Characterization of Fowlpox Virus.

The complex cytoplasmic DNA virus known as the fowlpox virus (FWPV) is a member of the avipoxvirus genus, Subfamily Chordopoxvirinae, and Family Poxviridae. The large genome size of FWPV makes it a potential vector for the creation of vaccines against a range of serious veterinary and human ailments. It also allows for multiple gene insertion and the generation of abortive infection in mammalian cells. The virus, which causes fowlpox in chickens and turkeys, is mainly transmitted to poultry through aerosols or biting insects. Fowlpox is a highly contagious disease that affects both domestic and wild birds, causing cutaneous and/or diphtheritic illnesses. To control the illness, strict hygiene practices and immunization with FWPV attenuated strains or antigenically similar pigeon pox virus vaccines are employed. Recent years have seen an increase in fowlpox outbreaks in chicken flocks, primarily due to the introduction of novel forms of FWPV. It is believed that the pathogenic characteristics of these strains are enhanced by the integration of reticuloendotheliosis virus sequences of variable lengths into the FWPV genome. The standard laboratory diagnosis of FPV involves histopathological analysis, electron microscopy, virus isolation on chorioallantoic membrane (CAM) of embryonated chicken eggs or cell cultures, and serologic techniques. For quick and consistent diagnosis, polymerase chain reaction (PCR) has proven to be the most sensitive method. PCR is used in concert with restriction endonuclease enzyme analysis (REA) to identify, differentiate, and characterize the molecular makeup of isolates of the fowlpox virus. Sequencing of the amplified fragments is then done.

Development of a Multiplex PCR Assay for Rapid Differentiation of Fowlpox and Pigeonpox Viruses.

The aim of this study was to develop a multiplex PCR assay capable of rapidly differentiating two major Avipoxvirus (APV) species, Fowlpox virus (FWPV) and Pigeonpox virus (PGPV), which cause disease in bird species. Despite the importance of a rapid differentiation assay, no such assay exists that can differentiate the APV species without sequencing. To achieve this, species-specific target DNA fragments were selected from the fpv122 gene of FWPV and the HM89_gp120 gene of PGPV, which are unique to each genome. Nine samples collected from unvaccinated chickens, pigeons, and a turkey with typical pox lesions were genetically identified as FWPV and PGPV. The designed primers and target DNA fragments were validated using in silico analyses with the nucleotide Basic Local Alignment Search Tool. The multiplex PCR assay consisted of species-specific primers and previously described PanAPV primers (genus-specific) and was able to differentiate FWPV and PGPV, consistent with the phylogenetic outputs. This study represents the first successful differentiation of FWPV and PGPV genomes using a conventional multiplex PCR test. This assay has the potential to facilitate the rapid diagnosis and control of APV infections.

Desarrollo de un ensayo de PCR múltiple para la diferenciación rápida de los virus de la viruela aviar y la viruela de paloma. El objetivo de este estudio fue desarrollar un ensayo de PCR múltiple capaz de diferenciar rápidamente dos especies principales de Avipoxvirus (APV) (viruela del pollo), el Fowlpox virus (FWPV) y el Pigeonpox virus (PGPV), (viruela de la gallina), que causan enfermedades en especies de aves. A pesar de la importancia de un ensayo de diferenciación rápida, no existe ningún ensayo que pueda diferenciar las especies de APV sin secuenciación. Para lograr esto, se seleccionaron fragmentos blanco de ADN específicos de especie del gene fpv122 de FWPV y el gene HM89_gp120 de Pigeonpox virus, que son únicos para cada genoma. Nueve muestras recolectadas de pollos, palomas y un pavo que no fueron vacunados con lesiones típicas de la viruela se identificaron genéticamente como FWPV y PGPV. Los iniciadores diseñados y los fragmentos de ADN blanco se validaron mediante análisis in silico mediante la herramienta de búsqueda de alineación local básica de nucleótidos (BLAST). El ensayo de PCR múltiple consistió en iniciadores específicos de especie y cebadores PanAPV previamente descritos (específicos de género) y fue capaz de diferenciar entre Fowlpox virus y Pigeonpox virus, de acuerdo con los resultados filogenéticos. Este estudio representa la primera diferenciación exitosa de los genomas de Fowlpox virus y Pigeonpox virus utilizando una prueba de PCR múltiple convencional. Este ensayo tiene el potencial de facilitar el diagnóstico rápido y el control de las infecciones por Avipoxvirus.

