Molecular Diagnosis of Beak and Feather Disease in Native Brazilian Psittacines

The incidence of the psittacine beak and feather disease virus (BFDV) was investigated in Brazilian native parrots with normal feathering arriving at rescue and triage centers for wild animals (CETAS, IBAMA) in the state of Minas Gerais, Brazil. BFDV DNA was investigated by previously described PCR technique for the partial amplification of BFDV ORF-1 in DNA extracts from blood, cloacal swab or liver of psittacines. Some birds provided more than one sample. Nine species of psittacines were sampled between January 2009 and October 2010. Blood (n=46) or cloacal swab (n=128) samples were obtained from psittacines immediately upon arrival at the triage centers. Liver samples were collected from necropsied birds dead on arrival (n=167). All swab samples were negative, except for one Ara ararauna individual (n=3) which blood presented the BFDV DNA. On the other hand, 11 liver samples were positive for BFDV DNA, with a prevalence of 7.8% in Amazona aestiva (n=140). No BFDV DNA was detected in the liver of Amazona amazonica (n=11), A. vinacea (n=5), A. rhodochorytha (n=4), Anodorhynchus hyacinthinus (n=3), Ara ararauna, (n=3), Aratinga leucophtalma (n=2), Guarouba guarouba (n=1) and Pionus maximiliani (n=1). In most cases, alopecia was not associated with BFDV detection in liver, and liver histopathology was inconclusive. Although all cloacal swab samples were negative, a few psittacines (n=19) that died at CETAS-Belo Horizonte were retested, and 21% were detected as positive in liver. A group of psittacines (n=16) was clinically evaluated, and despite showing feather dystrophy, all birds were negative in the cloacal swabs, except for one, which blood sample was positive (A. ararauna). The obtained sequences of the BFDV strains BH 215 and BH 732 were deposited in the GenBank (JQ649409 and JQ649410). A 98% similarity with strain sequences described in Australia, Japan, and New Zealand was observed. It is possible that these strains arrived in Brazil through the legal and illegal trade of parrots. However, it was not possible to associate BFDV infection with the geographical origin of birds and no local marker was detected. The rates of detection, although similar to other studies, indicate the tendency of a high incidence of the disease, possibly associated with stress, and high bird density and wide transmission in captivity conditions.

Molecular Diagnosis of Beak and Feather Disease in Native Brazilian Psittacines

The incidence of the psittacine beak and feather disease virus (BFDV) was investigated in Brazilian native parrots with normal feathering arriving at rescue and triage centers for wild animals (CETAS, IBAMA) in the state of Minas Gerais, Brazil. BFDV DNA was investigated by previously described PCR technique for the partial amplification of BFDV ORF-1 in DNA extracts from blood, cloacal swab or liver of psittacines. Some birds provided more than one sample. Nine species of psittacines were sampled between January 2009 and October 2010. Blood (n=46) or cloacal swab (n=128) samples were obtained from psittacines immediately upon arrival at the triage centers. Liver samples were collected from necropsied birds dead on arrival (n=167). All swab samples were negative, except for one Ara ararauna individual (n=3) which blood presented the BFDV DNA. On the other hand, 11 liver samples were positive for BFDV DNA, with a prevalence of 7.8% in Amazona aestiva (n=140). No BFDV DNA was detected in the liver of Amazona amazonica (n=11), A. vinacea (n=5), A. rhodochorytha (n=4), Anodorhynchus hyacinthinus (n=3), Ara ararauna, (n=3), Aratinga leucophtalma (n=2), Guarouba guarouba (n=1) and Pionus maximiliani (n=1). In most cases, alopecia was not associated with BFDV detection in liver, and liver histopathology was inconclusive. Although all cloacal swab samples were negative, a few psittacines (n=19) that died at CETAS-Belo Horizonte were retested, and 21% were detected as positive in liver. A group of psittacines (n=16) was clinically evaluated, and despite showing feather dystrophy, all birds were negative in the cloacal swabs, except for one, which blood sample was positive (A. ararauna). The obtained sequences of the BFDV strains BH 215 and BH 732 were deposited in the GenBank (JQ649409 and JQ649410). A 98% similarity with strain sequences described in Australia, Japan, and New Zealand was observed. It is possible that these strains arrived in Brazil through the legal and illegal trade of parrots. However, it was not possible to associate BFDV infection with the geographical origin of birds and no local marker was detected. The rates of detection, although similar to other studies, indicate the tendency of a high incidence of the disease, possibly associated with stress, and high bird density and wide transmission in captivity conditions.(AU)

Occurrence of Aviadenovirus in chickens from the poultry industry of Minas Gerais / Ocorrência de Aviadenovirus em aves da industria avícola de Minas Gerais

