Avian Poxvirus Research Articles

DISTRIBUTION AND PREVALENCE OF MOSQUITO-BORNE DISEASES IN O’AHU ’ELEPAIO

Abstract ABSTRACT The endemic Hawaiian avifauna is one of the most imperiled on earth, and diseases have been one of the most serious causes of species declines. From 1995–2005, we mist-netted and banded 266 endangered O’ahu ’Elepaio (Chasiempis sandwichensis ibidis) from 27 sites, examined them for visible symptoms of avian pox virus (Poxvirus avium), and screened blood samples for avian malaria (Plasmodium relictum). Pox-like lesions and malaria were found in all ’Elepaio populations on O’ahu; no parts of the island were free of these mosquito-borne diseases. Each year, 20% ± 4% of ’Elepaio had active lesions likely caused by pox and an additional 16% ± 4% had deformities and missing toes indicative of healed pox lesions. Prevalence of malaria was 87% over all years combined. Pox prevalence varied among years and was associated with annual rainfall, presumably due to greater abundance of mosquito breeding sites in wet years. Rainfall amounts at least as high as those associated with pox epizootics in 19...

Increasing prevalence of avian poxvirus in Darwin's finches and its effect on male pairing success

Increasing prevalence of avian poxvirus in Darwin's finches and its effect on male pairing success

Phylogenetic Analysis of Avian Poxviruses Among Free-Ranging Birds of Virginia

Polymerase chain reaction was used to amplify a portion of the avian poxvirus core 4b gene of infected free-ranging birds that presented at the Wildlife Center of Virginia during the 2003 and early 2004 years. The species of bird infected were a great blue heron (Ardea herodias), two American crows (Corvus brachyrhyncos), two American robins (Turdus migratorius), two mourning doves (Zenaida macroura), a red-tailed hawk (Buteo jamaicensis), a blue-gray gnatcatcher (Polioptila caerulea), a northern mockingbird (Mimus polyglottos), a house finch (Carpodacus mexicanus), and a northern cardinal (Cardinalis cardinalis). Phylogenetic analysis was performed using the consensus sequences determined for each avian case in Virginia in combination with avian poxvirus core 4b gene sequence from isolates previously described in Europe and that of vaccinia virus. Alignment of DNA sequences identified areas of point mutations and, in the case of a single mourning dove, the incorporation of a triplet of nucleotides. Maximum-likelihood analysis grouped the 2003-2004 Virginia avian poxviruses into a clade distinct from those reported in European free-ranging birds, with the exception of a single case in a mourning dove that clustered within one European clade. The cladogram that resulted from our analysis of the European isolates is in agreement with those previously published. This study identified a distinct clade of avian poxvirus unique from four clades previously described and associated with epornitics in free-ranging birds, where the core 4b gene DNA sequence has been the basis of comparison.

A new LAGE-1 peptide recognized by cytolytic T lymphocytes on HLA-A68 tumors.

Antigens encoded by genes of the LAGE family, including LAGE-1 and NY-ESO-1, are of interest for cancer immunotherapy because they are tumor-specific and shared by tumors of different histological types. Several clinical trials are in progress with NY-ESO-1 peptides, protein, recombinant poxviruses, and dendritic cells pulsed with peptides. In this study, CD8 T lymphocytes from an individual without cancer were stimulated with dendritic cells infected with a recombinant avian poxvirus encoding a complete LAGE-1 protein. A CTL clone was isolated that recognized a new LAGE-1 peptide, ELVRRILSR, which corresponds to position 103-111 of the protein sequence. It is presented by HLA-A6801 molecules. When tumor cells expressing LAGE-1 were transfected with HLA-A68, they were lysed by the CTL clone, indicating that the peptide is processed in tumor cells. These results indicate that the LAGE-1.A68 peptide can be used for antitumoral vaccination. We observed also that specific T cells could be detected in a blood sample with a high sensitivity by using an A68/LAGE-1 fluorescent multimer.

COMBINED INFECTION BY AVIAN POXVIRUS AND COLLYRICLUM FABA IN AN AMERICAN CROW (CORVUS BRACHYRHYNCHOS)

An adult American crow (Corvus brachyrhynchos) from Virginia, USA, was diagnosed with combined infection of avian poxvirus and the skin fluke Collyriclum faba. The flukes and viral inclusions were combined in a large (4 x 4 cm) multilobulated proliferative mass on the ventrum just cranial to the cloaca. The flukes were identified using light microscopy of organisms obtained by antemortem wedge biopsy. Intraepithelial cytoplasmic inclusions consistent with poxvirus infection were seen on histopathologic examination of the mass.

