Anna Orłowska

First detection of bluetongue virus serotype 14 in Poland

Here, we present the first detected cases of bluetongue virus (BTV) in native cattle from Poland. The virus was found in animals located near the Polish-Belarusian and Polish-Lithuanian borders. The positive animals were detected through an official epidemiological surveillance program. A combination of type-specific real-time RT-PCR and phylogenetic tests revealed the presence of BTV serotype 14 (BTV-14). This serotype is highly homologous to the vaccine strain and BTV-14 present in Russia, Lithuania, and Spain (from an animal imported from Lithuania). The most probable route of virus introduction to Poland was transmission through midges. All of the cases were subclinical.

The effect of selected molecules influencing the detrimental host immune response on a course of rabies virus infection in a murine model

Rabies is invariably fatal, when post-exposure prophylaxis is administered after the onset of clinical symptoms. In many countries, rabies awareness is very low and the availability of post-exposure prophylaxis, as recommended by WHO guidelines, is very limited or non-existent, probably as a consequence of high cost. Therefore, new concepts for rabies therapy are needed. Innate immune mechanisms involving the production of pro-inflammatory cytokines and chemokines, activated after rabies infection, are thought to be involved in the neuropathogenesis of rabies. These mechanisms can contribute to a detrimental host response to the rabies virus (RABV) infection. The use of inhibitors of cytokines/chemokines are supposed to extend the survival of a sick individual. Inhibitors of TNF-α, IL-6 and MAPKs were used in RABV inoculated mice to define their influence on the survival time of rabid mice. The study demonstrated that all inhibitors extended mice survival, but at different rates. A log-rank test confirmed the statistically significant survival of mice treated with TNF-α (p = .0087) and MAPKs inhibitors (p = .0024). A delay in the time of onset of rabies was also recorded, in mice given TNF-α and MAPKs inhibitors. The highest virus load was found in the spinal cord and the lowest in the cortex, regardless of the experimental group. Significant TNF-α (p ≤ .0001) and IL-6 (p ≤ .0001) gene upregulation was observed in mice, as a consequence of RABV infection. Regarding MAPKs pathways, there was significant upregulation of the caspase 3 (p = .012, p = .0026) and Mcl-1 (p = .0348, p = .0153) genes, whereas significant downregulation of the cytochrome C (p ≤ .0001), Bcl2 (p = .0002, p = .0007) and JNK3 (p = .042) genes. Rabies pathogenesis is multifactorial, involving both virus and host influences on the course of the infection.

Rabies epidemiological situation in Poland in 2009 and 2010

Abstract The paper describes the data concerning rabies in domestic animals and in wildlife as well as in bats in Poland in 2009 and 2010. Analysis of rabies situation was based on species involved and geographical distribution of rabies outbreaks. Favourable decreasing trend in rabies epidemic in 2009 was stopped by the outbreak of rabies in the Malopolska province in 2010. This resulted in dramatic increase in the number of rabies cases. Emergency vaccination in the zone of rabies outbreak with increased number of vaccines per km2 in bordering areas of the province has improved epizootic situation, which returned to the state before the outbreak. To monitor rabies situation a strict supervision of all elements of the ORV and surveillance of rabies is necessary.

The effect of combined drugs therapy on the course of clinical rabies infection in a murine model

Rabies is a fatal disease of all mammals causing almost 60,000 human deaths every year. To date, there is no effective treatment of clinical rabies once the symptoms appear. Here, we describe the promising effect of combination therapy composed of molecules that target replication of the rabies virus (RV) at different stages of life cycle and molecules that inhibit some pathways of the innate host immune response accompanied by a blood-brain barrier opener on the outcome of RV infection. The study reports statistically significant extension of survival of mice treated with the drug cocktail containing T-705, ribavirin, interferon α/β, caspase-1 inhibitor, TNF-α inhibitor, MAPKs inhibitor and HRIG compared to the survival of mice in the virus control group (p = 0.0312). Furthermore, the study points to the significant impact of interferon α/β on the survival of RV-infected mice. We have shown a significant down regulation of pro-inflammatory molecules (caspase-1 and TNF-a) in the CNS in RV-infected mice treated with a combination of drugs including interferon α/β.