The occurrence of Aviadenovirus (FAdV) was investigated in chickens from the poultry industry of Minas Gerais state, Brazil. The investigation was conducted due to the scarcity of recent data in the country and its description in neighboring countries. For this purpose, livers were collected from layer chicks (n=25), older layers (n=25), broilers (n=300), and livers (n=25) and stool (n=25) samples from broiler breeders, representing the major poultry regions of the state. FAdV DNA was demonstrated using a previously described PCR protocol for amplifying part of the hexon gene encoding sequence. FAdV was found in layer chicks (36 percent), widespread (100 percent) in older layers, and with regional differences in broilers (24-86 percent). Although all broiler breeder stools were negative, FAdV DNA was detected in livers (16 percent, 4/25) of stool-negative birds. In order to obtain additional information on the circulation of the infection, livers of subsistence chickens collected from one poultry intensive region, were evaluated (n = 12), with FAdV being detected in all samples. FAdV was found in young and old layers, broilers, broiler breeders and free-range chickens, and results suggest the circulation of FAdV among different types of chickens. The detection in older layer chickens may indicate an extended risk of horizontal transmission in regions of Minas Gerais with mixed activity of egg and meat type chickens and poor biosecurity strategies. The infection in breeders may indicate vertical transmission and the continuous production of infected progenies. The hexon-gene-targeted PCR amplicon sequences aligned with FAdV of species D of Aviadenovirus. Results indicate the necessity for biosecurity, especially for breeders, separating flocks according to origin, age and health status, which will be an advantage regarding any pathogen.(AU)

Descreve-se a ocorrência de Aviadenovirus (FAdV) na avicultura mineira. Foram amostrados fígados de poedeiras jovens (n=25) e velhas (n=25) e de frangos de corte (n=300). Em matrizes pesadas foram amostrados fígados (n=25) e fezes (n=25). O estudo envolveu as principais regiões avícolas do Estado de Minas Gerais. O DNA de FAdV foi pesquisado por PCR universal, descrito para a amplificação do gene que codifica o hexon de Aviadenovirus, usando FAdV Phelps como referência. Foi demonstrada a presença do DNA de FAdV em 100 por cento (25/25) das poedeiras velhas (78 semanas de idade) e em 36 por cento (9/25) das jovens (18 dias). Em frangos de corte, a detecção variou entre 24 e 86 por cento. Embora as fezes das matrizes tenham sido negativas, foi obtido o amplicon específico em 4/25 dos fígados dessas mesmas aves, indicando menor sensibilidade para detecção nas fezes. Em amostras da avicultura familiar (fígado), colhidas de uma das regiões de avicultura intensificada, foi detectado o genoma de FAdV em 100 por cento das galinhas (n=12). A constatação de alta disseminação de FAdV em aves da avicultura industrial e familiar de Minas Gerais contribui para o entendimento da epidemiologia de Aviadenovirus. As sequências nucleotídicas dos produtos de PCR alinharam com FAdV da espécie D de Aviadenovirus. A demonstração de FAdV em reprodutores indica risco de transmissão vertical e reforça a necessidade de biosseguridade estrita nesses plantéis. A presença de FAdV em diversos setores avícolas, incluindo poedeiras comerciais, frangos de corte, reprodutores e galinhas da avicultura familiar, recomenda a biosseguridade estrita entre as criações de mesmo tipo e de tipos diferentes de aves. A detecção em matrizes pode indicar a continuada geração de progênies infectadas.(AU)