Causes of Mortality in Sea Ducks (Mergini) Necropsied at the USGS-National Wildlife Health Center

A number of factors were identified as causes of mortality in 254 (59%) of 431 sea ducks submitted for necropsy at the USGS-National Wildlife Health Center, Madison, Wisconsin from 1975 until 2003. Bacteria causing large outbreaks of mortality were Pasteurella multocida and Clostridium botulinum Type E. Starvation was responsible for large mortality events as well as sporadic deaths of individuals. Lead toxicity, gunshot and exposure to petroleum were important anthropogenic factors. Other factors that caused mortality were avian pox virus, bacteria (Clostridium botulinum Type C, Riemerella anatipestifer and Clostridium perfringens), fungi (Aspergillus fumigatus and an unidentified fungus), protozoans (unidentified coccidia), nematodes (Eustrongylides spp.), trematodes (Sphaeridiotrema globulus and Schistosoma spp.), acanthocephalans (Polymorphus spp.), predation, cyanide and trauma (probably due to collisions). There were also a number of novel infectious organisms in free-living sea ducks in North America, which were incidental to the death, including avipoxvirus and reovirus, bacteria Mycobacterium avium, protozoans Sarcocystis sp. and nematodes Streptocara sp. Apart from anthropogenic factors, the other important mortality factors listed here have not been studied as possible causes for the decline of sea ducks in North America.

PREVALENCE OF POX-LIKE LESIONS AND MALARIA IN FOREST BIRD COMMUNITIES ON LEEWARD MAUNA LOA VOLCANO, HAWAII

Abstract Introduced avian pox virus and malaria have had devastating impacts on native Hawaiian forest birds, yet little has been published about their prevalence and distribution in forest bird communities outside of windward Hawaii Island. We surveyed native and non-native forest birds for these two diseases at three different elevations on leeward Mauna Loa Volcano at the Kona Forest Unit of Hakalau Forest National Wildlife Refuge. Prevalence of malaria by both serology and microscopy varied by elevation and ranged from 28% at 710 m to 13% at 1830 m. Prevalence of pox-like lesions also varied by altitude, ranging in native species from 10% at 710 m to 2% at 1830 m. Native species at all elevations had the highest prevalence of malarial antibody and pox-like lesions. By contrast, pox-like lesions were not detected in individuals of four non-native species and only 5% of Japanese White-eye (Zosterops japonicus) was positive for malaria. A significantly high proportion of birds with pox-like lesions also ...

Investigation of the morphology of cell clones, derived from the mammalian EBTr cell line and their susceptibility to vaccine avian poxvirus strains FK and Dessau

The ability for replication of vaccine avian pox viral strains FK and Dessau in cell clones, derived from the EBTr cell line, derived from embryonic bovine trachea, was studied. The derived seven cell clones showed different morphological characteristics and diverse sensitivity to both vaccine avian pox viral strains. Hence, the EBTr-derived cell clones could be used for cultivation, as well as for differentiation of vaccine avian pox viral strains. In addition, studies have been undertaken to elucidate the possible use of cultivated strains in these heterologous cell culture system's vaccine avian pox viral strains for biotechnology, as well as for solving problems, related to infection of people with avian viruses.

The impact of vaccines and the future of genetically modified poxvirus vaccines for poultry

The regular use of live or killed vaccines against infectious agents has remarkably improved the efficiency of poultry production. In some cases eradication of disease has been possible when the pathogen is antigenically stable and confined to a certain geographical area. In other instances monovalent or polyvalent live or killed vaccines have been effective in reducing mortality and morbidity. Many conventional vaccines are developed by trial and error and basic information about their genetic make-up is not known. While the poultry industry has benefited from the regular use of conventional vaccines, there is need for a new generation of effective vaccines that require minimal handling of birds during administration. Using molecular techniques, it is possible to identify the genes associated with virulence and protection. In genetically engineered vaccines, genes that encode protective antigens can be expressed in bacterial or viral vectors. In this regard, avianpox virus vectors appear to be promising for the generation of polyvalent vaccines expressing antigens from multiple pathogens.

