Synthetic Particulate Subunit Vaccines for the Prevention of Q Fever.

Coxiella burnetti is an intracellular bacterium that causes Q fever, a disease of worldwide importance. Q-VAX® , the approved human Q fever vaccine, is a whole cell vaccine associated with safety concerns. Here a safe particulate subunit vaccine candidate is developed that is ambient-temperature stable and can be cost-effectively manufactured. Endotoxin-free Escherichia coli is bioengineered to efficiently self-assemble biopolymer particles (BPs) that are densely coated with either strings of 18 T-cell epitopes (COX-BP) or two full-length immunodominant antigens (YbgF-BP-Com1) all derived from C. burnetii. BP vaccine candidates are ambient-temperature stable. Safety and immunogenicity are confirmed in mice and guinea pig (GP) models. YbgF-BP-Com1 elicits specific and strong humoral immune responses in GPs with IgG titers that are at least 1 000 times higher than those induced by Q-VAX® . BP vaccine candidates are not reactogenic. After challenge with C. burnetii, YbgF-BP-Com1 vaccine leads to reduced fever responses and pathogen burden in the liver and the induction of proinflammatory cytokines IL-12 and IFN-γ inducible protein (IP-10) when compared to negative control groups. These data suggest that YbgF-BP-Com1 induces functional immune responses reducing infection by C. burnetii. Collectively, these findings illustrate the potential of BPs as effective antigen carrier for Q fever vaccine development.

Pathogenic Leptospira Species Are Present in Urban Rats in Sydney, Australia.

Leptospirosis is an emerging disease among people and dogs in Sydney, Australia. However, the routes of Leptospira transmission in these cases, and in particular the possible role of rats as reservoirs of infection in Sydney, are unknown. Rats were collected within the City of Sydney Council area and their kidneys were tested for pathogenic Leptospira DNA by real-time (q)PCR. A subset of rats also had qPCR testing performed on whole blood and urine, and Microscopic Agglutination Testing (MAT) that included a panel of 10 Leptospira serovars from nine different Leptospira serogroups was performed on a subset of serum samples. Based on qPCR testing, the proportion of rats with Leptospira DNA in their kidneys was 9/111 (8.1%). qPCR testing of blood samples (n = 9) and urine (n = 4) was negative. None of the 10 serum samples tested MAT positive. A primary cluster of qPCR-positive locations was detected based on six infected rats, which partially overlapped with a previously identified cluster of canine leptospirosis cases in Sydney. These findings suggest that rats in Sydney might play a role in the transmission of leptospirosis to dogs and people. Further testing of rats in Sydney and investigation into other possible wildlife reservoirs of infection and environmental sources of leptospires are needed.

Spatial and Temporal Movements of Free-Roaming Cats and Wildlife in Two Local Government Areas in Greater Sydney, Australia.

Free-roaming cats pose a risk to their own health and welfare, as well as to the health and welfare of wildlife and humans. This study aimed to monitor and quantify area-specific free-roaming cat movement. Two local government areas (LGAs) in Greater Sydney were included, Campbelltown (CT) and the Blue Mountains (BM). Motion-capture cameras were installed on 100 volunteer properties (50 per LGA) to indirectly capture animal movements over two months. Transect drives were completed eight times (four per LGA) to directly observe roaming cats in residential areas. The cameras and transects both identified higher free-roaming cat numbers in CT (density of 0.31 cats per ha, resulting in an estimated abundance of 361 cats in the 1604 ha of residential area) than the BM (density of 0.21 cats per ha, resulting in an estimated abundance of 3365 cats in the 10,000 ha of residential area). More wildlife events were captured in the BM (total = 5580) than CT (total = 2697). However, there was no significant difference between CT and the BM for cat events (p = 0.11) or wildlife events (p = 0.32) observed via the cameras. Temporally, cats were observed via the cameras throughout the entire day with peaks at 9:30 am and 8:00 pm in the BM, and 7:00 am and 12:00 pm in CT. Overlaps in activity times were recorded for free-roaming cats with bandicoots (BM), possums (BM), and small mammals (BM and CT). This study demonstrates that camera monitoring on private property and transect drives are useful methods to quantify free-roaming cat abundance to inform cat management interventions.