Health assessment of raptors in triage in Belo Horizonte, MG, Brazil

Falconiformes (n=82), Strigiformes (n=84) and Cathartiformes (n=14) at a triage center (CETAS-Belo Horizonte, IBAMA, Brazil) were examined between 2008 and 2010 . No bird was reactive at hemagglutination-inhibition (HI) for antibodies against Mycoplasma gallisepticum (Mg). Two Caracara plancus (2/68) had HI titers (16-32) against Newcastle disease virus. No Chlamydophila psittaci DNA was detected in the liver (PCR; n=95). Blood smears (Giemsa; n=89) and spleen fragments (PCR; n=82) were 13.5% and 8.5% positive, respectively, for Haemoproteus only. Necropsy of Cathartiformes (n=10), Falconiformes (n=42) and Strigiformes (n=57) showed that trauma injuries were the main cause (63.3%) of admission and death, being fractures (38.5%) of the thoracic limbs (57.1%) the most frequent. Nematode (12.8%), cestode (1.8%), trematode (0.9%), and acanthocephalan (2.7%) parasite infections were relevant. Mites (Acari) were the most frequent (17.4%) external parasites, particularly Ornithonyssus sylviarum in Asio clamator and Amblyomma cajennense in Tyto alba. Chewing lice (10.1%) and Pseudolynchia spp. (9.2%) were also found. Histomonas spp. (6.4%) was found in the ceca of Bubo virginianus, Athene cunicularia, Tyto alba, and Asio clamator, but not in Falconiformes or Cathartiformes. Trichomonas spp. (oral cavity, pharynx and upper esophagus; 9.1%) was detected in Falconiformes and Strigiformes, but not in Cathartiformes. Trichomonas spp. were found in A. cunicularia, Asio clamator, Glaucidium brasilianum and Tyto alba (Strigiformes), and in Rupornis magnirostris, Milvago chimachima, Falco femoralis, Falco sparverius and Caracara plancus (Falconiformes). Coccidia (9.1%) (Sarcocystis spp., 6.4%) and mycosis were observed in most Tyto alba (70%). The evaluated Orders may not pose risks for commercial poultry production. Habitat loss and urban adaptation may be increasingly affecting raptors.

Health assessment of raptors in triage in Belo Horizonte, MG, Brazil

Falconiformes (n=82), Strigiformes (n=84) and Cathartiformes (n=14) at a triage center (CETAS-Belo Horizonte, IBAMA, Brazil) were examined between 2008 and 2010 . No bird was reactive at hemagglutination-inhibition (HI) for antibodies against Mycoplasma gallisepticum (Mg). Two Caracara plancus (2/68) had HI titers (16-32) against Newcastle disease virus. No Chlamydophila psittaci DNA was detected in the liver (PCR; n=95). Blood smears (Giemsa; n=89) and spleen fragments (PCR; n=82) were 13.5% and 8.5% positive, respectively, for Haemoproteus only. Necropsy of Cathartiformes (n=10), Falconiformes (n=42) and Strigiformes (n=57) showed that trauma injuries were the main cause (63.3%) of admission and death, being fractures (38.5%) of the thoracic limbs (57.1%) the most frequent. Nematode (12.8%), cestode (1.8%), trematode (0.9%), and acanthocephalan (2.7%) parasite infections were relevant. Mites (Acari) were the most frequent (17.4%) external parasites, particularly Ornithonyssus sylviarum in Asio clamator and Amblyomma cajennense in Tyto alba. Chewing lice (10.1%) and Pseudolynchia spp. (9.2%) were also found. Histomonas spp. (6.4%) was found in the ceca of Bubo virginianus, Athene cunicularia, Tyto alba, and Asio clamator, but not in Falconiformes or Cathartiformes. Trichomonas spp. (oral cavity, pharynx and upper esophagus; 9.1%) was detected in Falconiformes and Strigiformes, but not in Cathartiformes. Trichomonas spp. were found in A. cunicularia, Asio clamator, Glaucidium brasilianum and Tyto alba (Strigiformes), and in Rupornis magnirostris, Milvago chimachima, Falco femoralis, Falco sparverius and Caracara plancus (Falconiformes). Coccidia (9.1%) (Sarcocystis spp., 6.4%) and mycosis were observed in most Tyto alba (70%). The evaluated Orders may not pose risks for commercial poultry production. Habitat loss and urban adaptation may be increasingly affecting raptors.(AU)

A retrospective PCR investigation of avian Orthoreovirus, chicken infectious anemia and fowl Aviadenovirus genomes contamination in commercial poultry vaccines in Brazil / Estudo retrospectivo de vacinas avícolas vivas por PCR para os vírus da anemia das galinhas, Aviadenovirus aviário e Orthoreovirus aviário

Vacinas avícolas vivas comerciais produzidas entre 1991 e 2005 foram examinadas para a presença de genomas dos vírus da anemia infecciosa das galinhas (Gyrovirus CAV), da hepatite por corpúsculo de inclusão (Aviadenovirus FAdV) e da artrite viral/síndrome da má absorção (Orthoreovirus aviário ARV). Vinte e seis partidas de vacinas vivas liofilizadas de oito fabricantes com lacre original foram examinadas. As extrações de DNA e PCR de CAV e FAdV, e de RNA e RT-PCR para ARV, foram descritas previamente. Contaminações triplas de ARV, CAV e FAdV foram detectadas em vacinas de mesmo fabricante, produzidas em 1991 e 1992 contra a doença de Newcastle (DN), e para a encefalomielite aviária, produzida em 1994. ARV e CAV em co-infecção foram encontrados em vacinas contra a doença de Marek liofilizadas produzidas em 1996 por dois fabricantes diferentes. Genoma de ARV foi detectado em vacinas contra a bronquite infecciosa de setembro e dezembro de 1998, doença infecciosa bursal, de dezembro de 1998 e DN de janeiro de 1998. Três dos oito fabricantes apresentaram vacinas com contaminação e cinco nunca apresentaram vacinas contaminadas. Nenhuma vacina produzida a partir de 2001 apresentou contaminação. Cogita-se um papel epidemiológico para vacinas vivas, como fonte de infecção para ARV, CAV e FAdV e, potencialmente determinante da atual alta disseminação destes.(AU)

