Health impact evaluation of alternative management systems in vicuña (Vicugna vicugna mensalis) populations in Peru

DOI 10.1007/s11250-014-0543-3 REGULAR ARTICLES Health impact evaluation of alternative management systems in vicuña (Vicugna vicugna mensalis) populations in Peru Veronica Risco-Castillo & Jane Collins Wheeler & Raúl Rosadio & Francisco Javier García-Peña & Ignacio Arnaiz-Seco & Domingo Hoces & Hugo Castillo & Álvaro Veliz & Luis Miguel Ortega-Mora Accepted: 20 January 2014 # Springer Science+Business Media Dordrecht 2014 Abstract To determine the impact of farming over vicuña population in Peru, serum samples were collected from 207 vicuñas (126 captive vicuñas and 81 free-ranging vicuñas) and 614 domestic South American camelids (571 alpacas and 43 llamas), in ten Andean communities at the Salinas y Aguada Blanca reserve, province of Arequipa, southern Peru. Samples were tested for the presence of leptospirosis, foot and mouth disease (FMD), bovine viral diarrhea (BVD), bovine herpesvirus type 1 (BHV-1), brucellosis, bluetongue disease (BT), paratuberculosis, and neosporosis. Serological results showed that 1.9 % (4/207) of vicuñas, 18.6 % (106/571) of alpacas, and 23.3 % (10/43) of llamas were positive to one or more Leptospira serovars. One percent of vicuñas (2/207) and 2.4 % of domestic camelids (15/614) had Neospora caninum antibodies tested by ELISA, but only two vicuñas and two V. Risco-Castillo (*): L. M. Ortega-Mora SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain e-mail: vrisco@gmail.com J. C. Wheeler CONOPA Instituto de Investigación y Desarrollo de Camélidos Sudamericanos, Lima, Peru R. Rosadio: D. Hoces : H. Castillo : Á. Veliz Faculty of Veterinary Medicine, Universidad Nacional Mayor de San Marcos, Avda. Circunvalación s/n San Borja, Lima, Peru F. J. García-Peña Laboratory of Animal Health of Algete, National Reference Laboratory for Animal Leptospirosis, Madrid, Spain I. Arnaiz-Seco Laboratory of Animal Health and Production of Galicia, Lugo, Spain Present Address: V. Risco-Castillo INSERM, UMR-S 945, Paris, France alpacas were confirmed by Western blot. Epidemiological evaluation found an association of leptospirosis to sex and age (p<0.001), with female subjects older than 2.5 years at higher risk of infection. Interestingly, antibodies against Leptospira serovars were only found in captive vicuñas. This is the first study where health status of free-ranging and captive vicuñas has been compared. Results indicate minimal tonilpresenceoffmd,bvd,bhv-1,brucellosis,bt, paratuberculosis, and neosporosis allied to health disorders in our sample. The detection of seropositive animals against Leptospira, however, unveils the likely significance of leptospirosis in wild and domestic South American camelids, the impact of mixed husbandry over vicuña population and the risk to human health. Keywords Free-ranging. Captive. Domestic camelids. Leptospirosis. Arequipa Introduction The wild South American camelids, vicuña (Vicugna vicugna) and guanaco (Lama guanicoe), and their domestic forms, the alpaca (Vicugna pacos) and llama (Lama glama), are native species inhabiting the Andean plateau of South America many millennia ago (Wheeler 2012). Today, camelids rearing in Peru and Bolivia provide fiber, meat, and fuel to impoverished Andean herders, who hold most of these livestock (Wheeler and Hoces 1997). Current vicuña distribution is limited to areas of extreme elevation in the Andes (Wheeler 2012), and for several decades, vicuña has been in danger due to the economic value of their fiber (Appendix II of Convention on International Trade in Endangered Species (CITES)), but protection laws from involved countries in the last 20 years have succeeded on recovering the population (Cox 2003). Several management systems

have been established to ease recollection of their fiber and to avoid a likely negative ecological impact if forcing them to be reared as domestic animals (Sahley et al. 2007). Nevertheless, they allow their temporary or permanent captivity, placing vicuñas in contact to domestic animals and increasing the likelihood of disease transmission between species (Lichtenstein et al. 2002; Vilá 2002; Wheeler and Hoces 1997). International organizations recommend monitoring camelid health and determination of their susceptibility to diseases affecting domestic livestock (Office International des Epizooties (OIE) 2010). Although research regarding the role of domestic camelids as reservoirs of diseases is increasing worldwide (Kapil et al. 2009; Tibary et al. 2006), knowledge of vicuña health status is still limited. There are few reports of infectious diseases in wild vicuñas (Marcoppido et al. 2010), and most results are based on detection in a limited number of individuals (Celedon et al. 2001; Chavez-Velasquez et al. 2005). In domestic camelids, the presence of antibodies against bovine viral diarrhea virus (BVDV), bovine herpesvirus type 1 (BHV-1), bluetongue virus (BTV), Mycobacterium avium subsp. paratuberculosis, Brucella spp., and Toxoplasma