Field Performance of a Rapid Test to Detect Progressive, Regressive, and Abortive Feline Leukemia Virus Infections in Domestic Cats in Australia and Germany.

Different feline leukemia virus (FeLV) infection outcomes are possible in cats following natural exposure, such as progressive infections (persistent viremia), regressive infections (transient or no viremia followed by proviral persistence) and abortive infections (presence of only antibodies). Laboratory-based testing is currently required for categorization of infection outcomes in cats. The aim of this study was to evaluate the field performance of a novel, rapid, combination point-of-care (PoC) test kit commercially available in Europe (v-RetroFel®Ag/Ab; 2020-2021 version) to determine different FeLV infection outcomes by concurrent detection of FeLV antigen (p27) and antibodies against FeLV transmembrane envelope protein (p15E). A secondary aim was to evaluate the performance of the same test kit (v-RetroFel®FIV) to determine positive/negative feline immunodeficiency virus (FIV) infection status by the detection of antibodies to FIV capsid protein (p24) and transmembrane glycoprotein (gp40). Two cohorts of domestic cats were recruited and tested with v-RetroFel® using plasma or serum, including cats in Australia (n = 200) and cats in Germany (n = 170). Results from p27 antigen PoC testing, proviral DNA PCR, and neutralizing antibody testing or testing for antibodies against non-glycosylated surface unit envelope protein (p45) were used to assign cats to groups according to different FeLV infection outcomes. Testing with a laboratory-based FeLV p15E antibody ELISA was also performed for comparison. In the first cohort, v-RetroFel®Ag/Ab correctly identified 89% (109/122) FeLV-unexposed cats and 91% (21/23) progressive infections, but no regressive (0/23) or abortive (0/32) infections. In the second cohort, v-RetroFel®Ag/Ab correctly identified 94% (148/158) FeLV-unexposed cats and 100% (4/4) progressive infections, but no regressive (0/2) and only 17% (1/6) abortive infections. There was test agreement between v-RetroFel®Ab and the p15E laboratory ELISA in 58.9% of samples. As a secondary outcome of this study, the sensitivity and specificity of v-RetroFel®FIV testing in cohort 1 were 94.7% (18/19) and 98.3% (178/181), and in cohort 2, 30.0% (3/10) and 100.0% (160/160), respectively. Prior history of FIV vaccination did not produce any false-positive FIV results. In conclusion, v-RetroFel®Ag/Ab (2020-2021 version) was unable to accurately determine different FeLV infection outcomes in the field. Improvements of the test prior to application to field samples are required.

Evaluation of a Point-of-Care Feline Immunodeficiency Virus (FIV) Test Kit (RapidSTATUS™ FIV) to Determine the FIV Status of FIV-Vaccinated and FIV-Unvaccinated Pet Cats in Australia.

Feline immunodeficiency virus (FIV) is a retrovirus that can cause immunosuppression, co-morbidities, and neoplasia in infected cats, and is commonly tested for in veterinary clinics and animal shelters in Australia. FIV diagnosis using point-of-care (PoC) kits to detect FIV antibodies in Australia is complicated by the commercial availability of an inactivated whole-FIV vaccine. The aim of this study was to determine the accuracy of the RapidSTATUS™ FIV antibody test kit in FIV-vaccinated and FIV-unvaccinated cats in Australia. Plasma from pet cats of known FIV vaccination and FIV infection statuses (n = 361), comprised of 57 FIV-uninfected cats annually vaccinated against FIV, 10 FIV-uninfected cats with lapsed FIV vaccination histories, 259 FIV-unvaccinated/FIV-uninfected cats, and 35 FIV-infected cats, was tested. RapidSTATUS™ FIV testing had sensitivity of 97.1% (34/35) and specificity of 100% (326/326), with an overall accuracy of 99.7% (360/361). Additional testing was undertaken using plasma from FIV-uninfected cats recently administered a primary FIV vaccination course (n = 12) or an annual booster FIV vaccination (n = 10). RapidSTATUS™ FIV was 98.8% (81/82) accurate and 100% (32/32) accurate in cats recently administered primary or annual FIV vaccinations, respectively. The high level of accuracy of RapidSTATUS™ FIV (98.8-100%) therefore establishes this PoC kit as a DIVA (differentiating infected from vaccinated animals) test. RapidSTATUS™ FIV is recommended to aid animal shelters, veterinarians, and researchers in Australia to accurately determine FIV infection status, irrespective of FIV vaccination history.

