Koala retrovirus (KoRV) infects the majority of Australia’s koalas (Phascolarctos cinereus) and has been linked to several life-threatening diseases such as lymphoma and leukemia, as well as Chlamydia and thus poses a threat to the continued survival of this species. While quarantine and antiretroviral drug treatment are possible control measures, they are impractical, leaving vaccination as the only realistic option. In this study, we examined the effect of a recombinant envelope protein-based anti-KoRV vaccine in two groups of South Australian koalas: KoRV infected or KoRV free. We report a successful vaccination response in the koalas with no vaccine-associated side effects. The vaccine induced a significant humoral immune response as well as the production of neutralizing antibodies in both groups of koalas. We also identified B-cell epitopes that were differentially recognized in KoRV-infected versus KoRV-free koalas following vaccination. Importantly, we also showed that vaccination had a therapeutic effect on koalas infected exogenously with KoRV by reducing their circulating viral load. Together, this study highlights the possibility of successfully developing a vaccine against KoRV infection in koalas.
Koala retrovirus (KoRV) displays features of both an endogenous and exogenous virus and is linked to neoplasia and immunosuppression in koalas. This study explores the apparent differences in the nature and impact of KoRV infection between geographically and genetically separated "northern" and "southern" koala populations, by investigating the disease status, completeness of the KoRV genome and the proviral (DNA) and viral (RNA) loads of 71 northern and 97 southern koalas. All northern animals were positive for all KoRV genes (gag, pro-pol and env) in both DNA and RNA forms, whereas many southern animals were missing one or more KoRV genes. There was a significant relationship between the completeness of the KoRV genome and clinical status in this population. The proviral and viral loads of the northern population were significantly higher than those of the southern population (P < 0.0001), and many provirus-positive southern animals failed to express any detectable KoRV RNA. Across both populations there was a positive association between proviral load and neoplasia (p = 0.009). Potential reasons for the differences in the nature of KoRV infection between the two populations are discussed. Koala retrovirus (KoRV) was first identified in tissues from a leukemic koala in 1988 1 and the full genome sequence was published in 2000 2. This is a unique endogenous retrovirus which is also capable of being transmitted horizontally 3. It is thought that KoRV plays a role in the pathogenesis of lymphoid neoplasia as well as causing immunosuppression, making koalas more susceptible to developing overt chlamydial disease 4 , which is a major threat to koala populations. However while there appears to be a strong association between KoRV infection and these diseases in koalas, there is currently little if any published data proving a causal link 3,5,6. Anecdotally, there are differences in disease prevalence between koala populations in northern and southern Australia. The prevalence of lymphoid neoplasia is high in northern koalas 7 compared to southern koalas where lymphosarcoma has only recently been reported 8. The prevalence of Chlamydia pecorum infection and overt chlamydial disease is also high in northern koalas in comparison to southern populations, with chlamydial disease reported in 52% of more than 20,000 koala admissions to wildlife hospitals in South East Queensland (SE QLD) 9. In contrast, a recent necropsy study in South Australia (SA) reported only 21% of koalas with clinically overt disease 10. Conversely, SA kocalas have a higher prevalence of oxalate nephrosis than QLD koalas, with 55% of SA koalas demonstrating renal dysfunction due to oxalate nephrosis 11. These differences in disease prevalence may be the result of genetic differences between northern southern koala populations 12. Animals in the southern part of the range (the states of Victoria, South Australia [SA] and southern New South Wales [NSW]) have undergone a severe genetic bottleneck as a result of hunting pressures in the late 1800's. Thes...
Background Mycoplasma mastitis is increasingly posing significant impact on dairy industry. Although the effects of major conventional mastitis pathogens on milk components has been widely addressed in the literature, limited data on the effects of different Mycoplasma and Acholeplasma spp. on milk quality and quantity is available. The aim of this study was to determine the casual relationship of Mycoplasma spp. and A. laidlawii to mastitis and compare them to subclinical mastitis caused by conventional mastitis pathogens from a single dairy herd in South Australia; Mycoplasma spp. and A. laidlawii were detected using PCR applied directly to milk samples. The herd had mastitis problem with high somatic cell count and low response rate to conventional antimicrobial therapy. A total of 288 cow-level milk samples were collected aseptically and used in this study.ResultsConventional culture showed a predominance of coagulase-negative staphylococci, followed by coagulase-positive staphylococci, Streptococcus spp., Enterococcus spp., E. coli, and Klebsiella spp. PCR results showed a high prevalence of mycoplasmas (76.7%), including A. laidlawii (10.8%), M. bovis (6.2%), M. bovirhinis (5.6%), M. arginini (2%), and (52.1%) of cows were co-infected with two or more Mycoplasma and Acholeplasma species. Mycoplasma co-infection significantly increased somatic cell counts (SCC) similar to conventional mastitis pathogens and compared to non-infected cows with 389.3, 550.3 and 67.3 respectively; and decreased the milk yield with 29.0, 29.9 and 34.4 l, respectively. Mycoplasma co-infection caused significant increase in protein percentage, and significant decrease in fat percentage and total milk solids, similar to other conventional mastitis pathogens. In contrast, changes in milk composition and yield caused by various individual Mycoplasma species were non-significant.Conclusions Mycoplasma mastitis had on-farm economic consequences similar to common conventional mastitis pathogens. Results of our study indicate that co-infection Mycoplasma mastitis caused similar effect on milk composition to other mastitis pathogens and we hope these findings raise the awareness of the importance of their detection on routine diagnostic panels.
