Feline immunodeficiency virus, feline leukemia virus and Bartonella species in stray cats on St Kitts, West Indies

Abstract

Stray cats trapped in various areas of Basseterre, the capital of St Kitts in the West Indies, were tested for infection with feline immunodeficiency virus (FIV) and feline leukemia virus (FeLV) using commercial kits. Of 99 (51 male and 48 female) cats trapped in 2006/7, 15% (12 males and three females) were positive for FIV while none were positive for FeLV. Of 72 (41 males and 31 females) cats trapped in 2009, 14% (nine males and one female) were positive for FIV while none were positive for FeLV. Polymerase chain reaction analysis revealed DNA of Bartonella species in whole blood collected from 60/95 (63%) cats trapped in 2006/7. Sequencing of the 16S–23S rRNA gene intergenic transcribed spacer (ITS) region of a convenience sample of nine amplicons and the 11 isolates made from 43 blood samples which were cultured using Bartonella alpha Proteobacteria (BAPGM) enrichment medium revealed B henselae (14) and B clarridgeiae (six).

A recent survey has shown that stray cats (unowned animals that inhabit urban areas) are common on the Caribbean island of St Kitts. ¹ Although cats are generally liked by local people, principally because they remove vermin, stray cats pose problems relating to animal welfare, damage to various wildlife populations and intrusion on natural ecosystems. They may also serve as reservoirs of important infectious agents for domesticated pet cats, such as feline immunodeficiency virus (FIV) and feline leukemia virus (FeLV). These retroviruses occur widely and are important causes of morbidity and mortality in cats. ² Although commercial vaccines are available, they are expensive, require repeated administration and do not induce protection in all vaccinates. ² If cats do become infected and develop clinical signs, treatment regimes have been described, but these are expensive and time consuming, can have severe side-effects, and may only slow the progression of the disease. ²

Stray cats can also threaten public health as they may carry a variety of zoonotic diseases. Stray cats on St Kitts are commonly infected with Toxoplasma gondii ³ but there is no data on other important agents, in particular Bartonella henselae which is a very common vector borne zoonosis. ⁴ While Bartonella species infections in cats are mostly asymptomatic, some animals might show clinical signs ⁴ and, in people, infections can result in a variety of clinical conditions including cat scratch disease, bacillary angiomatosis–peliosis, endocarditis, bacteremia and, potentially, chronic arthralgia and neurological and neurocognitive diseases. ^5–7

To determine if stray cats on St Kitts are infected with FIV, FeLV and Bartonella species, we tested stray cats captured during two neutering programs carried out between 2006/7 and 2009. Our results are reported below.

Materials and methods

As part of a stray cat neutering program operated by Ross University Veterinary School Community Practice (RUSVMCP) in 2006/7 and 2009, cats were trapped in ‘Easy Set Racoon Traps’ (Havahart, Lititz, PA, USA) and delivered to a private veterinary clinic in Basseterre or the RUSVMCP where they were anesthetized with ketamine (20–25 mg/kg) and xylazine (0.5 mg/kg) given intramuscularly. Whole blood was collected aseptically in EDTA and tested for antibodies to FIV (p15 and p24) and antigens of FeLV (p27group-specific) with a commercial kit (Snap Combo FeLV Ag/FIV Antibody Test, Idexx Laboratories Westbrook, ME, USA).

After freezing at −20°C for up to 9 months, whole blood was thawed at room temperature for processing in the Intracellular Pathogens Research Laboratory at North Carolina State University. DNA for polymerase chain reaction (PCR) amplification was extracted using the QIAamp DNA Mini Kit (Qiagen, Valencia, CA). Also, 43 blood samples with an adequate sample volume of 1–2 ml, were inoculated into 10 ml of Bartonella alpha Proteobacteria (BAPGM) liquid enrichment growth medium as previously described. ^8,9 BAPGM is a novel, chemically-modified, insect-cell culture based liquid medium that supports the growth of at least seven Bartonella species. ⁸ Following incubation for 7 days, subcultures were established by sub-inoculation from the liquid medium onto an agar plate and DNA extracted from isolates as described above.

PCR was performed with primers which amplify portions of the 16S–23S rRNA intergenic transcribed spacer (ITS) region as described previously. ¹⁰ The ITS region PCR assay used in this study has a sensitivity of 0.5 bacterial genome copies per micro-liter of sample DNA template. ⁸ Amplicons were cloned with the pGEM-T Easy Vector System (Promega, Madison, WI) and the sequences determined by Davis Sequencing (Davis, CA). Sequences obtained were compared with those in GenBank database using AlignX software (Vector NTI Suite 6.0, InforMax).

Cats with positive FIV test results were humanely euthanased regardless of clinical condition. Cats free of FIV and FeLV were neutered using standard methods and released or adopted.

The Fisher's exact test (http://www.matforsk.no/ola/fisher.htm) was used to compare prevalence data and P values ≤0.05 were considered significant.

Results

The only cats included in the study were those regarded as being adults based on their physical appearance and the presence of permanent teeth. A total of 171 adult domestic shorthair cats were trapped at 30 sites over the duration of the study; 99 from 19 sites in 2006/7 and 72 from 17 sites in 2009. The overall prevalence of positive test results was 14.6% (25/171) for FIV and 0% for FeLV (0/171). Although administration of the FIV vaccine can result in false positive test results, ² the vaccine has not been introduced onto St Kitts and we regarded cat's positive on the FIV test to be infected. The overall prevalence of FIV infection was essentially unchanged between 2006/7 (15%; 15/99) and 2009 (14%; 10/72). In both 2006/7 and 2009 most of the infected animals were male (80% and 90%, respectively). The majority of FIV positive male cats had thickened facial skin from old fight wounds and/or had abscesses, cellulitis, and/or puncture wounds from recent fights.