Clinicopathological and Epidemiological Findings in Pet Cats Naturally Infected with Feline Immunodeficiency Virus (FIV) in Australia.

Feline immunodeficiency virus (FIV) infection in experimentally infected domestic cats produces characteristic clinical manifestations including hematological changes, neurological disease, neoplasia (most notably lymphoma) and lymphopenia-mediated immunodeficiency predisposing cats to a range of secondary infections. Conflicting reports exist, however, with regard to disease associations and survival time in naturally FIV-infected cats. The purpose of this retrospective case−control study was to investigate the effect of natural FIV infection on hematological, blood biochemical and urinalysis parameters and survival time in three cohorts of pet cats in Australia. Cohorts 1 and 2 were recruited from a large veterinary hospital in Melbourne, Victoria (n = 525 and 282), while a third cohort consisted of cats recruited from around Australia as part of a FIV field vaccine efficacy trial (n = 425). FIV-infected cats in cohorts 1, 2 and 3 were found to have 15/37 (41%), 13/39 (33%) and 2/13 (15%) clinicopathological parameters significantly different to FIV-uninfected cats, respectively. Two changes in FIV-infected cats in cohort 1, hypochromia (low hemoglobin) and hyperglobulinemia, were outside the supplied reference intervals and should serve as diagnostic triggers for FIV testing. Kaplan−Meier survival analysis of cats in cohorts 1 and 2 combined did not find any difference between FIV-infected and FIV-uninfected cats, however a confounding factor was a large euthanasia rate within the first 12 months in both groups. Three significant (p < 0.05) spatial clusters of FIV infection were identified in Melbourne. A possible relationship between FIV infection status and socioeconomic disadvantage was discovered, based on three government indices of socioeconomic status (p < 0.001). Until longitudinal field studies are performed in Australia to further investigate the long-term effects of natural FIV infection, Australian veterinarians should consider FIV to be an important infection of pet cats, and recommend measures to prevent FIV infection.

Leptospirosis is an emerging infectious disease of pig-hunting dogs and humans in North Queensland.

BACKGROUND: Leptospirosis is a zoonotic disease with a worldwide distribution, caused by pathogenic serovars in the genus Leptospira. Feral pigs are known carriers of Leptospira species and pig hunting using dogs is a common recreational activity in Queensland, Australia. METHODOLOGY AND PRINCIPAL FINDINGS: This study aimed to determine the seroprevalence of Leptospira spp. serovars in pig-hunting dogs above the Tropic of Capricorn in Queensland and by establishing the geographic distribution, serovars and incidence of human cases of leptospirosis in Queensland, identify potential overlap between human and canine exposure. We also explored the knowledge and risk-taking behaviours of pig-hunting dog owners towards zoonotic diseases. Ninety-eight pig-hunting dogs deemed healthy by physical examination and owned by 41 people from Queensland had serum submitted for Microscopic Agglutination Testing (MAT) to determine antibody titres against Leptospira serovars, while 40/41 dog owners completed a survey on their knowledge of diseases relating to pig hunting. Human leptospirosis cases (n = 330) notified to Queensland Health between 2015-2018 were analysed. Approximately one quarter (23/87; 26%) of unvaccinated pig-hunting dogs were seropositive to Leptospira spp. Although harder to interpret, 8/11 (73%) vaccinated dogs were seropositive to Leptospira spp. Pig hunters may be more likely to contract leptospirosis compared with the general Queensland population, based on responses from surveyed hunters. The highest concentration of human leptospirosis was in the wet tropics region of Far North Queensland. There was little overlap between the serovars dogs were exposed to and those infecting humans. The dominant serovar identified in unvaccinated dogs was Australis (13/23; 57%), with serovar Arborea (36/330; 10.9%) responsible for the highest number of human leptospirosis cases. Topaz was the second most common serovar in both humans and dogs and was previously unrecorded in Australian dogs. Most hunters surveyed used hand washing as a zoonotic disease risk reduction technique. CONCLUSIONS: Leptospirosis is an emerging disease of growing significance. The infection requires a 'one health' approach to understand its epidemiology. With shifting climatic patterns influencing human-animal-environment interactions, ongoing monitoring of diseases like leptospirosis is critical to helping prevent infection of individuals and disease outbreaks.