Isolation of Reticuloendotheliosis Virus from Chorio-allantoic Membrane of SPF Chicken Eggs inoculated with Fowl Pox Virus.

Fowl Pox Viruses (FPV) infect chickens and turkeys giving rise to pock lesions on various body parts like combs, wattles, legs, shanks, eyes, mouth etc. The birds, affected with FPV, also show anemia and ruffled appearance which are clinical symptoms of Reticuloendotheliosis. Interestingly, the field strains of FPV are integrated with the provirus of Reticuloendotheliosis Virus (REV). Due to this integration, the infected birds, upon replication of FPV, give rise to free REV virions, causing severe immunosuppression and anemia. Pox scabs, collected from the infected birds, not only show positive PCR results upon performing FPV-specific 4b core protein gene PCR but also show positive results for the PCR of REV-specific env gene and FPV-REV 5'LTR junction. Homogenized suspension of the pock lesions, upon inoculating to the Chorio-allantoic Membrane (CAM) of 10 days old specific pathogen-free embryonated chicken eggs, produces characteristic pock lesions in serial passages. But the lesions also harbor REV mRNA or free virion, which can be identified by performing REV-specific env gene PCR using REV RNA from FPV-infected CAMs. The study suggests successful replication and availability of REV mRNA and free virion alongside the FPV virus, although the CAM is an ill-suited medium for any retroviral (like REV) growth and replication.

Occurrence of enteric viruses causing clinical diarrhea in small ruminants in northern Indian plains: a reverse transcription PCR based molecular study.

Fowlpox virus (FPV) infects chickens and turkeys giving rise to pock lesions on various body parts like combs, wattles, legs, shanks, eyes, mouth, etc. The birds, affected with FPV, also show anemia and a ruffled appearance which are clinical symptoms of reticuloendotheliosis. Interestingly, the field strains of FPV are integrated with the provirus of reticuloendotheliosis virus (REV). Due to this integration, the infected birds, upon replication of FPV, give rise to free REV virions, causing severe immunosuppression and anemia. Pox scabs, collected from the infected birds, not only show positive PCR results upon performing FPV-specific 4b core protein gene PCR but also show positive results for the PCR of REV-specific env gene and FPV-REV 5'LTR junction. Homogenized suspension of the pock lesions, upon inoculating to the chorio-allantoic membrane (CAM) of 10-day-old specific pathogen-free embryonated chicken eggs, produces characteristic pock lesions in serial passages. However, the lesions also harbor REV mRNA or free virion, which can be identified by performing REV-specific env gene PCR using REV RNA from FPV-infected CAMs. The study suggests successful replication and availability of REV mRNA and free virion alongside the FPV, although the CAM is an ill-suited medium for any retroviral (like REV) growth and replication.

Whole-genome based strain identification of fowlpox virus directly from cutaneous tissue and propagated virus.