The occurrence of Orthoreovirus, Rotavirus and chicken anemia virus in chickens of the poultry industry in Minas Gerais, Brazil / Ocorrência de Orthoreovirus, Rotavirus e vírus da anemia das galinhas em frangos de corte da indústria avícola de Minas Gerais

Fifty-four fecal samples taken from broiler chickens from 1 to 45 days of age, and of pullets from 10 to 13 weeks of age, original from eight different poultry regions in the state of Minas Gerais, Brazil, were collected from March 2008 to January 2010 for avian Orthoreovirus (ARV) and avian Rotavirus (AvRV) analyses. For the assay of ARV, RNA was immediately extracted (Trizolâ) and transcribed into cDNA for assaying in a nested-PCR with ARV-specific primers. For AvRV, polyacrylamide gel electrophoresis (PAGE) was performed with RNA extracts obtained by phenol-chloroform extraction. CAV was additionally investigated through a nested-PCR of thymus and spleen. Results found 5.55% positive for ARV and 9.25% for AvRV. Also, CAV and ARV genomes were detected in co-infection, in a highly prostrated and claudicating chicken flock. No ARV or AvRV infections were detected in pullets. Material of a clinically affected flock was inoculated into SPF embryos, resulting in embryonic hemorrhage, whitish foci in the chorio-allantoic membrane and death. Sequencing of ARV amplicons and isolate cDNA grouped local strains with the ARV S1133 strain, historically used in live vaccines, suggesting the continued circulation of this vaccine virus strain in intensive poultry regions. Detection rates for ARV and AvRV, as well as the presence of CAV, were additionally indicative of failing biosecurity strategies for the intensive poultry regions examined.(AU)

Avaliou-se a ocorrência de Orthoreovirus (ARV) e Rotavirus (AvRV) aviários na avicultura industrial de Minas Gerais. Foram colhidas cinquenta e quatro amostras de fezes de frangos de corte entre um e 45 dias e de frangas de postura de 10 a 13 semanas de idade. Para análise de ARV, o RNA foi imediatamente extraído (Trizol), transcrito em cDNA e avaliado em uma PCR com oligonucleotídeos iniciadores específicos para ARV. Para a investigação de AvRV, os extratos de RNA foram obtidos por fenol-clorofórmio e submetidos à eletroforese em gel de poliacrilamida. Todas as amostras foram também avaliadas para o DNA do vírus da anemia das galinhas (CAV) em uma nested-PCR específica. Em frangos de corte, a positividade encontrada para ARV foi de 5,55% e para AvRV de 9,25%. CAV foi detectado em coinfecção em um plantel com refugagem, claudicação e prostração. Nenhuma amostra de poedeiras foi positiva para ARV ou AvRV. Material de plantel com sinais clínicos foi purificado e inoculado em ovos SPF embrionados, sendo obtidas lesões hemorrágicas e focos brancos na membrana cório-alantóide. O sequenciamento dos produtos de PCR e de embrião agrupou os isolados de ARV com a estirpe S1133, historicamente usada como vacina viva. Os resultados sugerem a continuada circulação da infecção por estirpes assemelhadas a ARV S1133 nas regiões de avicultura industrial. Os índices de detecção de ARV, AvRV e CAV indicam que a intensificação nas regiões produtoras tem resultado em falhas de biosseguridade.(AU)

Purificação de IgG de codorna a partir de soro e gema de ovos / Purification of Japanese quail IgG from serum and egg yolk

The purification of Japanese quail IgG from serum was performed using caprilic acid and ammonium sulfate, and from egg yolk using PEG-6000 and ethanol. After confirming the purification and the concentration of IgG, the yolk samples had twice the amount of protein compared to serum samples. The IgG extracts were analyzed by SDS-PAGE and western blot showing similar results as the ones of chicken IgG used as the standard. So, these methodologies can be used for purifying quail serum or egg yolk IgG, which would enable the development of diagnostic assays.(AU)

Ostrich (Struthio camelus) gastric diseases in Minas Gerais, Brazil, from 1997 to 2009 / Doenças gástricas de avestruzes em Minas Gerais