2020 AAHA/AAFP Feline Vaccination Guidelines.

The guidelines are a consensus report on current recommendations for vaccination of cats of any origin, authored by a Task Force of experts. The guidelines are published simultaneously in the Journal of Feline Medicine and Surgery (volume 22, issue 9, pages 813-830, DOI: 10.1177/1098612X20941784) and the Journal of the American Animal Hospital Association (volume 56, issue 4, pages 249-265, DOI: 10.5326/JAAHA-MS-7123). The guidelines assign approved feline vaccines to core (recommended for all cats) and non-core (recommended based on an individualized risk-benefit assessment) categories. Practitioners can develop individualized vaccination protocols consisting of core vaccines and non-core vaccines based on exposure and susceptibility risk as defined by the patient's life stage, lifestyle, and place of origin and by environmental and epidemiologic factors. An update on feline injection-site sarcomas indicates that occurrence of this sequela remains infrequent and idiosyncratic. Staff education initiatives should enable the veterinary practice team to be proficient in advising clients on proper vaccination practices and compliance. Vaccination is a component of a preventive healthcare plan. The vaccination visit should always include a thorough physical exam and client education dialog that gives the pet owner an understanding of how clinical staff assess disease risk and propose recommendations that help ensure an enduring owner-pet relationship.

Pig-hunting dogs are an at-risk population for canine heartworm (Dirofilaria immitis) infection in eastern Australia.

BACKGROUND: Canine heartworm disease, caused by Dirofilaria immitis, has global veterinary importance. In Australia, the prevalence of canine heartworm infection decreased markedly following the introduction of over-the-counter macrocyclic lactones. We aimed to estimate the prevalence of canine heartworm infection in at-risk populations of dogs in eastern Australia and analyse published prevalence data from Australia. METHODS: In total, 566 dogs from eastern Australia were tested for the presence of D. immitis antigen. Four cohorts were studied: pig-hunting dogs from Queensland (Cohort 1, n = 104), dogs from remote New South Wales (NSW) (Cohort 2, n = 332), urban pets from rural NSW (Cohort 3, n = 45) and ex-racing Greyhounds from Sydney, NSW (Cohort 4, n = 85). Serum samples were screened for D. immitis antigen using a reference laboratory microwell-based assay (DiroChek®) or a point-of-care immunochromatography test kit (Anigen Rapid®). Risk factors associated with the odds of D. immitis antigen seropositivity were identified using binary logistic regression models. Seropositive blood samples were tested for the presence and quantity of D. immitis DNA using a species specific real-time (q)PCR assay. A metanalysis of the Australian canine heartworm literature was conducted. RESULTS: The prevalence of dirofilariasis in pig-hunting dogs from Queensland (Cohort 1) was 12.5% (95% CI: 6.5-18.9%), with a subpopulation of dogs from Central Queensland having a prevalence of 21% (95% CI: 12.3-33.4%). Age was significantly associated with D. immitis antigen seropositivity (increased risk with increased age). The odds of being > 5 years versus ≤ 5 years was 3.7-times (95% CI: 1.1-12.5) greater in antigen positive versus antigen negative dogs. No D. immitis antigen positive dogs were detected in dogs from NSW (Cohorts 2-4). The Australian canine heartworm disease literature includes 98 peer-reviewed publications (1901-2019) with 30 studies reporting on D. immitis prevalence in dogs. Throughout the publication peak period (1980s), the primary antemortem diagnostic test was detection of microfilariae. CONCLUSIONS: Canine heartworm infection in dogs used for pig hunting is a previously unexplored topic in Australia. Pig-hunting dogs are infected with canine heartworm in Queensland, Australia, placing pet dogs and cats at increased risk of infection.