gondii (Acosta et al. 1972; Chavez-Velasquez et al. 2005; Puntel et al. 1999; Rivera et al. 1987; Rosadio et al. 1993) has been reported. It is likely that the low reproductive profiles in alpacas and llamas may be due to infectious agents such as BVDV (Rosadio et al. 2003) orparasitessuchast. gondii and Neospora caninum (Chavez-Velasquez et al. 2004; Serrano- Martinez et al. 2004, 2007). Vulnerability of vicuñas to those diseases cannot be overruled. In recent years, there has been an increase in reports of the role of domestic and wild animals as reservoirs of zoonotic agents such as Leptospira in developing countries, and hence their potential as a source of infection in humans (Jimenez- Coello et al. 2010; Jorietal.2009). Some authors have detected exposure to several Leptospira serovars (Hill and Wyeth 1991; Santos et al. 2009) in llamas and alpacas, and it has been suggested an association with reproductive problems as well as a potential zoonotic risk (Marin et al. 2007). The goal of this study was to determine and compare the health status of free-ranging and captive vicuñas and to establish the potential exposure to infectious agents from the neighboring domestic camelids reared in the Salinas and Aguada Blanca National Reserve. Our results will be a starting point in the understanding of the effect of mixed husbandry and health condition in both wild and captive vicuñas. Materials studied Area description, animals, and sample collection We sampled ten Andean communities located at 2,800 6,050 m above sea level, in 366.94 10 3 ha Salinas and Aguada Blanca National Reserve (Fig. 1), in the province of Arequipa, Peru (71 13 W, 16 5 S), between October and December of 2005. Mean temperature ranged from 5 to 15 C and we recorded no precipitation (mean 0 mm 3 ). At this season, the local population practices the traditional chaccu during which both wild and captive vicuñas are rounded up for shearing. We also sampled llamas and alpacas located in areas adjacent to vicuñas (Table 1). We used the software Win Episcope (v. 2.0) to determine the number of samples to collect, considering a prevalence of at least 5 % for each disease under study, with a confidence interval of 99 %. Additionally, we calculated the sampling size for each species in each community in relation to the total number of animals in the region (2004 livestock survey, Ministry of Agriculture, Peru). Trained personnel collected blood samples from either the jugular or femoral veins and checked the animals for any sign of disease. Sample tubes were immediately stored on ice and transported to the laboratory for serum recovery. Ten serum samples were also collected from alpacas born and raised at a high-biosecurity farm in Simplon village, south Switzerland, to be included as negative controls for all diseases tested. Sample analysis We performed enzyme-linked immunosorbent assays (ELISA), following manufacturer s recommendation for the detection of BVD, BHV-1, foot and mouth disease (FMD), BTV, paratuberculosis, brucellosis (Brucella abortus, Brucella ovis, and Brucella melitensis), and neosporosis (N. caninum; Table 2). On the other hand, diagnosis of leptospirosis was carried out by the microscopic agglutination test (MAT) and using 17 serovars (Leptospira borgpetersenii serovars Australis, Ballum, Bataviae, Djasiman, Hardjo, Javanica, and Tarassovi; Leptospira interrogans serovars Autumnalis, Bratislava, Canicola, Copenhageni, and Pomona; Leptospira kirschneri serovars Cynopteri and Grippotyphosa; Leptospira noguchi serovars Louisiana and Panama; and Leptospira weilii serovar Sarmin). Samples positive to brucellosis or neosporosis by ELISA were confirmed by the rose bengal test or by Western blotting (Chavez-Velasquez et al. 2004) with small modifications, respectively. In the case of neosporosis test confirmation, nitrocellulose membranes containing soluble extract of N. caninum were incubated for 1 h at 37 C in agitation with the serum samples at 1:100 dilution in phosphate-buffered saline (PBS), washed three times, and incubated for 1 h at 37 C in agitation with a peroxidase-conjugated sheep antillama antibody at 1:1,000 dilution in PBS. Finally, membranes were developed with 4-chloro-1-naphthol. Statistical analysis We tabulated and analyzed the results statistically with the SAS system software (v. 7) and the GraphPad Prism software