Differentiation of Avian Poxvirus Strains on the Basis of Nucleotide Sequences of 4b Gene Fragment

Investigations for detection and differentiation of nine avian poxviruses (APVs) were carried out by the use of a polymerase chain reaction (PCR) combined with restriction enzyme analysis (REA) and further nucleotide sequence analysis. With one primer set, which framed a region within the fowl poxvirus 4b core protein gene, we were able to detect APV-specific DNA from 19 tested strains and isolates belonging to five defined Avipoxvirus species and four previously undefined isolated species. PCR results revealed no recognizable differences in size of amplified fragments among the different APVs. REA of PCR products with MseI and EcoRV allowed us to differentiate most of the tested avipox species. Nucleotide sequence analysis of the amplified fragments showed a nucleotide similarity of 72%-100% among the different species. Phylogenetic analysis documented five distinguishable sequence clusters in accordance with results obtained by REA. PCR in combination with REA and sequencing of the amplified fragments is a rapid and effective diagnostic system, and it is a new approach to refine epidemiologic studies of APV infections.

DEMOGRAPHY OF HAWAI‘I ‘ELEPAIO: VARIATION WITH HABITAT DISTURBANCE AND POPULATION DENSITY

From 1994 to 1997, I monitored ‘Elepaio (Chasiempis sandwichensis), a forest bird endemic to Hawaii, in three sites that differed in either degree of habitat disturbance or population density. The goal was to determine whether areas with disturbed forest and reduced population density due to disease were lower-quality habitats that acted as population sinks. Sites where the forest had been disturbed by cattle grazing and timber harvesting contained larger ‘Elepaio territories, supported less dense ‘Elepaio populations, and had a higher proportion of subadult birds in the breeding population and, thus, can be considered lower-quality habitat for ‘Elepaio. However, ‘Elepaio populations in disturbed sites did not have lower survival, reproduction, or site fidelity than a population in denser forest, and they were self-sustaining. Disturbed sites were not population sinks maintained by immigration; they simply had a lower carrying capacity. Annual survival of territory-holders was high and was slightly higher in males (0.86 ± 0.04 [mean ± 1 se]) than in females (0.79 ± 0.05). Fecundity was low, with pairs producing 0.56 ± 0.06 fledglings per year. Site fidelity was high in males (0.98) and females (0.93), and most birds remained on the same territory for the duration of the study or until their presumed death. Population growth was stable at two sites (λ = 0.96 ± 0.03, 1.02 ± 0.04) and increasing (1.12 ± 0.04) at a site recovering from an epizootic of avian poxvirus. Young, subadult-plumaged males defended smaller territories and produced fewer fledglings than older males, but female age did not affect territory size or reproductive success. Recruitment age was younger where population density was lower.

Descriptive study of an avian pox outbreak in wild red-legged partridges (Alectoris rufa) in Spain.

This study describes the dynamics and epidemiology of an outbreak of avian pox in free-living, red-legged partridges (Alectoris rufa) in southern Spain. Between March 2000 and January 2001, 115 free-living, red-legged partridges (70 juveniles, 45 adults) were captured and radio-tagged. This, together with the necropsy of 44 carcasses (10 juveniles, 34 adults) found in the study area, and the inspection of 108 shot birds (74 juveniles, 34 adults) after a hunting drive in October, permitted a close monitoring of the course of the disease. Forty-one per cent of radio-tagged juveniles but none of 45 radio-tagged adults showed pox-like lesions at the time of capture, recapture, or necropsy. At least 40% of the juveniles that survived into the hunting season, but only 2.9% of the adults inspected at the same time, showed lesions suggestive of infection with avian poxvirus. The survival of juveniles during the peak of the outbreak was much lower than that of the adults, but we found no significant differences between the survival probabilities of juvenile partridges with and without pox-like lesions. Nevertheless, some birds may have developed lesions after their capture. The occurrence and course of the disease in a managed area with intense predator control underlines the need for studies on the combined influence of diseases and predators on population dynamics. Also the need for early detection of diseases for the management of game species is emphasized.

Characterization of an avianpox virus isolated from an Andean condor ( Vultur gryphus)

A novel pox virus, condorpox virus (CPV) isolated from the spleen of an Andean condor ( Vultur gryphus) by inoculation of chorioallantoic membranes (CAM) of specific pathogen free (SPF) chicken embryos was compared biologically, antigenically and genetically with fowlpox virus (FPV), the type species of the genus Avipoxvirus. Susceptible chickens inoculated with CPV developed only mild localized lesions but were not protected against subsequent challenge with FPV. Based on Western blotting, in addition to the presence of cross-reacting antigens, distinct differences in antigenic profiles of CPV and FPV were observed. Sequence analysis of a 4.5 kb HindIII fragment of CPV genomic DNA revealed the presence of eight co-linear genes corresponding to FPV open reading frame (ORF)193–198, 201 and 203. Interestingly, reticuloendotheliosis virus (REV) sequences present in the genome of all FPV were absent in CPV. Although, the results of a phylogenic analysis suggested that CPV is a member of the genus Avipoxvirus, its unique antigenic, biologic and genetic characteristics distinguish it from FPV to be considered as a new member of this genus.