Descrevem-se as doenças gástricas de avestruzes diagnosticadas em um Laboratório de Doenças das Aves, Belo Horizonte-MG, entre 1997 e 2009. As afecções gástricas corresponderam a 46,2% de todos os casos de doença, com quadros que incluíram impactação (83,3%), infecções e parasitoses (16,7%). As impactações foram causadas por material não alimentar diversificado e as infecções e parasitoses incluíram o fungo Macrorhabdus ornithogaster (megabacteria) e o nematódeo Libyostrongylus douglassii.(AU)

Apoptose e expressão de VP2 e GAPDH na infecção precoce pelo vírus da doença infecciosa da bursa de Fabricius em pintos SPF / Apoptosis and expression of VP2 and GADPH in an experimental infectious bursal disease in SPF chicks

Vinte e nove pintos SPF de um dia foram inoculados com o vírus da doença infecciosa da bursa de Fabricius (VDIB) para avaliar a ocorrência precoce de apoptose e a expressão da proteína viral 2 (VP2) e da enzima gliceraldeído fosfato dehidrogenase (GAPDH). Os animais foram distribuídos em cinco grupos: 1-controle; e 2 a 5- com 24, 48, 72 e 96 horas pós-inoculação, respectivamente. Fragmentos da bursa de Fabricius foram colhidos para processamento histológico e extração de RNA. Lâminas coradas em HE e TUNEL (marcação in situ da fragmentação do genoma com transferase terminal de deoxinucleotídeo) foram utilizadas na morfometria do índice apoptótico. Amostras de mRNA foram testadas para a expressão dos genes VP2 e GAPDH utilizando-se transcrição reversa e RT-PCR. Utilizou-se um kit SYBR GREEN PCR, e a reação foi desenvolvida em ABI Prism 7000 SDS. Os índices apoptóticos cresceram progressivamente indicando uma relação na atrofia bursal causada pelo VDIB. Paralelamente, os resultados da PCR em tempo real demonstraram queda da carga viral nas células linfóides da bursa nos diferentes intervalos de tempo do experimento. Esses resultados sugerem um papel protetor da apoptose na diminuição da replicação viral.(AU)

Twenty-nine SPF 1-day-old chicks were inoculated with infectious bursal disease virus (IBDV) to evaluate early apoptosis and the expression of viral protein 2 (VP2) and glyceraldehyde-3-phosphate dehydrogenease (GAPDH). Five groups were formed: G1-control -and G2 to G5, - 24, 48, 72 and 96 hours post inoculation, respectively. Half of each BF was fixed and processed by routine techniques. To quantify apoptosis, 5µm-thick sections were stained with HE and submitted to TUNEL (terminal transferase UDP nick end labeling) technique. mRNA was extracted from pooled samples of 3 animals/group and used for the expression of VP2 and GADPH genes using the reverse transcription and real-time polymerase chain reaction (RT-PCR). A SYBR GREEN PCR kit was used and the reaction was carried out in an ABI Prism 7000 SDS. Apoptotic indexes progressively increased indicating a role of IBDV in inducing hypotrophy of the BF. Also, it was showed that as long as apoptosis increased, viral protein expression decreased, which suggests that apoptosis plays a role as a defense mechanism against viral replication.(AU)

ELISA indireto para detecção de IgG antivírus da doença de Newcastle em soro de codorna / Indirect ELISA for the detection of IgG specific to Newcastle disease virus in quail serum

An indirect ELISA for the detection of japanese quail IgG specific to Newcastle disease virus (NDV) was developed. The secondary anti-quail IgG was produced in Balb/c mice, by inoculating Freund's complete adjuvant emulsified japanese quail-IgG extract. The purification of IgG was achieved using the caprilic acid method. The ELISA was compared to the haemagglutination-inhibition (HI) test for antibodies to NDV. ELISA cut-off point was established through TG-ROC analysis. Total correlation was observed between the ELISA and the HI, being the ELISA efficient in the identification of positive and negative sera, with high sensitivity and specificity (100 percent). These results validate the use of the indirect ELISA as an alternative for the detection of NDV-specific IgG in japanese quail sera, with the advantage of high sensitivity and automation(AU)

Macrorhabdus ornithogaster in ostrich, rhea, canary, zebra finch, free range chicken, turkey, guinea-fowl, columbina pigeon, toucan, chuckar partridge and experimental infection in chicken, japanese quail and mice / Macrorhabdus ornithogaster em avestruzes, ema, canário, mandarim, galinha, peru, galinha da Angola, pombo doméstico, rolinha, tucano, perdiz de chuckar e infecção experimental em galinha, codorna e camundongo