Fig. 1 Geographic location of Salinas y Aguada Blanca Natural Reserve and Andean community samples (v. 5), and Fisher s exact test was applied to determine the association between a given diagnosed disease and sex (male or female), age (less than 2 years old, 2 5 years old, or more than 5 years old), species and management system (freeranging vicuña, captive vicuña, or domestic camelids), and location (Andean community) or geographical region within the National Park (north, south, east, or west). Results The results showed that 1.9 % (4/207) of vicuñas had antibodies against different serovars of Leptospira, while seroprevalence in the domestic camelids was of 18.9 % (116/614), including 106 of 571 alpacas and 10 of 43 llamas. The serovars detected were Pomona, Autumnalis, Copenhageni, and Bratislava. The antibody titers ranged between 1:100 and 1:30,000; 93 of alpacas positive to Pomona were also positive to Autumnalis (n=64), Bratislava (n=8), or Copenhageni (n= 20), 19 alpacas were positive to three serovars, and 4 alpacas were positive to the four serovars, while 4 llamas were positive to two serovars (Pomona and Autumnalis). Among vicuñas, two animals were positive to two serovars (Pomona and Autumnalis or Copenhageni) and one was positive for the four serovars. Furthermore, an association between seropositivity and species (p <0.0001), sex (p =0.0017), age (p<0.001), and region of origin (p=0.069) was observed, with the highest risk of infection found in alpacas, in female Table 1 Location and distribution of animals sampled Location at reserve Andean community Free-ranging vicuna Captive vicuna Alpaca Lama North Chalhuanca 27 62 4 North Chuca-Imata 14 70 1 North Colca Huallata 29 63 North Pillone 20 62 6 South Salinas Huito 2 31 East Carmen Chaclaya 21 5 East Tarucani 22 56 17 West Ampi 22 72 3 West Tambo Cañahuas 45 71 West Tocra 26 63 7 Total 81 126 571 43

Table 2 Serologic tests performed, methods used, and cutoff value established for the diseases studied Disease Technique Kit Manufacturer Sensibility Cutoff value BVD Competitive ELISA ELISA BVD/MD/BD Pourquier BVD/MD/BD P80 40 % inhibition BHV-1 Blocking ELISA HerdChek IBRgB Idexx BHV-1 gb 55 % blocking Paratuberculosis ELISA ID Screen ID.Vet M. avium paratuberculosis 70 % S/P FMD Competitive ELISA CHEKIT-FMD 3ABC Bommeli-Intervet Serotypes A, O and C According to the manufacturer Leptospirosis MAT NA In house 17 serovars 1:100 BT Competitive ELISA Ingezim BTV Compac Ingenasa 24 BTV serotypes 50 % inhibition Brucellosis Competitive ELISA Ingezim Brucella compac Ingenasa Multispecies (Brucella spp.) 60 % inhibition Neosporosis Competitive ELISA Chekit Neospora Idexx N. caninum According to the manufacturer BVD bovine viral diarrhea, BHV-1 bovine herpesvirus type 1, FMD foot and mouth disease, BT bluetongue disease, ELISA enzyme-linked immunosorbent assay camelids, and in animals older than 2.5 years from the eastern area of the National Reserve. Interestingly, all positive vicuñas came from the western region (Toccra, Ampi) and were kept in captivity. Regarding N. caninum, 1 % of the vicuñas (2/207) and 2.4 % of the domestic camelids (15/614) were found to have antibodies when tested by ELISA. However, only two vicuñas and two alpacas were confirmed by Western blotting to have antibodies specific to N. caninum (data not shown). No association was found with any of the variables analyzed, and the vicuñas and alpacas came from different Andean communities (alpacas came from the north region and vicuñas came from the east and west of the National Park). Regarding the serodiagnosis of Brucella sp., 2.9 % of vicuñas (6/207), 10.5 % of alpacas (60/571), and 9.3 % of llamas (4/43) were seropositive when tested by ELISA. Nevertheless, all samples resulted negative when retested using the reference rose bengal test. All animals were negative for the other diseases analyzed (FMD, BVD, BHV-1, BTV, and paratuberculosis). No clinical signs were observed in animals sampled during the study. Discussion To date, the health status of South American camelids grazed in shared pastures has been poorly investigated. This is the first study that analyzes and compares the health status of freeranging and captive vicuña with that of domestic camelids (llamas and alpacas) reared in the same area. An association of BVD infection with respiratory disease, abortion, ill thrift, and diarrhea has been described in llamas and alpacas (Belknap et al. 2000; Carman et al. 2005; Foster et al. 2007; Mattson et al. 2006), and serological findings report up to 11 % prevalence in Peruvian alpacas (Rivera et al. 1987). Most seropositive camelids in Peru have been found in mixed farms (with other livestock such as sheep and cattle) than in those where only camelids are bred (Rosadio et al. 2003). The seronegativity in the animal population of this study agrees, in the case of BTV, with a lack of competent vector speciesinthisregionduringthedryseason(lager2004) and confirms the importance of veterinary surveillance to avoid contact and disease transmission to naïve populations from infected livestock. This is especially true for free-ranging vicuña since any other control measure is unacceptable in wild species. Our results confirm for the first time the presence of the Leptospira serovars Pomona, Autumnalis, Bratislava, and Copenhageni among the captive vicuña populations in Peru and the likely circulation of this pathogenic agent between domestic and wild camelids. Similar seroprevalences have been found in other regions (Hodgin et al. 1984; Llorente et al. 2002),andhighantibodytitershavebeenassociatedto active infection during the rainy season (Santos et al. 2009). While Pomona and Autumnalis were the most seroprevalent