Felis Catus Gammaherpesvirus 1 DNAemia in Whole Blood from Therapeutically Immunosuppressed or Retrovirus-Infected Cats.

Gammaherpesviruses are major co-pathogens of human immunodeficiency virus (HIV) infection, making the interactions between feline immunodeficiency virus (FIV) and Felis catus gammaherpesvirus 1 (FcaGHV1) pertinent to both human and veterinary medical research. FIV-infected cats are at increased risk of FcaGHV1 DNAemia and consistently harbor higher FcaGHV1 loads than FIV-uninfected cats. Whether immune deficiencies unrelated to FIV are associated with similar risks is unknown. Using whole blood FcaGHV1 qPCR, we found no difference in the frequency of DNAemia or DNA load in therapeutically immunosuppressed (P1, n = 18) or feline leukemia virus (FeLV)-infected (P2, n = 57) patients compared with age- and sex-matched controls (C1, n = 58; C2, n = 57). In contrast, FIV/FeLV-co-infected cats (P3, n = 5) were at increased risk of FcaGHV1 DNAemia compared to retrovirus uninfected controls (C3, n = 39; p = 0.0068), and had a higher median FcaGHV1 DNA load, although the latter was not significant. FIV/FeLV-co-infected cats (P3) had a similar frequency of FcaGHV1 DNAemia reported compared to FIV-infected controls (C4). In conclusion, we found no evidence that cats with therapeutic immunosuppression or FeLV infection were at greater risk of FcaGHV1 DNAemia or had higher FcaGHV1 DNA load in whole blood. The risk of DNAemia in FIV/FeLV-co-infected cats was similar to that documented previously in cats infected with FIV alone.

Searching for Lyme borreliosis in Australia: results of a canine sentinel study.

BACKGROUND: Lyme borreliosis is a common tick-borne disease of the northern hemisphere that is caused by bacterial spirochaetes of the Borrelia burgdorferi (sensu lato) (Bbsl) complex. To date, there has been no convincing evidence for locally-acquired Lyme borreliosis on the Australian continent and there is currently a national debate concerning the nature and distributions of zoonotic tick-transmitted infectious disease in Australia. In studies conducted in Europe and the United States, dogs have been used as sentinels for tick-associated illness in people since they readily contact ticks that may harbour zoonotic pathogens. Applying this principle, we used a combination of serological assays to test dogs living in tick 'hot spots' and exposed to the Australian paralysis tick, Ixodes holocyclus, for evidence of exposure to B. burgdorferi (s.l.) antigens and other vector-borne pathogens. RESULTS: Altogether, 555 dogs from four demographic groups were recruited into this study. One dog had evidence of exposure to Anaplasma spp. but no other dog was positive in screening tests. A total of 122 dogs (22.0%) had a kinetic ELISA (KELA) unit value > 100, and one dog with a high titre (399.9 KELA units) had been vaccinated against B. burgdorferi (sensu stricto) before travelling to Australia. Older dogs and those with a history of tick paralysis were significantly more likely to have a KELA unit value > 100. Line immunoassay analysis revealed moderate-to-weak (equivocal) bands in 27 (4.9%) dogs. CONCLUSIONS: Except for a single dog presumed to have been exposed to Anaplasma platys, infection with Anaplasma spp. B. burgdorferi (s.l.), Ehrlichia spp., and Dirofilaria immitis, was not detected in the cohort of Australian dogs evaluated in this study. These results provide further evidence that Lyme borreliosis does not exist in Australia but that cross-reacting antibodies (false positive results) are common and may be caused by the transmission of other tick-associated organisms.

Comparison of three feline leukaemia virus (FeLV) point-of-care antigen test kits using blood and saliva.

