Modulation of the host Th1 immune response in pigeon protozoal encephalitis caused by Sarcocystis calchasi

Pigeon protozoal encephalitis (PPE) is an emerging central-nervous disease of domestic pigeons (Columba livia f. domestica) reported in Germany and the United States. It is caused by the apicomplexan parasite Sarcocystis calchasi which is transmitted by Accipter hawks. In contrast to other members of the Apicomplexa such as Toxoplasma and Plasmodium, the knowledge about the pathophysiology and host manipulation of Sarcocystis is scarce and almost nothing is known about PPE. Here we show by mRNA expression profiling a significant down-modulation of the interleukin (IL)-12/IL-18/interferon (IFN)-γ axis in the brains of experimentally infected pigeons during the schizogonic phase of disease. Concomitantly, no cellular immune response was observed in histopathology while immunohistochemistry and nested PCR detected S. calchasi. In contrast, in the late central-nervous phase, IFN-γ and tumor necrosis factor (TNF) α-related cytokines were significantly up-modulated, which correlated with a prominent MHC-II protein expression in areas of mononuclear cell infiltration and necrosis. The mononuclear cell fraction was mainly composed of T-lymphocytes, fewer macrophages and B-lymphocytes. Surprisingly, the severity and composition of the immune cell response appears unrelated to the infectious dose, although the severity and onset of the central nervous signs clearly was dose-dependent. We identified no or only very few tissue cysts by immunohistochemistry in pigeons with severe encephalitis of which one pigeon repeatedly remained negative by PCR despite severe lesions. Taken together, these observations may suggest an immune evasion strategy of S. calchasi during the early phase and a delayed-type hypersensitivity reaction as cause of the extensive cerebral lesions during the late neurological phase of disease

Repatriation of an old fish host as an opportunity for myxozoan parasite diversity: The example of the allis shad, Alosa alosa (Clupeidae), in the Rhine

Background: Wildlife repatriation represents an opportunity for parasites. Reintroduced hosts are expected to accumulate generalist parasites via spillover from reservoir hosts, whereas colonization with specialist parasites is unlikely. We address the question of how myxozoan parasites, which are characterized by a complex life-cycle alternating between annelids and fish, can invade a reintroduced fish species and determine the impact of a de novo invasion on parasite diversity. We investigated the case of the anadromous allis shad, Alosa alosa (L.), which was reintroduced into the Rhine approximately 70 years after its extinction in this river system. Methods: We studied parasites belonging to the Myxozoa (Cnidaria) in 196 allis shad from (i) established populations in the French rivers Garonne and Dordogne and (ii) repatriated populations in the Rhine, by screening the first adults returning to spawn in 2014. Following microscopical detection of myxozoan infections general myxozoan primers were used for SSU rDNA amplification and sequencing. Phylogenetic analyses were performed and cloned sequences were analyzed from individuals of different water sources to better understand the diversity and population structure of myxozoan isolates in long-term coexisting vs recently established host-parasite systems. Results: We describe Hoferellus alosae n. sp. from the renal tubules of allis shad by use of morphological and molecular methods. A species-specific PCR assay determined that the prevalence of H. alosae n. sp. is 100 % in sexually mature fish in the Garonne/Dordogne river systems and 22 % in the first mature shad returning to spawn in the Rhine. The diversity of SSU rDNA clones of the parasite was up to four times higher in the Rhine and lacked a site-specific signature of SNPs such as in the French rivers. A second myxozoan, Ortholinea sp., was detected exclusively in allis shad from the Rhine. Conclusions: Our data demonstrate that the de novo establishment of myxozoan infections in rivers is slow but of great genetic diversity, which can only be explained by the introduction of spores from genetically diverse sources, predominantly via straying fish or by migratory piscivorous birds. Long-term studies will show if and how the high diversity of a de novo introduction of host-specific myxozoans succeeds into the establishment of a local successful strain in vertebrate and invertebrate hosts

Absence of Mycoplasma spp. in nightingales (Luscinia megarhynchos) and blue (Cyanistes caeruleus) and great tits (Parus major) in Germany and its potential implication for evolutionary studies in birds

Mycoplasma spp. are important pathogens in poultry and cause high economic losses for poultry industry worldwide. In other bird species (e.g. white storks, birds of prey, and several waterfowl species), Mycoplasma spp. are regularly found in healthy individuals, hence, considered apathogenic or part of the microbiota of the upper respiratory tract. However, as Mycoplasma spp. are absent in healthy individuals of some wild bird species, they might play a role as respiratory pathogen in these bird species, e.g. Mycoplasma gallisepticum in house finches. The knowledge on the occurrence of Mycoplasma spp. in wild birds is limited. To evaluate the relevance of Mycoplasma spp. in free-ranging nightingales and tits, 172 wild caught birds were screened for the presence of mycoplasmas. The birds were sampled via choanal swabs and examined via molecular methods (n = 172) and, when possible, via culture (n = 142). The Mycoplasma sp. was determined by sequencing the 16S rRNA gene and 16S-23S Intergenic Transcribed Spacer Region. All birds were tested negative for mycoplasmas via PCR and/or mycoplasmal culture. Hence, free-ranging nightingales and tits do not show any mycoplasma in their microbial flora of the respiratory tract. Therefore, these songbird species may suffer from clinical mycoplasmosis when being infected. We hypothesize that birds relying on their vocal ability for reproduction have excluded mycoplasmas from their respiratory flora compared to other bird species

Detection of hepatitis E virus in wild boars of rural and urban regions in Germany and whole genome characterization of an endemic strain