Fowlpox (FP) is an economically important viral disease of commercial poultry. The fowlpox virus (FPV) is primarily characterised by immunoblotting, restriction enzyme analysis in combination with PCR, and/or nucleotide sequencing of amplicons. Whole-genome sequencing (WGS) of FPV directly from clinical specimens prevents the risk of potential genome modifications associated with in vitro culturing of the virus. Only one study has sequenced FPV genomes directly from clinical samples using Nanopore sequencing, however, the study didn't compare the sequences against Illumina sequencing or laboratory propagated sequences. Here, the suitability of WGS for strain identification of FPV directly from cutaneous tissue was evaluated, using a combination of Illumina and Nanopore sequencing technologies. Sequencing results were compared with the sequence obtained from FPV grown in chorioallantoic membranes (CAMs) of chicken embryos. Complete genome sequence of FPV was obtained directly from affected comb tissue using a map to reference approach. FPV sequence from cutaneous tissue was highly similar to that of the virus grown in CAMs with a nucleotide identity of 99.8%. Detailed polymorphism analysis revealed the presence of a highly comparable number of single nucleotide polymorphisms (SNPs) in the two sequences when compared to the reference genome, providing essentially the same strain identification information. Comparative genome analysis of the map to reference consensus sequences from the two genomes revealed that this field isolate had the highest nucleotide identity of 99.5% with an FPV strain from the USA (Fowlpox virus isolate, FWPV-MN00.2, MH709124) and 98.8% identity with the Australian FPV vaccine strain (FWPV-S, MW142017). Sequencing results showed that WGS directly from cutaneous tissues is not only rapid and cost-effective but also provides essentially the same strain identification information as in-vitro grown virus, thus circumventing in vitro culturing.

laying Farm: Up to Date Data on a Fowlpox Outbreak in Phylogenetic Analysis in Iran, 2018.

Fowlpox is an economically significant viral disease in poultry, characterized by two forms of clinical signs, including cutaneous and diphtheritic lesions. This infection can have several adverse effects on flock performance, such as a reduction in egg production and growth and an increase in mortality. In winter 2018, an infection suspected to fowlpox was reported from a Hy-line W-36 laying farm in Isfahan province, Iran. The birds were 38 weeks of age and showed obvious diphtheritic signs in mucous membranes with increased mortality and reduced egg production. In total, 20 samples were collected from diphtheritic lesions (Trachea and Esophagus) of infected birds. The Polymerase Chain Reaction method was used to amplify a 578 bp fragment of the poxvirus 4b core protein gene. Phylogenetic relationships of avian poxviruses are usually analyzed using the 4b core protein-coding gene sequences with molecular weights of 75.2 kDa. The major elements had the fowlpox genome, and sequencing was performed for one isolate as representative. The nucleotide sequence result showed that this isolate (FP\UT-POX-2018) had a similarity rate of 99.53% with the previous Iranian fowlpox isolate (FP\GHPCRLAB.3) sequenced in the GenBank.Moreover, there was a 100% similarity among the current isolate nucleotide sequence, FP/NobilisVarioleW, and FP/FPV-VR250. The derived phylogenetic tree showed that these isolates were clustered in A1 subclades. Therefore, Iranian isolates of fowlpox virus have remained in the same subclade of phylogenetic classification (subclade A1), and they show high genomic similarity with previous isolates of Iran. Veterinarians and farmers must not underestimate fowlpox. However, they should consider the importance of vaccination against this disease like any other disease care.

An atypical clinicopathological manifestation of fowlpox virus associated with reticuloendotheliosis virus in commercial laying hen flocks in Brazil.

Fowlpox (FP) is a common epitheliotropic disease in chickens that is usually controlled by live attenuated vaccines. However, there have been some reports of outbreaks of FP in recent years, even in vaccinated flocks, presenting as atypical lesions and feathering abnormalities in chickens. These findings can be associated with fowlpox virus (FPV) with the reticuloendotheliosis virus (REV) integrated into its genome. In the present study, outbreaks of atypical FP were explored in vaccinated commercial laying hen flocks to determine the nature of the causative agent by histopathologic and molecular approaches. FPV and REV were detected and classified into subclade A1 of the genus Avipoxvirus and subtype 3 of REV (REV3), respectively. Additionally, heterogeneous populations of FPV with partial (containing only a remnant long terminal repeat-LTR) or total (all functional genes) integration of REV were identified by heterologous PCRs and detected considering reference integration sites. These results indicate the mechanism of chimeric genome FPV-REV associated with outbreaks and atypical clinicopathological manifestations in commercial laying hens for the first time in Brazil and in South America. In addition, this study demonstrates the emergence of REV integrated in the FPV genome in Brazilian chicken flocks.

Efficacy of novel recombinant fowlpox vaccine against recent Mexican H7N3 highly pathogenic avian influenza virus.