Feline leukaemia virus (FeLV) can be a challenging infection to diagnose due to a complex feline host-pathogen relationship and occasionally unreliable test results. This study compared the accuracy of three point-of-care (PoC) FeLV p27 antigen test kits commonly used in Australia and available commercially worldwide (SNAP FIV/FeLV Combo, Witness FeLV/FIV and Anigen Rapid FIV/FeLV), using detection of FeLV provirus by an in-house real-time polymerase chain reaction (qPCR) assay as the diagnostic gold standard. Blood (n=563) and saliva (n=419) specimens were collected from a population of cats determined to include 491 FeLV-uninfected and 72 FeLV-infected individuals (45 progressive infections [p27 and qPCR positive], 27 regressive infections [p27 negative, qPCR positive]). Sensitivity and specificity using whole blood was 63% and 94% for SNAP Combo, 57% and 98% for Witness, and 57% and 98% for Anigen Rapid, respectively. SNAP Combo had a significantly lower specificity using blood compared to the other two kits (P=0.004 compared to Witness, P=0.007 compared to Anigen Rapid). False-positive test results occurred with all three kits using blood, and although using any two kits in parallel increased specificity, no combination of kits completely eliminated the occurrence of false-positive results. We therefore recommend FeLV proviral PCR testing for any cat that tests positive with a PoC FeLV antigen kit, as well as for any cat that has been potentially exposed to FeLV but tests negative with a FeLV antigen kit, before final assignment of FeLV status can be made with confidence. For saliva testing, sensitivity and specificity was 54% and 100%, respectively, for all three test kits. The reduced sensitivity of saliva testing compared to blood testing, although not statistically significant, suggests saliva testing with the current generation of PoC FeLV antigen kits is unsuitable for screening large populations of cats, such as in shelters.

Duration of antibody response following vaccination against feline immunodeficiency virus.

Objectives Recently, two point-of-care (PoC) feline immunodeficiency virus (FIV) antibody test kits (Witness and Anigen Rapid) were reported as being able to differentiate FIV-vaccinated from FIV-infected cats at a single time point, irrespective of the gap between testing and last vaccination (0-7 years). The aim of the current study was to investigate systematically anti-FIV antibody production over time in response to the recommended primary FIV vaccination series. Methods First, residual plasma from the original study was tested using a laboratory-based ELISA to determine whether negative results with PoC testing were due to reduced as opposed to absent antibodies to gp40. Second, a prospective study was performed using immunologically naive client-owned kittens and cats given a primary FIV vaccination series using a commercially available inactivated whole cell/inactivated whole virus vaccine (Fel-O-Vax FIV, three subcutaneous injections at 4 week intervals) and tested systematically (up to 11 times) over 6 months, using four commercially available PoC FIV antibody kits (SNAP FIV/FeLV Combo [detects antibodies to p15/p24], Witness FeLV/FIV [gp40], Anigen Rapid FIV/FeLV [p24/gp40] and VetScan FeLV/FIV Rapid [p24]). Results The laboratory-based ELISA showed cats from the original study vaccinated within the previous 0-15 months had detectable levels of antibodies to gp40, despite testing negative with two kits that use gp40 as a capture antigen (Witness and Anigen Rapid kits). The prospective study showed that antibody testing with SNAP Combo and VetScan Rapid was positive in all cats 2 weeks after the second primary FIV vaccination, and remained positive for the duration of the study (12/12 and 10/12 cats positive, respectively). Antibody testing with Witness and Anigen Rapid was also positive in a high proportion of cats 2 weeks after the second primary FIV vaccination (8/12 and 7/12, respectively), but antibody levels declined below the level of detection in most cats (10/12) by 1 month after the third (final) primary FIV vaccination. All cats tested negative using Witness and Anigen Rapid 6 months after the third primary FIV vaccination. Conclusions and relevance This study has shown that a primary course of FIV vaccination does not interfere with FIV antibody testing in cats using Witness and Anigen Rapid, provided primary vaccination has not occurred within the previous 6 months. Consequently, Witness and Anigen Rapid antibody test kits can be used reliably to determine FIV infection status at the time of annual booster FIV vaccination to help detect 'vaccine breakthroughs' and in cats that have not received a primary course of FIV vaccination within the preceding 6 months. The duration of antibody response following annual booster FIV vaccination and the resulting effect on antibody testing using PoC kits needs to be determined by further research. The mechanism(s) for the variation in FIV antibody test kit performance remains unclear.