Inhibition of caspase-1 prolongs survival of mice infected with rabies virus

Rabies virus infects almost all mammals resulting in lethal disease. To date there is no treatment available for symptomatic rabies and there is an urgent need to develop treatment strategies that would prolong survival, thereby providing a window of opportunity for the host to mount a protective immune response. We hypothesized that both virus and excessive immune response contribute to disease and that interfering with both is necessary to prevent lethal disease. Here, we have inhibited the pro-inflammatory response associated with pyroptosis and showed that inhibition of CASP-1 had a beneficial effect on survival time. Our results confirm that some inflammatory responses may be involved in the pathogenesis of severe disease and the results suggest that effective intervention includes inhibition of virus and host response.

Bokeloh bat lyssavirus isolation in a Natterer’s bat, Poland

In recent years, Bokeloh bat lyssavirus (BBLV), a member of the novel lyssavirus genus Bokeloh bat lyssavirus in the family Rhabdoviridae, has been detected in Germany (five cases) and France (two cases). Here, we report the isolation of BBLV in a Natterers bat (Myotis nattereri) in Poland. The bat brain tested positive for rabies using classical diagnostics tests (FAT and RTCIT) and then subsequently confirmed by molecular techniques. Viral RNA was found in all peripheral organs tested, and the highest viral loads were detected in brain, the salivary gland and bladder. Phylogenetic analysis performed on complete viral genome sequences revealed the closest homology to representatives of BBLV lineage B, isolated previously in southern Germany. This case provides further evidence that BBLV is widespread in Europe.

Combination drug treatment prolongs survival of experimentally infected mice with silver-haired bat rabies virus

Rabies is a lethal disease in humans and animals, killing approximately 60,000 people every year. Currently, there is no treatment available, except post-exposure prophylaxis (PEP) that can be administered whenever exposure to a rabid animal took place. Here we describe the beneficial effects of a combination treatment initiated at day 4 post infection, containing anti-viral drugs and immune modulators in infected mice. Combination therapy resulted in significant increase in survival time (P < 0.05) and significantly lowers viral RNA in the brain and spinal cord (P < 0.05). Furthermore, treatment influenced markers of pyroptosis and apoptosis and early inflammatory response as measured by the levels of TNF-α. Morphological lesions were absent in rabies virus infected mice with few signs of inflammation. However, these were not significant between the different groups.

Diagnostic reliability of different RT-PCR protocols for the detection of bluetongue virus serotype 14 (BTV-14)

Abstract Introduction: The reverse transcription polymerase chain reaction (RT-PCR) is one of the most extensively used methods for identification of animals infected with bluetongue virus (BTV). There are several RT-PCR protocols published and several real-time RT-PCR (rtRT-PCR) commercial kits available on the market. Because Poland faced BTV-14 infection in 2012, different protocols were implemented in the country to confirm the RT-PCR results positive for this virus. The article presents a comparative study of several RT-PCR protocols and discusses their diagnostic reliability and applicability. Material and Methods: Six rtRT-PCR/RT-PCR protocols were compared for the laboratory diagnostic of fourteen BTV-14 isolates circulating in Poland in 2012–2014. Results: All 14 isolates were positive in the protocols of Shaw et al. (18), a commercial LSI NS3 kit, and Eschbaumer et al. (5). Four out of fourteen BTV-14 isolates gave positive results in Hoffmann’s 2 and 6 protocols and none of the 14 isolates yielded positive results in Maan et al. (8) method. Phylogenetic study of a short fragment of 450 nt of BTV segment 2 (258–696 positions) revealed 100% identity within Polish variants and with Russian and Spanish isolates. Conclusion: The paper points to the possible false negative results in the diagnosis of BTV infections depending on the protocol used.