Since 2012, H7N3 highly pathogenic avian influenza (HPAI) has produced negative economic and animal welfare impacts on poultry in central Mexico. In the present study, chickens were vaccinated with two different recombinant fowlpox virus vaccines (rFPV-H7/3002 with 2015 H7 hemagglutinin [HA] gene insert, and rFPV-H7/2155 with 2002 H7 HA gene insert), and were then challenged three weeks later with H7N3 HPAI virus (A/chicken/Jalisco/CPA-37905/2015). The rFPV-H7/3002 vaccine conferred 100% protection against mortality and morbidity, and significantly reduced virus shed titers from the respiratory and gastrointestinal tracts. In contrast, 100% of sham and rFPV-H7/2155 vaccinated birds shed virus at higher titers and died within 4 days. Pre- (15/20) and post- (20/20) challenge serum of birds vaccinated with rFPV-H7/3002 had antibodies detectable by hemagglutination inhibition (HI) assay using challenge virus antigen. However, only a few birds (3/20) in the rFPV-H7/2155 vaccinated group had antibodies that reacted against the challenge strain but all birds had antibodies that reacted against the homologous vaccine antigen (A/turkey/Virginia/SEP-66/2002) (20/20). One possible explanation for differences in vaccines efficacy is the antigenic drift between circulating viruses and vaccines. Molecular analysis demonstrated that the Mexican H7N3 strains have continued to rapidly evolve since 2012. In addition, we identified in silico three potential new N-glycosylation sites on the globular head of the H7 HA of A/chicken/Jalisco/CPA-37905/2015 challenge virus, which were absent in 2012 H7N3 outbreak virus. Our results suggested that mutations in the HA antigenic sites including increased glycosylation sites, accumulated in the new circulating Mexican H7 HPAIV strains, altered the recognition of neutralizing antibodies from the older vaccine strain rFPV-H7/2155. Therefore, the protective efficacy of novel rFPV-H7/3002 against recent outbreak Mexican H7N3 HPAIV confirms the importance of frequent updating of vaccines seed strains for long-term effective control of H7 HPAI virus.

Characterization of Fowlpox virus in chickens and bird-biting mosquitoes: a molecular approach to investigating Avipoxvirus transmission.

Avian pox is a highly contagious avian disease, yet relatively little is known about the epidemiology and transmission of Avipoxviruses. Using a molecular approach, we report evidence for a potential link between birds and field-caught mosquitoes in the transmission of Fowlpox virus (FWPV) in Singapore. Comparison of fpv167 (P4b), fpv126 (VLTF-1), fpv175-176 (A11R-A12L) and fpv140 (H3L) gene sequences revealed close relatedness between FWPV strains obtained from cutaneous lesions of a chicken and four pools of Culex pseudovishnui, Culex spp. (vishnui group) and Coquellitidea crassipes caught in the vicinity of the study site. Chicken-derived viruses characterized during two separate infections two years later were also identical to those detected in the first event, suggesting repeated transmission of closely related FWPV strains in the locality. Since the study location is home to resident and migratory birds, we postulated that wild birds could be the source of FWPV and that bird-biting mosquitoes could act as bridging mechanical vectors. Therefore, we determined whether the FWPV-positive mosquito pools (n=4) were positive for avian DNA using a polymerase chain reaction-sequencing assay. Our findings confirmed the presence of avian host DNA in all mosquito pools, suggesting a role for Cx. pseudovishnui, Culex spp. (vishnui group) and Cq. crassipes mosquitoes in FWPV transmission. Our study exemplifies the utilization of molecular tools to understand transmission networks of pathogens affecting avian populations, which has important implications for the design of effective control measures to minimize disease burden and economic loss.

Detection of Fowlpox virus carrying distinct genome segments of Reticuloendotheliosis virus.