Since 2000, Macrorhabdus ornithogaster "megabacteriosis" has been diagnosed in the avian diseases laboratory in a diversity of avian species and varied spectrum of disease. The disease in some species (chickens, turkeys, guinea fowls) was clinically characterized by emaciation, prostration, loss of appetite, cachexia and death, with a typically chronic course. A more acute disease was observed in finches (canary-Serinus and zebra-Taeniopygia) and budgerigars (Melopsittacus undulatus). The large rod shaped organism, visible from 100 times magnification, with and without staining, could be detected in sick and also in reasonably normal individuals of some species, such as chickens, turkeys, quails and pigeons. In rheas (Rhea americana), ostriches (Struthio camelus), canaries, zebra-finches, guinea-fowl (Numida meleagris) and budgerigars. The disease was severe, causing to up to 100% mortality. The infection could be detected in some species along with other infectious or disease problems, such as endoparasites (helminths, coccidia) and ectoparasitism (order Mallophaga or/and order Acarina). The cultivation of M. ornithogaster was successfully achieved in solid and liquid media, originated from chickens (four isolates), guinea fowl (1 isolate), chuckar partridge (1 isolate) and canary (1 isolate). A very interesting finding at microscopy was motility of M. ornithogaster, as detected both in cultures obtained on agar for pathogenic fungi and passaged into thioglycolate broth, as well as on samples observed in wet preparations from in vivo. Differences in colony aspects were noted among the isolates. Experimental infections were attempted in chicken and japanese quail, using a chicken isolate, allowing the detection of the organism in the proventriculus and liver in apparently normal birds. One chicken isolate was injected intraperitoneally in Balb/c mice and resulted in 100% mortality.(AU)

Desde 2000, diversos casos de infecção e doença por Macrorhabdus ornithogaster (megabacteria) foram diagnosticados no Setor de Doenças das Aves (Escola de Veterinária da UFMG). A doença clínica foi caracterizada por emagrecimento, prostração, perda do apetite, caquexia e morte, em curso crônico, embora com forma mais aguda em canários e periquitos. O microrganismo grande, em forma de bastão, visível a partir de 100 aumentos sem e com coloração, pode também ser detectado em aves de aspecto clínico normal, principalmente galinhas, perus, codornas e pombos. Em emas (Rhea), avestruzes (Struthio camelus), canários, mandarins, galinhas da Angola (Numida meleagris) e periquitos Australianos (Melopsittacus undulatus), a severidade da doença foi sempre maior, ocasionando até 100% de mortalidade em alguns plantéis. Na maioria das espécies a doença foi detectada em aves com endo e/ou ectoparasitismo. O cultivo de M. ornithogaster foi obtido em meio sólido (ágar para fungos patogênicos) e subcultivado em meio líquido (thioglicolato), do proventriculo de galinha, galinha da Angola, perdiz de chuckar e canário. O resultado mais surpreendente na microscopia de M. ornithogaster foi a presença de motilidade, detectada tanto de cultivos in vitro quanto de preparações úmidas de in vivo. Diferenças nos aspectos das colônias foram notadas entre os isolados. Infecções experimentais em galinha (SPF) e codorna japonesa permitiram a detecção do organismo nos proventrículos das aves de aspecto normal. Nas codornas, à necropsia notaram-se hemorragias hepáticas. A infecção experimental em camundongos via intraperitoneal resultou em 100% de mortalidade, também com lesões hepáticas. Aspectos do cultivo, a importância da doença, as espécies de aves susceptíveis e seu papel na epidemiologia são discutidos.(AU)

Afinidades antigênicas de amostras de campo do vírus da bronquite infecciosa das galinhas com a amostra Massachusetts M41 / Antigenic affinities of infectious bronchitis virus field isolates to Massachusetts M41 strain

Com o objetivo de avaliar as afinidades antigênicas entre 14 amostras de vírus da bronquite infecciosa das galinhas (VBIG) isoladas de casos clínicos ocorridos entre 1972 e 1989 no Estado de Minas Gerais, sua reatividade frente a dois anticorpos monoclonais (AcMs) específicos contra a glicoproteína S1(subscrito) do sorotipo Massachusetts de VBIG foi examinada em ELISA. As 14 amostras de campo estudadas foram agrupadas,de acordo com o relacionamento antigênico aos AcMs, em relacionadas (três amostras) e não relacionadas (onze amostras) à amostra M41 do sorotipo Massachusetts. As amostras de campo não reconhecidas, considerando a alta especificidade dos AcMs à amostra M41, compõem uma diversidade que pode variar de integrantes do sorotipo Massachusetts de origem vacinal a sorotipos heterólogos. Amostras com afinidade antigênica à M41 (208-1972, PM1-1987 e PM2-1987) foram detectadas, o que configura a preservação da amostra no campo, apesar da alta variabilidade da glicoproteína S1(subscrito), já que foram isoladas de surtos de doença natural nas regiões de avicultura de Minas Gerais. A detecção de antígenos de alta variabilidade que caracterizam a amostra M41, apesar das pressões da imunidade dos plantéis e da mutabilidade, pode indicar que os antígenos de alta afinidade aos receptores celulares (best fit) que atingiram alto estágio evolutivo podem estar sendo preservados(AU)