Diagnosing feline immunodeficiency virus (FIV) infection in FIV-vaccinated and FIV-unvaccinated cats using saliva.

We recently showed that two immunochromatography point-of-care FIV antibody test kits (Witness FeLV/FIV and Anigen Rapid FIV/FeLV) were able to correctly assign FIV infection status, irrespective of FIV vaccination history, using whole blood as the diagnostic specimen. A third FIV antibody test kit, SNAP FIV/FeLV Combo (an enzyme-linked immunosorbent assay [ELISA]), was unable to differentiate antibodies produced in response to FIV vaccination from those incited by FIV infection. The aim of this study was to determine if saliva is a suitable diagnostic specimen using the same well characterized feline cohort. FIV infection status of these cats had been determined previously using a combination of serology, polymerase chain reaction (PCR) testing and virus isolation. This final assignment was then compared to results obtained using saliva as the diagnostic specimen utilizing the same three point-of-care FIV antibody test kits and commercially available PCR assay (FIV RealPCR). In a population of cats where one third (117/356; 33%) were FIV-vaccinated, both immunochromatography test kits accurately diagnosed FIV infection using saliva via a centrifugation method, irrespective of FIV vaccination history. For FIV diagnosis using saliva, the specificity of Anigen Rapid FIV/FeLV and Witness FeLV/FIV was 100%, while the sensitivity of these kits was 96% and 92% respectively. SNAP FIV/FeLV Combo respectively. SNAP FIV/FeLV Combo had a specificity of 98% and sensitivity of 44%, while FIV RealPCR testing had a specificity of 100% and sensitivity of 72% using saliva. A revised direct method of saliva testing was trialed on a subset of FIV-infected cats (n=14), resulting in 14, 7 and 0 FIV positive results using Anigen Rapid FIV/FeLV, Witness FeLV/FIV and SNAP FIV/FeLV Combo, respectively. These results demonstrate that saliva can be used to diagnose FIV infection, irrespective of FIV vaccination history, using either a centrifugation method (Anigen Rapid FIV/FeLV and Witness FeLV/FIV) or a direct method (Anigen Rapid FIV/FeLV). Collection of a saliva specimen therefore provides an acceptable alternative to venipuncture (i) in fractious cats where saliva may be easier to obtain than whole blood, (ii) in settings when a veterinarian or trained technician is unavailable to collect blood and (iii) in shelters where FIV testing is undertaken prior to adoption but additional blood testing is not required.

Seroprevalence of feline immunodeficiency virus and feline leukaemia virus in Australia: risk factors for infection and geographical influences (2011-2013).

OBJECTIVES: Our aim was to: (i) determine the current seroprevalence of feline immunodeficiency virus (FIV) and feline leukaemia virus (FeLV) in three large cohorts of cats from Australia; and (ii) investigate potential risk factors for retroviral infection. METHODS: Cohort 1 (n = 2151 for FIV, n = 2241 for FeLV) consisted of cats surrendered to a shelter on the west coast of Australia (Perth, Western Australia [WA]). Cohort 2 (n = 2083 for FIV, n = 2032 for FeLV) consisted of client-owned cats with outdoor access recruited from around Australia through participating veterinary clinics. Cohort 3 (n = 169 for FIV, n = 166 for FeLV) consisted of cats presenting to Murdoch University Veterinary Hospital for a variety of reasons. Fresh whole blood was collected and tested using a commercially available point-of-care lateral flow ELISA kit that detects p27 FeLV antigen and antibodies to FIV antigens (p15 and p24) (cohorts 1 and 2), or one of two lateral flow immunochromatography kits that detect p27 antigen and antibodies to FIV antigen (p24 and/or gp40) (cohort 3). Data recorded for cats in cohort 2 included signalment, presenting complaint and postcode, allowing investigation of risk factors for FIV or FeLV infection, as well as potential geographical 'hot spots' for infection. RESULTS: The seroprevalence of FIV was 6% (cohort 1), 15% (cohort 2) and 14% (cohort 3), while the seroprevalence of FeLV was 1%, 2% and 4% in the same respective cohorts. Risk factors for FIV infection among cats in cohort 2 included age (>3 years), sex (male), neutering status (entire males) and location (WA had a significantly higher FIV seroprevalence compared with the Australian Capital Territory, New South Wales and Victoria). Risk factors for FeLV infection among cats in cohort 2 included health status ('sick') and location (WA cats were approximately three times more likely to be FeLV-infected compared with the rest of Australia). No geographical hot spots of FIV infection were identified. CONCLUSIONS AND RELEVANCE: Both FIV and FeLV remain important infections among Australian cats. WA has a higher seroprevalence of both feline retroviruses compared with the rest of Australia, which has been noted in previous studies. A lower neutering rate for client-owned male cats is likely responsible for the higher seroprevalence of FIV infection in WA cats, while the reason for the higher seroprevalence of FeLV in WA cats is currently unknown.