Fowlpox virus (FWPV), the type species of the genus Avipoxvirus family Poxviridae, is a large double-stranded DNA virus that causes fowlpox in chickens and turkeys. Notably, sequences of the avian retrovirus reticuloendotheliosis virus (REV) are frequently found integrated into the genome of FWPV. While some FWPV strains carry remnants of the REV long terminal repeats (LTRs), other strains have been shown to contain insertions of nearly the full-length REV provirus in their genome. In the present study we detected heterogeneous FWPV populations carrying the REV LTR or the near full-length REV provirus genome in a Merriam's wild turkey (Meleagris gallopavo merriami). The bird presented papules distributed throughout the non-feathered areas of the head. Avipoxvirus-like virions were observed in the lesions by transmission electron microscopy and the presence of FWPV was confirmed by DNA sequencing. Metagenomic sequencing performed on nucleic acid extracted from the skin lesions revealed two FWPV genome populations carrying either a 197-nt remnant of the REV LTR or a 7939-nt long fragment corresponding to the full-length REV provirus. Notably, PCR amplification using primers targeting FWPV sequences flanking the REV insertion site, confirmed the natural occurrence of the heterogeneous FWPV genome populations in one additional clinical sample from another turkey affected by fowlpox. Additionally, sequencing of a historical FWPV isolate obtained from chickens in the US in 2000 also revealed the presence of the two FWPV-REV genome populations. Results here demonstrate distinct FWPV populations containing variable segments of REV genome integrated into their genome. These distinct genome populations are likely a result of homologous recombination events that take place during FWPV replication.

Spotlight on avian pathology: fowlpox virus.

Fowlpox virus is the type species of an extensive and poorly-defined group of viruses isolated from more than 200 species of birds, together comprising the avipoxvirus genus of the poxvirus family. Long known as a significant poultry pathogen, vaccines developed in the early and middle years of the twentieth century led to its effective eradication as a problem to commercial production in temperate climes in developed western countries (such that vaccination there is now far less common). Transmitted mechanically by biting insects, it remains problematic, causing significant losses to all forms of production (from backyard, through extensive to intensive commercial flocks), in tropical climes where control of biting insects is difficult. In these regions, vaccination (via intradermal or subcutaneous, and increasingly in ovo, routes) remains necessary. Although there is no evidence that more than a single serotype exists, there are poorly-described reports of outbreaks in vaccinated flocks. Whether this is due to inadequate vaccination or penetrance of novel variants remains unclear. Some such outbreaks have been associated with strains carrying endogenous, infectious proviral copies of the retrovirus reticuloendotheliosis virus (REV), which might represent a pathotypic (if not newly emerging) variant in the field. Until more is known about the phylogenetic structure of the avipoxvirus genus (by more widespread genome sequencing of isolates from different species of birds) it remains difficult to ascertain the risk of novel avipoxviruses emerging from wild birds (and/or by recombination/mutation) to infect farmed poultry.

Molecular characterization of avipoxviruses circulating in Mozambique, 2016-2018.

Samples from 45 chickens, two turkeys, one peacock and one quail with symptoms of fowlpox were collected in Mozambique between November 2016 and January 2018. Phylogenetic analysis revealed that the samples contained avipoxviruses belonging to both clade A1 and clade A2. In addition, all of the Clade A1 viruses were positive by PCR for the integration of reticuloendotheliosis virus, while the clade A2 avipoxvirus samples were negative. This study confirms the circulation of clade A1 avipoxviruses in Mozambique in addition to identifying clade A2 for the first time in the country.

Identification of Clade E Avipoxvirus, Mozambique, 2016.

Analysis of scab samples collected from poultry during outbreaks of fowlpox in Mozambique in 2016 revealed the presence of clade E avipoxviruses. Infected poultry were from flocks that had been vaccinated against fowlpox virus. These findings require urgent reevaluation of the vaccine formula and control strategies in this country.

Current and future vaccines and vaccination strategies against infectious laryngotracheitis (ILT) respiratory disease of poultry.