Characterization of an avianpox virus isolated from an Andean condor (Vultur gryphus)

A novel pox virus, condorpox virus (CPV) isolated from the spleen of an Andean condor (Vultur gryphus) by inoculation of chorioallantoic membranes (CAM) of specific pathogen free (SPF) chicken embryos was compared biologically, antigenically and genetically with fowlpox virus (FPV), the type species of the genus Avipoxvirus. Susceptible chickens inoculated with CPV developed only mild localized lesions but were not protected against subsequent challenge with FPV. Based on Western blotting, in addition to the presence of cross-reacting antigens, distinct differences in antigenic profiles of CPV and FPV were observed. Sequence analysis of a 4.5 kb HindIII fragment of CPV genomic DNA revealed the presence of eight co-linear genes corresponding to FPV open reading frame (ORF)193–198, 201 and 203. Interestingly, reticuloendotheliosis virus (REV) sequences present in the genome of all FPV were absent in CPV. Although, the results of a phylogenic analysis suggested that CPV is a member of the genus Avipoxvirus, its unique antigenic, biologic and genetic characteristics distinguish it from FPV to be considered as a new member of this genus.

Avian pox infection in Spanish Imperial eagles (Aquila adalberti)

A cutaneous lesion, previously known as "warts", affecting the featherless parts of face and legs has long been recognized in juvenile Spanish Imperial eagles (Aquila adalberti). This paper describes the presentation, microbiological, histopathological, and electron microscopic findings of lesions and diagnosis of poxvirus infection in nine juveniles. Lesions consisted of single or multiple nodules with a crust and surrounded by skin swelling. Seventy-eight percent of the swabs taken from lesions yielded bacterial growth, with Escherichia coli being the most common bacterium isolated. Histopathology revealed typical pox lesions in all cases. Histopathological changes found consisted of proliferative epithelium, with ballooning degeneration of keratinocytes and lymphocyte infiltrates extending into underlying dermis. Avianpox virus was confirmed by the presence of eosinophilic intracytoplasmatic inclusion bodies in the affected cells on light microscopy, and diagnosis confirmation was performed by electron microscopy of biopsies from all nine eagles.

Serum chemistry and antibody status to some avian pathogens of free‐living and captive condors (Vultur gryphus) of central Chile

Summary This study provides information on serology and serum chemistry of the Andean condor (Vultur gryphus). Twenty condors living under natural conditions were captured, blood sampled, measured, and released. In addition, 12 captive condors maintained at the Metropolitan Zoo of Santiago, Chile, were included as a comparison with free‐living birds. All sera were negative to antibodies against reference strains of avian paramyxoviruses of serotypes 1 to 9, avian influenza and avian pox virus. Free‐living condors had total protein, albumin, globulin and Mg values significantly (P< 0.05) lower than those of captive birds. The haematological values obtained in free‐living condors are of particular interest since they may correspond to the optimum values of the species.

Diversified prime and boost protocols using recombinant vaccinia virus and recombinant non-replicating avian pox virus to enhance T-cell immunity and antitumor responses

Recombinant vaccinia viruses containing tumor associated genes represent an attractive vector to induce immune responses to weak immunogens in cancer immunotherapy protocols. The property of intense immunogenicity of vaccinia proteins, however, also serves to limit the number of inoculations of recombinant vaccinia viruses. Host immune responses to the first immunization have been shown to limit the replication of subsequent vaccinations and thus reduce effectiveness of boost inoculations. The use of recombinant avian pox viruses (avipox) such as the canarypox (ALVAC) or fowlpox are potential candidates for immunization protocols in that they can infect mammalian cells and express the inserted transgene, but do not replicate in mammalian cells. We report here the construction and characterization of a canarypox (ALVAC) recombinant expressing the human carcinoembryonic antigen (CEA) gene (designated ALVAC-CEA). Antibody, lymphoproliferative, and cytolytic T-cell responses as well as tumor inhibition were shown to be elicited by the ALVAC-CEA recombinant in a murine model. The utilization of a diversified immunization scheme using a recombinant vaccinia virus followed by recombinant avian pox virus was shown to be far superior than the use of either one alone in eliciting CEA-specific T-cell responses. Experiments were conducted to determine if the use of a diversified immunization scheme using a recombinant vaccinia virus (rV-CEA) and ALVAC-CEA would be superior to the use of either one alone in eliciting CEA-specific T-cell responses. When mice were immunized with rV-CEA and then ALVAC-CEA, CEA-specific T-cell responses were at least four times greater, and far superior to those achieved with three immunizations of ALVAC-CEA. Multiple boosts of ALVAC-CEA following rV-CEA immunization further potentiated anti-tumor effects and CEA specific T-cell responses. These studies demonstrate the proof of concept of the advantage of diversified immunization protocols employing both recombinant vaccinia and recombinant avipox vectors.