Available avian influenza (AIV) serological diagnostic tests cannot distinguish vaccinated from naturally infected birds. Differentiation of vaccinated from infected animals (DIVA) is currently advocated as a means of achieving the full control of H5N1. In this study, for the first time, recombinant ectodomain of M2 protein (M2e) of avian influenza virus (H5N1 strain) was used for the DIVA serology test. M2e was cloned into pMAL-P4X vector and expressed in E. coli cells. We used Western blot to recognize the expressed M2e-MBP protein by chicken antisera produced against live H5N1 virus. Also, the specificity of M2e-MBP protein was compared to the M2e synthetic peptide via ELISA. In M2e-MBP ELISA, all sera raised against the live avian influenza viruses were positive for M2e antibodies, whereas sera from killed virus vaccination were negative. Furthermore, M2e-MBP ELISA of the field sera obtained from vaccinated and non-vaccinated chickens showed negative results, while challenged vaccinated chickens demonstrated strong positive reactions. H5N1-originated recombinant M2e protein induced broad-spectrum response and successfully reacted with antibodies against other AIV strains such as H5N2, H9N2, H7N7, and H11N6. The application of the recombinant protein instead of synthetic peptide has the advantages of continues access to an inexpensive reagent for performing a large scale screening. Moreover, recombinant proteins provide the possibility of testing the DIVA results with an additional technique such a Western blotting which is not possible in the case of synthetic proteins. All together, the results of the present investigation show that recombinant M2e-MBP can be used as a robust and inexpensive solution for DIVA test.
The long-term survival of the koala is under serious threat from multiple factors, including infectious disease agents such as Chlamydia and koala retrovirus (KoRV). KoRV is present in both exogenous and endogenous forms, depending on the geographical location of the population. In the northern half of Australia, it is present as an endogenous infection in all koalas, making a case for an urgent need to develop a therapeutic vaccine that might prevent KoRV-associated pathologies in these koalas. To this end, we determined the therapeutic effects of vaccinating koalas harbouring endogenous KoRV with a recombinant KoRV Env protein combined with a Tri-adjuvant. We found that vaccination led to a significant increase in circulating anti-KoRV IgG levels, as well as increase in neutralising antibodies. Our study also showed that post-vaccination antibodies were able to recognize epitopes on the Env protein that were unrecognised pre-vaccination, as well as resulting in an increase in the recognition of the previously recognised epitopes. The vaccine also induced antibodies that were cross-reactive against multiple KoRV-subtypes. Finally, we found a complete clearance of KoRV-A in plasma from koalas that had detectable levels of KoRV-A pre-vaccination. Similarly, there was a significant reduction in the expression of KoRV-B viral RNA levels post-vaccination. Collectively, this study showed that koalas harbouring endogenous KoRV can benefit from prophylactic vaccination against KoRV using a recombinant KoRV-A Env protein and that the mechanism of this protection might be through the boosting of natural anti-KoRV antibodies and expanding the breadth of the recognised epitopes.
Canine parvovirus (CPV-2) is an important cause of hemorrhagic enteritis in dogs. In Australia the disease has been associated with CPV-2a and CPV-2b variants. A third more recently emerged variant overseas, CPV-2c, has not been detected in surveys of the Australian dog population. In this study, we report three cases of canine parvoviral enteritis associated with CPV-2c infection; case 1 occurred in an 8-week-old puppy that died following acute hemorrhagic enteritis. Cases 2 and 3 were an 11-month-old female entire Saint Bernard and a 9-month-old male entire Siberian husky, respectively, both which had completed vaccination schedules and presented with vomiting or mild diarrhea only. Full genomic sequencing of parvoviral DNA from cases 1, 2, and 3 revealed greater than 99% homology to known CPV-2c variants and predicted protein sequences from the VP2 region of viral DNA from all three cases identified; glutamic acid residues at the 426 amino acid residue, characteristic of the CPV-2c variant. Veterinary professionals should be aware that CPV-2c is now present in Australia, detected in a puppy and vaccinated young adult dogs in this study. Further characterization of CPV-2c-associated disease and its prevalence in Australian dogs requires additional research.
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