Infectious laryngotracheitis (ILT) is an economically important respiratory disease of poultry that affects the industry worldwide. Vaccination is the principal tool in the control of the disease. Two types of vaccines, live attenuated and recombinant viral vector, are commercially available. The first generation of GaHV-1 vaccines available since the early 1960's are live viruses, attenuated by continuous passages in cell culture or embryos. These vaccines significantly reduce mortalities and, in particular, the chicken embryo origin (CEO) vaccines have shown to limit outbreaks of the disease. However, the CEO vaccines can regain virulence and become the source of outbreaks. Recombinant viral vector vaccines, the second generation of GaHV-1 vaccines, were first introduced in the early 2000's. These are Fowl Pox virus (FPV) and Herpes virus of turkeys (HVT) vectors expressing one or multiple GaHV-1 immunogenic proteins. Recombinant viral vector vaccines are considered a much safer alternative because they do not regain virulence. In the face of challenge, they improve bird performance and ameliorate clinical signs of the disease but fail to reduce shedding of the challenge virus increasing the likelihood of outbreaks. At the moment, several new strategies are being evaluated to improve both live attenuated and viral vector vaccines. Potential new live vaccines attenuated by deletion of genes associated with virulence or by selection of CEO viral subpopulations that do not exhibit increased virulence upon passages in birds are being evaluated. Also new vector alternatives to express GaHV-1 glycoproteins in Newcastle diseases virus (NDV) or in modified very virulent (vv) serotype I Marek's disease virus (MDV) were developed and evaluated.

Concurrent Fowlpox and Candidiasis Diseases in Backyard Chickens with Unusual Pox Lesions in the Bursa of Fabricius.

Concurrent fowlpox and candidiasis diseases occurred in a backyard chicken flock. Four deceased chickens (one Nagoya breed and three white silkie chickens) were examined for diagnosis. At necropsy, white curd-like plaques were observed in the crop. Fungal elements that stained positive for Candida albicans with immunohistochemistry were distributed throughout the tongue, choanal mucosa, esophagus, and crop. Typical fowlpox lesions, composed of proliferating epithelial cells with ballooning degeneration and viral intracytoplasmic inclusions, were observed in the conjunctiva, nasal mucosa, and skin around the cloaca. Interestingly, hyperplastic interfollicular epithelium with rare virus inclusions was observed in the bursa of Fabricius (BF). Some bursal follicles were replaced by proliferating epithelial cells. These proliferating cells immunohistochemically stained positive for cytokeratin. PCR and subsequent genetic sequencing detected the C. albicans gene in the crop, and fowlpox virus genes in the BF. These results indicate that this outbreak was a rare presentation of fowlpox in spontaneously infected chickens, with unusual pox lesions in the BF.

Relationship Between Values of Fowlpox ELISA and the Presence of "Takes" After Vaccination.

Values from an ELISA for evaluating the immune response induced by a commercial vaccine against fowlpox virus and the lesion at the site of inoculation (i.e., swelling of the skin or a pox where the vaccine was applied) were compared. The ELISA was carried out with an antigen prepared by precipitation of a cell culture-propagated virus suspension with ammonium sulfate and concentration by centrifugation. A 0.1 M acetate buffer (pH 5) was used as the sensitizing solution for maximum specific binding of the antigen to the microplate plastic well. Four experiments were conducted where the birds were bled once a week before and after vaccination and then were examined simultaneously for evidence of "takes." This study showed that there is a relationship between the ELISA values to the fowlpox vaccine that are considered positive and the presence of postvaccination lesions.

Outbreaks of Pox Disease Due to Canarypox-Like and Fowlpox-Like Viruses in Large-Scale Houbara Bustard Captive-Breeding Programmes, in Morocco and the United Arab Emirates.