Aiming to the evaluation of antigenic relationships among isolates of infectious bronchitis virus (IBV) through their reactivity against Massachusetts M41 S1 glycopolypeptide specific monoclonal antibodies (Mab) an ELISA was developed. Fourteen IBV isolates obtained from field cases of disease, reported from 1972 to 1989 in Minas Gerais, Brazil, were examined. The IBV isolates could be grouped into related or not to M41, based on the reactivity to M41 S1 specific Mabs. The unrecognized field isolates conform a diversity of representatives, which may range from Massachusetts vaccine strains to unrelated heterologous IBV serotypes. The detection of three M41-related isolates (208-1972, PM1-1987 and PM2-1987) indicate that, despite the high variability and pressure for changes in the S1 protein, M41 clones are still present in the field and may point out that the survival strategies for isolates should include mechanisms that preserve the best fits for cellular receptors. (AU)

An outbreak of malaria by Haemoproteus columbae in pigeons

Relata-se um surto de malária por Haemoproteus columbae em pombos-correio (Columba livia), caracterizado por um quadro agudo com mortalidade de 3% ao dia em aves de aspecto saudável ou crônico com prostração e fraqueza. O exame clínico das aves doentes evidenciou sinusite e conjuntivite e à necropsia traqueíte hemorrágica, aerossaculite, esplenomegalia, nefromegalia e hepatomegalia com hemorragias. O díptero hematófago da família Hippoboscidae Pseudolynchia canariensis foi encontrado inserido entre as penas dos indivíduos. Esfregaços de sangue periférico e cardíaco e impressões de baço e fígado corados por Giemsa permitiram a visualização de inclusões intraeritrocitárias características de Haemoproteus columbae

Fatal sarcosporidiosis in a passerine of species Gnorimopsar chopi chopi / Sarcosporidiose fatal em pássaro-preto (Gnorimopsar chopi chopi)

Relata-se um caso fatal de sarcosporidiose em pássaro-preto (melro), Gnorimopsar chopi chopi, caracterizado por intensa prostração, respiração superficial e decúbito lateral. Embora nao tenham sido observadas alterações significativas no sistema respiratório, o quadro clínico confundia-se com doença nesse sistema, em decorrência do comprometimento dos músculos ligados à respiração.(AU)

Cryopreservation of an avian spirochete strain in liquid nitrogen

Soros de aves experimentalmente infectadas, contendo espiroquetas viáveis, foram submetidos a dois procedimentos antes da criopreservação: glicerol na diluição de 1/2 (v/v), designado como soro com glicerol a 50% (GS), e dimetilsulfóxido na proporção de 1/10 (v/v), designado como soro com DMSO a 10% (DS). Após 15 meses de estocagem em nitrogênio líquido, amostras dos tratamentos GS e DS foram descongeladas e suas infectividades foram testadas em frangos susceptíveis. Apesar de ambos os procedimentos terem mantidos a infectividade da bactéria, DMSO a 10% no soro de frango apresentou-se mais satisfatório como criopreservante.

Resultados preliminares da utilização de cultivos de anéis de traquéia para o estudo de estirpes brasileiras do vírus da bronquite infecciosa das galinhas / A preliminary use of tracheal organ cultures for evaluating Brazilian infectious bronchitis virus strains

Tracheal organ cultures (TOC) were prepared and used for evaluating four Brazilian isolates of infectious bronchitis virus (IBV). IBV field isolates and vaccine strains were titrated in TOC and results compared to those from chicken embrionated eggs. Serum neutralization (SN) employing IBV strain-specific serum was performed for evaluating relationships between isolates. Titration results of tests performed in TOC or eggs were in mutual agreement and were considered for validating the adapted TOC methodology as alternative for virological studies in our laboratory. Sera specific to M41 (Massachusetts) or A5968 (Connecticut) did neutralize their respective IBV strains only. Field strains 208 and 29-78 specific sera did neutralize Massachusetts serotype strains M41 and H120, but PM2 serum did only M41. Strain PM4 specific serum did not neutralize any of the reference IBV analyzed, including M41, A5968 and H120 and may indicate that the isolate is serologically different from the Massachusetts serotype, currently adopted for vaccine strains in Brazil.(AU)

