Oral Vaccination of Captive Small Indian Mongoose (Herpestes auropunctatus) against Rabies

The small Indian mongoose (Herpestes auropunctatus), native to southern Asia, was introduced to many islands worldwide for control of pest animals in the late 19th or early 20th century (Barun et al., 2011). Most introductions occurred in tropical areas, but mongooses also have been successfully released on several islands in the Adriatic Sea (Tvrtkovic and Krystufek, 1990), with climatic conditions during the winter months well below the mongoose's thermal tolerance of 10 C (Nellis and McManus, 1974; Cavallini and Serafine, 1995). From these islands the animal has also invaded the mainland with fox-mediated rabies and is expanding its range southward (Cirovic et al., 2011). So far, no rabies cases in mongooses have been reported from this area. However, the species has been identified as a rabies reservoir on some of the Caribbean islands, including Cuba, The Dominican Republic, Grenada, and Puerto Rico, since the mid-20th century (Everard and Everard, 1992). Mongoose rabies on the islands is a result of independent spillover infections from rabid dogs (Nadin-Davis et al., 2006). Attempts to control mongoose rabies through culling had only a transient effect (Everard and Everard, 1992). Hence, oral vaccination of mongoose has been suggested as a more promising alternative (Blanton et al., 2006). Oral vaccination campaigns against rabies in several wildlife reservoir species have been successfully implemented in Europe and North America (Rupprecht et al., 2008). Experimental studies have shown that mongooses can be vaccinated against rabies by the oral route (Blanton et al., 2006). Also, field studies showed that bait was readily accepted by mongooses (Linhart et al., 1993; Creekmore et al., 1994). However, a well-accepted bait does not automatically make it an efficient vaccine delivery vehicle for oral administration. A major limitation is the small size of the animal; hence, existing oral rabies vaccine bait for larger carnivores such as foxes (Vulpes vulpes) and raccoons (Procyon lotor) are not optimal for vaccine delivery. The large size of the bait often leads to separation and consequently rejection of the vaccine container or spillage of the vaccine. Therefore, a bait specifically targeted to mongooses needs to be developed (Linhart et al., 1993). We developed and tested a suitable bait to deliver vaccine to mongooses based on their immune response to the vaccine. The required permits for the animal studies were obtained from the appropriate authorities (import permit 53-5-V-La-1004/12-42502; bait study 42502-3-647 IDT; immunogenicity study 42502-3-658 IDT). Six male mongooses were caught using baited box traps on the rabies-free island Korčula, Croatia, and transported to the Animal House at IDT Biologika GmbH, Germany (CVED reference CVEDA.SI.2012.0000298-V1).

During the bait studies, conducted in the outdoor animal enclosure, mongooses were kept individually in open wire cages (200×100×80 cm) with the bottom partially covered with plastic boards. The top was covered with sheets to protect the animals from rain and direct sunlight. Inside the cages trays filled with water or sand, tree branches, and observation posts were placed for environmental enrichment. A wooden box (65×95×60 cm) containing a sleeping box filled with straw was attached to each wire cage. Cages were cleaned at least once a day. Animals were fed daily with 40 g of a mixture of 26% poultry, 22% fish, and 29% cattle (heart/liver/kidney/rumen). Laboratory mice, fruit, and hard-boiled eggs were also offered. Water was provided ad libitum.

Several natural products (e.g., poultry, eggs, mice, fruits) were tested as bait attractants; the mongooses clearly preferred animal-derived products. During the next phase, selected substances were mixed with a proprietary unspecified bait matrix and offered initially simultaneously with the standard food, separate or homogeneously mixed. The best results were obtained with egg-flavored bait. Finally, a vaccine blister filled with dyed water was incorporated into the egg-flavored bait matrix and offered to each of the animals on two occasions. Direct observation of the animals was not possible; therefore, cameras (RCX-2, Leupold, Leupold & Stevens Inc., Beaverton, Oregon, USA) were placed in each cage. All animals accepted the bait and perforated the blister except for one animal during the second test run. Although the blister (28×20×9 mm) containing a volume of 1.0 mL was relatively small, in two of 11 attempts, considerable spillage of dyed water (>50%) onto the ground was observed.