Infectious diseases can be serious threats for the success of reinforcement programmes of endangered species. Houbara Bustard species (Chlamydotis undulata and Chlamydotis macqueenii), whose populations declined in the last decades, have been captive-bred for conservation purposes for more than 15 years in North Africa and the Middle East. Field observations show that pox disease, caused by avipoxviruses (APV), regularly emerges in conservation projects of Houbara Bustard, despite a very strict implementation of both vaccination and biosecurity. Data collected from captive flocks of Houbara Bustard in Morocco from 2006 through 2013 and in the United Arab Emirates from 2011 through 2013 were analysed, and molecular investigations were carried out to define the virus strains involved. Pox cases (n = 2311) were observed during more than half of the year (88% of the months in Morocco, 54% in the United Arab Emirates). Monthly morbidity rates showed strong variations across the time periods considered, species and study sites: Four outbreaks were described during the study period on both sites. Molecular typing revealed that infections were mostly due to canarypox-like viruses in Morocco while fowlpox-like viruses were predominant in the United Arab Emirates. This study highlights that APV remain a major threat to consider in bird conservation initiatives.

Highly pathogenic fowlpox virus in cutaneously infected chickens, China.

We investigated an acute outbreak of the cutaneous form of fowlpox among chickens in China in November 2009. Using pathologic and virologic methods, we identified a novel type of fowlpox virus that carried an integrated genomic sequence of reticuloendotheliosis virus. This highly pathogenic virus could lead to severe ecologic effects and economic losses.

Existence of variant strains Fowlpox virus integrated with Reticuloendotheliosis virus in its genome in field isolates in Tanzania.

Fowlpox virus (FPV) is one example of poultry viruses which undergoes recombination with Reticuloendotheliosis virus (REV). Trepidation had been raised, and it was well established on augmented pathogenicity of the FPV upon integration of the full intact REV. In this study, we therefore intended at assessing the integration of REV into FPV genome of the field isolates obtained in samples collected from different regions of Tanzania. DNA extraction of 85 samples (scabs) was performed, and FPV-specific PCR was done by the amplification of the highly conserved P4b gene. Evaluation of FPV-REV recombination was done to FPV-specific PCR positively identified samples by amplifying the env gene and REV long terminal repeats (5' LTR). A 578-bp PCR product was amplified from 43 samples. We are reporting for the first time in Tanzania the existence of variant stains of FPV integrated with REV in its genome as 65 % of FPV identified isolates were having full intact REV integration, 21 % had partial FPV-REV env gene integration and 5 % had partial 5' LTR integration. Despite of the fact that FPV-REV integrated stains prevailed, FPV-REV-free isolates (9 %) also existed. In view of the fact that full intact REV integration is connected with increased pathogenicity of FPV, its existence in the FPV genome of most field isolates could have played a role in increased endemic, sporadic and recurring outbreaks in selected areas in Tanzania.

Clinical and pathological findings of concurrent poxvirus lesions and aspergillosis infection in canaries.

OBJECTIVE: To investigate clinical, pathological and mycological findings in canaries, in which pox lesions and Aspergillus fumigatus (A. fumigatus) infection were observed simultaneously. METHODS: This study was performed on a breeding colony (about 100 canaries) affected by fatal wasting disease. Necropsy was undertaken on 10 severely affected canaries, and gross lesions were recorded. Samples from internal organs displaying lesions were obtained for histopathological evaluation. Tracheal swap samples of internal organs of the all infected animals with lesions at necropsy were cultured in Sabouraud Dextrose Agar for mycological examination. RESULTS: At necropsy, caseous foci were determined in the lungs, on the air sacs, liver, spleen, heart. Swelling of the eyelids, diffuse hemorrhages in the subcutaneous tissue with small papular lesions of the skin were other typical necropsy findings. Histopathologically, pathognomonic eosinophilic intracytoplasmic inclusion bodies, which called Bollinger bodies, in both skin cells and vacuolated air way epithelial cells confirmed canary pox infection. Moreover, histopathological examination of the white-yellowish caseous foci revealed necrotic granulomatous reaction consisting of macrophages, heterophil leukocytes and giant cells encapsulated with a fibrous tissue. After the culture of the tissue samples, the formation of bluish green colonies confirmed A. fumigatus infection. CONCLUSIONS: Canary pox has been known as the disease that can result in high losses in a short time, as a re-emerging disease that has not been present during recent years in canary flocks in Iran. So, the current paper provides useful information to prevent misdiagnosed of canary pox disease which can cause secondary mycotic infection.