Cats positive for FIV were found at 32% (6/19) of the sites in 2006/7 and 18% (3/17) of the sites in 2009, but cats were only trapped at six sites in both study period periods. Cats positive for FIV were found at two of these six sites with the number of infected cats increasing from 2006/7 to 2009 (2/10 (20%) to 7/16 (44%) for one site and 2/7 (29%) to 2/2 (100%) for the other) but the differences were not significant (P=0.21, P=0.42, respectively).

Sixty of the 95 (63%) whole blood samples examined by ITS region PCR for Bartonella species gave amplicons of an appropriate size (around 530 base pairs). ¹⁰ A higher percentage of the females cats (72%) were PCR positive than males (52%), but the difference was not significant (P=0.085). Sequences of nine randomly selected amplicons were identical to B henselae (six) or B clarridgeiae (three). Isolates were obtained from 11/43 blood samples (26%) and sequence analyses identified both B henselae (eight isolates; 73%) and B clarridgeiae (three isolates; 27%).

Data was only available on the fate of cats captured in 2009; one died under anesthesia for neutering, 30 (42%) were released (15 females; 15 males) and 31 (43%) were adopted (17 females, 14 males). Young adults, irrespective of gender, that became tame and affectionate during the hospitalization period were most likely to be adopted.

Discussion

Until our study, knowledge of FeLV and FIV on St Kitts was based on laboratory tests conducted on pet cats presenting to the RUSVMCP. As almost all of these cats belonged to students attending RUSVM and originated in the US, it was difficult to determine if affected animals were infected on St Kitts or in the US. Our inability to identify FeLV in the stray cats we studied (0%; 95% confidence interval 0–2.3%) indicates the virus is not endemic on the island or is present at a very low level. We suspect the latter as local experience at the RUSVMCP shows cats are very infrequently found with FeLV infection and these inevitably are cats that have been brought to the island from the US by their student owners. Also, a similar study in Grenada failed to reveal FeLV infected cats (0/229) ¹¹ indicating the virus may be absent or at a very low level in the Caribbean region as a whole. Natural low prevalences in geographical areas have been reported elsewhere and have been attributed to genetic resistance to infection, less virulent viral strains, climatic factors, behavioral differences amongst cat populations, and geographical isolation. ¹² St Kitts and other islands with no evidence of FeLV are fortunate in that their geographic isolation should greatly facilitate the maintenance of the situation. Prevention of importation of the virus can be achieved by veterinary authorities requiring that cats being introduced to the islands have negative tests for the disease and are vaccinated. ²

The prevalence of FIV we found (14.6%) is similar to that reported elsewhere, although there is great variation within and between countries, for example, 0–22% in Vietnam, ¹³ 10% in England, ¹⁴ 11% in Israel, ¹² 9 ¹⁵ to 22% in Australia, ¹⁶ 2–23% in North America, ¹⁷ and 29% in Grenada. ¹¹ This variability can be due to a number of factors including differences in population densities, ¹⁷ level of neutering, age, gender ratios and interactions between cat colonies. Most important, however, is the presence of roaming male cats as transmission of FIV is mainly through bite wounds ¹⁸ and male cats, irrespective of their neutering status, are more likely to show territorial aggression and be involved in fights. ^14,16,19,20 This is consistent with our results where significantly more males than females were infected and most had clinical evidence of fight wounds. Decreasing or eliminating FIV from Caribbean islands by testing and removal programs would pose many problems ² but would be facilitated by their geographical isolation.

The prevalence of Bartonella species infections in cats also varies considerably, being lowest in cold countries and highest in warm countries. ²¹ The level of infection we found on St Kitts is very high by world standards, and is consistent with that reported in two other islands, Trinidad (59%) ²² and Jamaica (19%). ²³ The high prevalence of infection based upon PCR testing is not unexpected as climatic factors on the island favor survival of the cat flea, Ctenocephalides felis, which is the major vector of B henselae and B clarridgeiae. ⁴ The relatively high prevalence of B henselae compared to B clarridgeiae is common ^21,24 and was present in stray cats in Trinidad (9:1). ²² In Jamaica, however, only B henselae was found. ²³ Although B clarridgeiae appears to only rarely cause clinical infections in people and animals, ^4,5 B henselae is an important human pathogen and cat scratch disease has been described in Cuba and Jamaica. ²³ Further, B henselae causes bacillary angiomatosis–peliosis and bacteremia in immunosuppressed people ⁵ and the Caribbean has the highest incidence of HIV/AIDS outside of sub-Saharan Africa. ²⁵ Further studies are indicated to determine the extent and importance of Bartonella species infections in people in the region.

In summary, our study has shown FIV and B henselae and B clarridgeiae infections are common in stray cats on St Kitts but FeLV appears to be absent from the island. This has important implications for veterinary and human health workers on the island and those dealing with tourists and animals that might visit the island. The data also adds to our knowledge of the distribution and prevalence of infections with these organisms.