirus infection in Indian sheep and goats. Journal of Entomology and Zoology Studies. 2018;6(1):1480-1481.
25. Sonawane GG, Tripathi BN, Kumar R, Kumar J. Diagnosis and prevalence of ovine pulmonary adenocarcinoma in lung tissues of naturally infected farm sheep. Veterinary World. 2016;9(4):365-370.
26. Tyagi RPS, Singh J. Ruminant surgery. CBS Publishers and Distributors, Delhi; 1993:318-338.
27. Valecha S. Pathology and molecular characterization of small ruminant lentiviruses (SRLVs) in sheep and goats. MVSc Thesis; 2022.
28. Waseem A, Pawaiya RVS, Singh R, Gupta VK, Rajukumar K, Mir MS, et al. Seroprevalence of caprine International Journal of Advanced Biochemistry Research https://www.biochemjournal.com arthritis encephalitis virus infection in Indian goats. 2015.
29. Zink MC, Narayan O, Kennedy PG, Clements JE. Pathogenesis of visna/maedi and caprine arthritisencephalitis: new leads on the mechanism of restricted virus replication and persistent inflammation. Veterinary Immunology and Immunopathology. 1987;15(1-2):167-180. doi:10.1016/0165-2427(87)90110-3