serovars among the alpaca, llama, and captive vicuñas, there were no animals seropositive to Hardjo, which is the more adapted to true ruminants (Gerritsen et al. 1994). These results coincide with previous studies (Santos et al. 2009; Suárez et al. 2007) and could infer the adaptation of at least Pomona to South American camelids, as previously documented for cattle from swine. Precautions must be taken regarding the risk of cross-reaction between serovars, as seen in animals with high titers against Pomona and lower titers against Autumnalis or Bratislava (Kingscote 1986). Interestingly, Leptospira seroprevalences in both captive vicuña and domestic camelids from Toccra were unexpectedly high, although no association between geographic location and management system was found. Also, Copenhageni antibodies were detected in the three species studied regardless of their location or age. Similar results have been observed by other authors (Rosadio et al. 2003), and while the source of transmission may come from contaminated watercourses (Felt et al. 2011; Karesh et al. 1998), this risk would have been reduced or nonexistent during our dry season study, so the detection of high titers against Pomona and Autumnalis could indicate

close contact with infected livestock, rodents, or even guinea pigs (Gressler et al. 2010). The high antibody titers found in female subjects older than 5 years during the dry season could also indicate a high risk of venereal transmission for these serovars (Little 1986). Interestingly, we found higher prevalence in adult female subjects than that in adult male subjects, in contrast to what has been observed by other authors (Santos et al. 2009). While literature reports a rather low seroprevalence of leptospirosis in cattle from this region (Arias et al. 2011), the role of wildlife as a source of transmission has been described. Wild rodents such as the northern viscacha and the colilargo or Andean pygmy mouse (Lagidium peruanum, Oligoryzomys andinus, Thylamys sp.) opossums (Didelphis marsupialis), weasels (Mustela frenata), or hog-nosed skunks (Conepatus chinga) could be implicated in Leptospira transmission to vicuñas (Bunnell et al. 2000; Vanasco et al. 2003). Pomona and Copenhageni have been isolated from renal tissue of wild mice of the Akodon genus (Zamora and Riedermann 1999), and the Andean fox (Lycalopex culpaeus) may also play a role in the epizootiology of Pomona and Icterohaemorrhagiae, as has been described elsewhere for other fox species (Millan et al. 2009). Although we report low seroprevalence rates for the southern region, our findings highlight the importance of establishing proper sanitary measures to protect both human and animal population health and well-being (Cespedes et al. 2006). Seroprevalence against neosporosis in the vicuñas and domestic camelids studied was very low. Previous studies have also detected a moderate to low seroprevalence of neosporosis in South American camelids (Chavez-Velasquez et al. 2004; Wolfetal.2005), and the possible reduced susceptibility of llamas has been suggested to propitiate their role as intermediate hosts (More et al. 2008). This possibility would agree with our results since we found no association between diagnosis and location of the animals, although the Neospora-positive vicuñas shared grazing areas with domestic camelids, or a higher risk of horizontal transmission via dogs or wild carnivores known to inhabit the region and predate South American camelids. The results presented here provide insights that could aid local and governmental authorities, together with Andean communities, to make informed decisions regarding management strategy of wild South American camelids in the future and to establish and make measures better adapted to protect their health status. Although the close presence of domestic camelids and vicuñas might not be a factor affecting the health status of the latter, it would also be advisable to carry out broader studies to analyze the zoonotic risk of leptospirosis and other diseases affectingsouthamericancamelids. Acknowledgments We are grateful to the following people who generously helped us in this study: C. Flores and D. Arias (CONACS), A. Cornejo and M. Avendaño (INRENA), H. Escarza (ACVIRA), and Andean communities from Salinas y Aguada Blanca National Reserve (Toccra, Tarucani, Chuca-Imata, Pillone, Chalhuanca, Ampi, Colca- Huallata, Tambo Cañahuas, Salinas Huito, and Carmen Chaclaya). We also thank J.M. Blasco (Center of Agrofood Technology and Research of Aragon, Spain) for analyzing samples against brucellosis by MAT analysis and Dr. J. Kuonen (Alpaka-Zucht Simplon) for kindly providing samples with negative control alpaca serum. This study was financed by a grant for cooperation for the development from the University Complutense of Madrid (projects for cooperation to development Complutense University of Madrid, ref. 14/05: health impact evaluation of new management systems on vicuña population of Peru). 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