Seroprevalence and risk factors for Toxoplasma gondii exposure in Australian feral and stray cats using an in-house modified agglutination test.

Toxoplasma gondii is a globally distributed zoonotic protist, capable of infecting all warm-blooded animals. In Australia, cats (Felis catus) are the only definitive host capable of spreading T. gondii infection via oocysts. Free-roaming cats are widespread in Australia and can play a central role in the ecology of T. gondii. Therefore, understanding the epidemiology of this parasite in stray and feral cats is essential to understanding the potential risk of infection in animals and humans. Due to a lack of easily accessible commercial kits, an in-house modified agglutination test (MAT) was established to test for IgG antibodies against T. gondii, using cell culture-derived T. gondii tachyzoites, and compared with a commercial MAT. A total of 552 serum samples collected during 2018 - 2021 from stray (n = 456) and feral cats (n = 90) (samples with missing data n = 6) from four Australian states, representing different age groups of both sexes, were screened for antibodies against T. gondii. Risk factors for T. gondii infection were assessed using multivariable logistic regression analysis. The in-house MAT had excellent agreement with the commercial MAT and provided a reliable and economical serological tool for T. gondii screening in animals. The overall observed seroprevalence for T. gondii in cats was 40.4 % (223/552). Bodyweight (as a proxy for age), geographical location, season and whether cats were feral or stray, were factors associated with T. gondii seropositivity in cats. Sex was not found to be a risk factor for T. gondii infection in feral and stray cats. This study shows that Australian stray and feral cats have a high T. gondii seroprevalence, which may translate to significant health impacts for wildlife species, livestock and the public.

Surveillance for Toxoplasma gondii, Brucella spp., and Chlamydia spp. in Australian Fur Seal (Arctocephalus pusillus doriferus) Abortions.

The intracellular pathogens Toxoplasma gondii, Brucella spp., and Chlamydia spp. are all known causative agents of abortion in wildlife. Both T. gondii and Brucella spp. have been identified in marine mammal abortions and a limited number of studies have detected their potential presence in Australian fur seals (Arctocephalus pusillus doriferus), but data are sparse for these pathogens in Australian fur seal breeding colonies. Australian fur seals have been shown to have a high degree of third-trimester pregnancy loss in one of their largest breeding colonies. Additionally, pup production has declined at the largest breeding colony for the species. This study surveyed the presence of T. gondii, Brucella spp., and Chlamydia spp. as potential infectious causes of this reproductive loss. Aborted fetuses were collected from two of the largest breeding colonies for the species, Seal Rocks (n=19) and Kanowna Island (n=34). These were examined grossly and through histopathological evaluation, in conjunction with molecular testing for all three pathogens. Placentas were collected from full-term births during the pupping season from Kanowna Island (n=118). These were used to compare the molecular prevalence of the three pathogens in presumed successful pregnancies. Chlamydia spp. was not detected in aborted fetuses in this study. Brucella spp. was detected with PCR in both aborted fetuses (9.4%) and placentas from full-term births (3.4%), and T. gondii was detected using routine histopathology (n=2/53), immunohistochemistry (n=3/4), and PCR (n=4/53) in tissues from aborted fetuses. Toxoplasma gondii was present in 7.5% of third-trimester abortions and absent from all full-term placentas. Brucella spp. was detected in both aborted fetuses and full-term placentas. This is the first description of vertical transmission of T. gondii in a marine mammal from the southern hemisphere.

Towards a pan marsupial sero-immunological tool in the demanding field of wildlife serology: Marsupial immunoglobulin-binding capability with protein A/G, protein L and anti-kangaroo antibody.

Detection of infections in wildlife species is increasingly important to reduce the risk of spreading zoonotic and economically important parasites, understand disease epidemiology and promote the conservation of wildlife species. Serological tests are key in disease diagnosis and surveillance by detecting immunoglobulins against infectious agents. However, the need for species-specific reagents has limited the application of serological tests in wildlife species. This study evaluated the serum immunoglobulin-binding capability of polyclonal anti-kangaroo antibody and two non-species-specific reagents, including protein A/G and protein L, with the largest range of Australian marsupial species so far, including 32 species representing three major marsupial orders. Immunoglobulin-binding capability was assessed using immunoblotting, enzyme-linked immunosorbent assay and Western blot techniques. Variation in immunoglobulin-binding capability was observed between the three reagents and across the species tested, both across but also within taxonomic groups. Taxonomic distance was thus not always a good predictor of immunoglobulin-binding affinity, emphasizing the need to validate these reagents for each species separately. However, all three reagents bound with the serum immunoglobulins of most marsupial species tested. The findings of this study provide a valuable reference for species differences in affinity to protein A/G, protein L and anti-kangaroo antibody, assisting in the selection of appropriate reagents and the development of sero-immunological assays in Australian marsupials.

Descriptive Comparison of ELISAs for the Detection of Toxoplasma gondii Antibodies in Animals: A Systematic Review.

Toxoplasma gondii is the zoonotic parasite responsible for toxoplasmosis in warm-blooded vertebrates. This systematic review compares and evaluates the available knowledge on enzyme-linked immunosorbent assays (ELISAs), their components, and performance in detecting T. gondii antibodies in animals. Four databases were searched for published scientific studies on T. gondii and ELISA, and 57 articles were included. Overall, indirect (95%) and in-house (67%) ELISAs were the most used types of test among the studies examined, but the 'ID Screen® Toxoplasmosis Indirect Multi-species' was common among commercially available tests. Varying diagnostic performance (sensitivity and specificity) and Kappa agreements were observed depending on the type of sample (serum, meat juice, milk), antigen (native, recombinant, chimeric) and antibody-binding reagents used. Combinations of recombinant and chimeric antigens resulted in better performance than native or single recombinant antigens. Protein A/G appeared to be useful in detecting IgG antibodies in a wide range of animal species due to its non-species-specific binding. One study reported cross-reactivity, with Hammondia hammondi and Eimeria spp. This is the first systematic review to descriptively compare ELISAs for the detection of T. gondii antibodies across different animal species.