<p>Abstract</p> <p>Background</p> <p>Hepatitis E is an increasingly diagnosed human disease in Central Europe. Besides domestic pigs, in which hepatitis E virus (HEV) infection is highly prevalent, wild boars have been identified as a possible source of human infection. In order to assess the distribution of HEV in the wild boar population of Germany, we tested liver samples originating from different geographical regions for the presence of the HEV genome and compared the detected sequences to animal and human HEV strains.</p> <p>Results</p> <p>A total of 148 wild boar liver samples were tested using real-time RT-PCR resulting in an average HEV detection rate of 14.9% (95% CI 9.6–21.6). HEV was detected in all age classes and all geographical regions. However, the prevalence of HEV infection was significantly higher in rural as compared to urban regions (p < 0.001). Sequencing of the PCR products indicated a high degree of heterogenicity of the detected viruses within genotype 3 and a grouping according to their geographical origin. The whole genome sequence of an HEV isolate (wbGER27) detected in many wild boars in the federal state of Brandenburg was determined. It belongs to genotype 3i and shows 97.9% nucleotide sequence identity to a partial sequence derived from a human hepatitis E patient from Germany.</p> <p>Conclusion</p> <p>The results indicate that wild boars have to be considered as a reservoir for HEV in Germany and that a risk of HEV transmission to humans is present in rural as well as urban regions.</p

Modulation of the host Th1 immune response in pigeon protozoal encephalitis caused by Sarcocystis calchasi

Pigeon protozoal encephalitis (PPE) is an emerging central-nervous disease of domestic pigeons (Columba livia f. domestica) reported in Germany and the United States. It is caused by the apicomplexan parasite Sarcocystis calchasi which is transmitted by Accipter hawks. In contrast to other members of the Apicomplexa such as Toxoplasma and Plasmodium, the knowledge about the pathophysiology and host manipulation of Sarcocystis is scarce and almost nothing is known about PPE. Here we show by mRNA expression profiling a significant down-modulation of the interleukin (IL)-12/IL-18/interferon (IFN)-γ axis in the brains of experimentally infected pigeons during the schizogonic phase of disease. Concomitantly, no cellular immune response was observed in histopathology while immunohistochemistry and nested PCR detected S. calchasi. In contrast, in the late central-nervous phase, IFN-γ and tumor necrosis factor (TNF) α-related cytokines were significantly up-modulated, which correlated with a prominent MHC-II protein expression in areas of mononuclear cell infiltration and necrosis. The mononuclear cell fraction was mainly composed of T-lymphocytes, fewer macrophages and B-lymphocytes. Surprisingly, the severity and composition of the immune cell response appears unrelated to the infectious dose, although the severity and onset of the central nervous signs clearly was dose-dependent. We identified no or only very few tissue cysts by immunohistochemistry in pigeons with severe encephalitis of which one pigeon repeatedly remained negative by PCR despite severe lesions. Taken together, these observations may suggest an immune evasion strategy of S. calchasi during the early phase and a delayed-type hypersensitivity reaction as cause of the extensive cerebral lesions during the late neurological phase of disease

Accipiter hawks and Common Woodpigeon in Germany

The apicomplexan parasite Sarcocystis calchasi (S. calchasi) triggers pigeon protozoal encephalitis, a neurologic disease in columbids. Accipiter hawks have been identified as the final host, and Columbidae and Psittaciformes as intermediate hosts. In this study, 368 free-ranging Accipiter hawks and 647 free-ranging common woodpigeons were sampled in a country-wide study in order to identify the prevalence of S. calchasi in these populations. A semi-nested PCR specific for S. calchasi tested positive in 7.3% (4.9–10.5) of submitted samples from Accipiter hawks. Juvenile Accipiter hawks (13.7%; 7.7–22.0) had a significantly higher infection rate with S. calchasi than adult Accipiter hawks (5.8%; 2.7–9.3). The prevalence of S. calchasi in common woodpigeons was 3.3% (5.4–9.7). Positive pigeons were identified in 14/16 federal states, and a region-dependency was detected, with higher rates of infection in the eastern parts of Germany. The results of this study suggest that the common woodpigeon is a natural reservoir for S. calchasi. In a study of one region for four consecutive years, an increase in prevalence was not detected. Findings indicate that the parasite is not newly introduced to Germany, but rather long established. The prevalence suggests that there is a substantial risk of S. calchasi infections in other free-ranging as well as captive host species

Susceptibility of adult pigeons and hybrid falcons to experimental aspergillosis

Aspergillosis caused by Aspergillus fumigatus seems to be more prevalent in some avian species than in others. We compared the development of aspergillosis in 8-month-old Gyr-Saker hybrid falcons and 8-month-old pigeons after a single intratracheal inoculation of different dosages of A. fumigatus conidia (107, 105 and 103). Clinical signs, including vomiting, discoloration of the urates, loss of appetite and dyspnoea, were observed in four out of five falcons and in four out of five pigeons inoculated with 107 A. fumigatus conidia. Necropsy revealed the presence of granulomas in the air sacs and/or lungs in four out of five falcons and in four out of five pigeons in the high dosage group. A. fumigatus was isolated from these granulomas in three falcons and in three pigeons. The presence of fungal hyphae was detected with Periodic acid Shiff reagent staining in three out of five falcons and in three out of five pigeons in the high dosage group. Avian respiratory macrophages were clearly present in and around the fungal granulomas. In the other dosage groups, no granulomas, positive A. fumigatus cultures or fungal hyphae were present, except for one falcon in the middle dosage group in which a sterile granuloma without fungal hyphae was noticed. In conclusion, the study shows that adult falcons and pigeons are susceptible to aspergillosis after inoculation of a single dose of conidia intratracheally