Determining the feline immunodeficiency virus (FIV) status of FIV-vaccinated cats using point-of-care antibody kits.

This study challenges the commonly held view that the feline immunodeficiency virus (FIV) infection status of FIV-vaccinated cats cannot be determined using point-of-care antibody test kits due to indistinguishable antibody production in FIV-vaccinated and naturally FIV-infected cats. The performance of three commercially available point-of-care antibody test kits was compared in a mixed population of FIV-vaccinated (n=119) and FIV-unvaccinated (n=239) cats in Australia. FIV infection status was assigned by considering the results of all antibody kits in concert with results from a commercially available PCR assay (FIV RealPCR™). Two lateral flow immunochromatography test kits (Witness FeLV/FIV; Anigen Rapid FIV/FeLV) had excellent overall sensitivity (100%; 100%) and specificity (98%; 100%) and could discern the true FIV infection status of cats, irrespective of FIV vaccination history. The lateral flow ELISA test kit (SNAP FIV/FeLV Combo) could not determine if antibodies detected were due to previous FIV vaccination, natural FIV infection, or both. The sensitivity and specificity of FIV RealPCR™ for detection of viral and proviral nucleic acid was 92% and 99%, respectively. These results will potentially change the way veterinary practitioners screen for FIV in jurisdictions where FIV vaccination is practiced, especially in shelter scenarios where the feasibility of mass screening is impacted by the cost of testing.

Differences in the faecal microbiome of non-diarrhoeic clinically healthy dogs and cats associated with Giardia duodenalis infection: impact of hookworms and coccidia.

The protozoan parasite Giardia duodenalis causes a waterborne diarrhoeal disease in animals and humans, yet many Giardia-infected hosts remain asymptomatic. Mixed parasite infections are common in both animals and humans with unknown consequences for Giardia or other parasites. We compared the composition and diversity of bacterial communities from 40 dogs, including free-roaming dogs, and 21 surrendered cats from Australia. The dog cohort included 17 (42.5%) dogs positive for Giardia and 13 (32.5%) dogs positive for dog hookworm (Ancylostoma caninum). The cat samples included eight positive for Giardia and eight positive for Cystoisospora. The V4 region of 16S rRNA was sequenced at an average of 36,383 high quality sequences (>200 bp) per sample using the Ion Torrent PGM platform. In dogs we found significant (P<0.05, AnoSim) difference between the Giardia-positive and -negative groups when evaluating bacterial genera. No such difference was demonstrated between Ancylostoma-positive and -negative dogs. However, there was a modest but not significant separation of the Giardia-negative and -positive dogs (P=0.09, UniFrac) using principal coordinate analysis. Removal of dogs with hookworms further separated Giardia-positive and -negative groupings (P=0.06, UniFrac). In cats, the presence of Giardia was not associated with a significant difference based on bacterial genera (P>0.05, AnoSim). Cystoisospora-positive cats, however, exhibited significantly different profiles from Cystoisospora-negative cats (P=0.02, AnoSim) and UniFrac showed significant separation of Cystoisospora-positive and -negative samples (P<0.01). The results suggest that in clinically heathy dogs and cats, helminths and protozoa are associated with different microbiomes and possibly variable gut microbiota functions. Understanding the association of parasites and microbiomes has important consequences for the administration of antiparasitic drugs in animals and humans.