For vaccination, the animals were sedated and relocated to individual cages within an isolation unit inside the animal house. A blood sample (B0; Table 1) was collected by claw clipping on the day of vaccination. Animals were sedated with 5 mg ketamine (Ketamin, WDT, Garbsen, Germany) and 0.1 mg Medetomidine (Domitor®, Janssen Animal Health, Neuss, Germany). Two of the six sedated and relocated animals received 1.0 mL by direct oral instillation with SPBN GASGAS (10^8.5 focus-forming units (FFU)/mL), a highly attenuated live rabies virus construct developed using reverse genetics (Faber et al., 2009). The other four animals were offered the selected vaccine bait 1 wk after relocation, which contained approximately 0.6 mL SPBN GASGAS (10^8.5 FFU/ml). At the end of the study, 28 days postvaccination, the animals were sedated with a mixture of 50 mg ketamine and 10 mg xylazine (Sedaxylan, Eurovet Animal Health, Bladel, the Netherlands) and euthanatized with 300 mg pentobarbital (Release®, WDT). A second blood sample (B1; Table 1) was collected. Blood samples were examined for rabies virus–neutralizing antibodies (VNA) using the rapid fluorescence focus inhibition test (Smith et al., 1973) with adaptations as described by Cox and Schneider (1976). Of the animals that seroconverted, the two animals receiving the vaccine construct by direct oral instillation had much higher VNA titers than the animals offered a vaccine bait (Table 1). Three of four mongooses offered a bait developed an immune response >0.5 IU/mL (range: 0.63–1.13 IU/mL), but the response was less pronounced than in the two animals offered 1.0 mL of the vaccine by direct oral instillation (1.91 and 4.79 IU/mL).

Previous studies in mongooses using very similar constructs showed that the presence of VNA was indicative for protective immunity (Blanton et al., 2006). SPBN GASGAS has been tested in other species such as raccoons, and animals with VNA were consistently protected against a relevant rabies infection (Blanton et al., 2007). Therefore, no challenge infection was performed. Although three of four mongooses offered a bait clearly seroconverted, it seems that improvement is needed in delivery of an adequate immunogenic dose, especially relative to vaccine spillage. Meanwhile, trials should be undertaken to test the suitability of this experimental oral rabies vaccine bait under field conditions in areas with mongoose-mediated rabies. Such studies would assess bait uptake under field conditions as well as bait depletion by nontarget species such as rats (Rattus spp.) and feral cats (Felis catus; Linhart et al., 1993). Also mongoose-specific baiting strategies must be developed in view of certain aspects of the behavioral ecology of this small carnivore that differ considerably compared with other species targeted for oral rabies vaccination. For example, the relatively long breeding season, dispersal peak of juveniles, and extended home range overlap need to be considered.

An additional consideration is the reported naturally acquired immunity against rabies, with more than 40% of a mongoose population having antibodies against rabies in one field study (Everard et al., 1981). Under such circumstances, a much lower vaccination coverage through bait uptake may be required to achieve sufficient herd immunity to interrupt transmission. Conversely, high levels of naturally acquired immunity may indicate that the contact rate among mongooses and subsequently transmission rate of rabies virus is extremely high, which may require much higher vaccination coverage than was required for the elimination of fox rabies. However, as illustrated in this study, successful immunization of mongoose through a single vaccine bait suggests that mongoose rabies control in the Caribbean by means of oral rabies vaccination may be feasible.

Special thanks to Hans-Joachim Bätza for his support in organizing this study. Also the dedicated work of the people looking after the animals is highly appreciated. We are grateful to the assistant editor and two anonymous reviewers for their comments and contributions to this manuscript. With the exception of Ivana Lojkic and Thomas Müller, all authors are full-time employees of a company producing, among other products, oral rabies vaccine bait.