Unusual lesions associated with avian poxvirus infection in rosy‐faced lovebirds (Agapornis roseicollis)

Summary An epornitic of avian pox occurred in rosy‐faced lovebirds (Agapomis roseicollis). The infected birds showed a variety of lesions including cutaneous, diphtheritic, systemic and oncogenic entities. Proliferative changes with cytoplasmic inclusion bodies in the cornea, bursa of Fabricius, and cranial and nasal bones which were found in the present cases have not been described previously. Electron microscopic examination of the skin, cornea, and cranial and nasal bones revealed poxvirus virions in the inclusions. Secondary infection of candidiasis was very common in cutaneous pox lesions.

Cross-Protection Test of an Avian Poxvirus Isolated from Houbara Bustards

An avian poxvirus was isolated previously from the houbara bustard (Chlamydotis undulata). We carried out a cross-protection test on 66 captive-bred canaries. Thirty-five canaries were vaccinated with a commercial canary poxvirus (CP) vaccine. Three weeks later all 66 birds were assigned randomly to six different groups: group Ia (n = 14) was vaccinated and challenged with houbara bustard poxvirus (HP) strain; group Ib (n = 13) was vaccinated and challenged with a CP strain; group Ic (n = 7) was vaccinated and not inoculated; group IIa (n = 14) was nonvaccinated and challenged with HP strain; group IIb (n = 11) was nonvaccinated and challenged with a CP strain; and group IIc (n = 7) was not vaccinated and not challenged. Vaccinated groups (Ia, Ib, Ic) had no losses and remained healthy. All of the birds (100%) in group IIb died within 10 days, and 10 birds (71.4%) of group IIa died within 20 days. The nonvaccinated control group (IIc) remained healthy. Poxvirus was isolated from the liver, digestive tract, lungs, and inoculation lesions of nonvaccinated dead CP- and HP-challenged birds. Secondary bacterial infections were higher among nonvaccinated HP-challenged birds (85.7%) than in nonvaccinated CP-challenged birds (25%). The results of this experiment reveal a degree of immunogenic relatedness between CP and HP strain and support the recommendation that houbara bustards be vaccinated with a CP vaccine.

Human safety and immunogenicity of a canarypox-rabies glycoprotein recombinant vaccine: an alternative poxvirus vector system

Avian poxvirus recombinants undergo abortive replication in nonavian cells, yet can achieve expression of extrinsic gene products. Canarypox-vectored vaccines have been innocuous and immunogenic in several mammalian species. ALVAC-RG, a canarypox recombinant expressing the rabies glycoprotein gene, was inoculated intramuscularly into adult volunteers on days 0, 28, and 180. Sequential cohorts received 10 3.5, 10 4.5, and 10 5.5 50% tissue culture infective doses (TCID 50); additional volunteers received the standard human diploid cell rabies vaccine (HDCV) on the same schedule. Reactogenicity of ALVAC-RG was minimal. The lowest dose of ALVAC-RG induced little antibody to rabies virus by ELISA or rapid fluorescent focus inhibition test (RFFIT), but 10 4.5 and 10 5.5 TCID 50 doses elicited significant responses in both assays. All recipients of 10 4.5 and 10 5.5 TCID 50 of ALVAC-RG attained RFFIT values above the presumed protective level. Canarypox-specific immune responses did not inhibit boosting of rabies-specific antibodies by the day 180 dose of ALVAC-RG. T cell proliferation in response to inactivated rabies virus in vitro was similar in HDCV and ALVAC-RG recipients after the first and second doses, although HDCV yielded superior results after the third dose. ALVAC-RG was safe in humans, induced functional antibody to rabies glycoprotein, elicited cellular responses to rabies virus, and could be used successfully for booster dosing at a 6 month interval.