Estudo comparativo da virulência de amostras de vacina do vírus da doença de Newcastle em galinhas SPF por meio da análise morfométrica da espessura traqueal

Virulence of three vaccinal lentogenic strains (La Sota, Ulster and VG-GA) of the Newcastle disease virus (NDV) was determined by morphometric analysis of tracheal thickness, in 45-day-old SPF chickens (n=12), free from NDV antibodies. Tracheal thickness was evaluated 1, 2, 3, 4, 6, and 8 days after intratracheal vaccination. La Sota strain induced higher tracheal swelling than the others, during most of the time of the experiment. Maximum swelling of tracheal mucosa occurred at the thirdday after vaccination. At that time, La Sota and Ulster had the same virulence, and both caused higher swelling of tracheal mucosa than VG-GA strain.

A virulência das amostras de vacinas lentogênicas La Sota, Ulster e VG-GA do vírus da doença de Newcastle (VDN) foi determinada por análise morfométrica da espessura da traquéia de galinhas (n=12) de 45 dias de idade, livres de anticorpos anti-VDN, vacinadas por via intra-traqueal. As traquéias foram avaliadas 1, 2, 3, 4, 6 e 8 dias pós-vacinação. A amostra La Sota induziu maior espessamento da traquéia no decorrer de todo período experimental. Ao terceiro dia pós-vacinação, período em que as traquéias se apresentaram mais espessas, as amostras La Sota e Ulster não diferiram em sua virulência, sendo ambas mais virulentas do que a amostra VG-GA, induzindo maior espessamento de traquéia e causando lesões histológicas mais severas.

Production of monoclonal antibodies against conserved components of infectious bronchitis virus

Murine hybridomas producing IgG1 monoclonal antibodies (Mabs) against N and S2 proteins (53KDa and 82KDa, respectively) from avian infection bronchitis virus (IBV) strain M41 were generated by the fusion of a myeloma cell line (Sp2/0-Ag14) with spleen cells from Balb/c mice previously immunized with whole virus IBV M41. Post-fusion screening criterion was by ELISA and 36 positive hybrids were generated after fusions. Two hybrids specific to N (N3F10) and S2 (S12B2) proteins from M41 (serotype Massachusetts) were selected by western blotting. These Mabs recognized the Ark-99 (serotype Arkansas) and A5968 (serotype Connecticut) IBV strains in addition to M41. By ELISA, the Mab against the S2 (S12B2) recognized all reference and Brazilian strains (M41, SE-17, H52, 297, 283, PM-1, PM-2, PM-3, 351, 29-78 E 327) studied, while the Mab against N recognized only six (M41, SE-17, H52, 283, 327 e 297) strains. The Mab against S2 may become a useful tool for IBV detection on the routine diagnosis of infectious bronchitis, especially for helping the differential diagnosis of clinically and pathologically confusing diseases, while the Mab against N (N3F10) recognized a probably less conserved region among the strains and may be interesting to comparing IBV isolates.

Foram produzidos anticorpos monoclonais (AcM) da subclasse IgG1 contra as proteínas N (53KDa) e S2 (82KDa) do vírus da bronquite infecciosa das galinhas (VBIG) amostra M41. Os híbridos secretores originaram-se da fusão entre células de mielomas da linhagem Sp2/0-Ag14 e linfócitos B de camundongos Balb/c previamente imunizados com o vírus completo. O primeiro critério de seleção foi por ELISA, no qual 36 híbridos originados de duas fusões reagiram positivamente; destes, foram selecionados dois AcM que reagiram contra as proteínas N (N3D4) e S2 (S12B2) do VBIG da amostra M41 (sorotipo Massachusetts) no western blotting. Os mesmos AcM foram também capazes de reconhecer as estirpes Ark-99 (sorotipo Arkansas) e A5968 (sorotipo Connecticut) do VBIG no western blotting. No ELISA, o AcM contra S2 (S12b2) reconheceu 11 estirpes das 11 estudadas (M41, SE-17, H52, 297, 283, PM-1, PM-2, PM-3, 351, 29-78 E 327), enquanto o AcM contra a proteína N reconheceu apenas seis estirpes (M41, SE-17, H52, 283, 327 e 297). O anticorpo monoclonal contra S2 comportou-se como uma boa ferramenta de diagnóstico do VBIG, independentemente do sorotipo que pode ser aplicado em ensaios de rotina laboratorial, especialmente no diagnóstico das doenças que se confundem com a BIG nas galinhas, enquanto o AcM contra N (N3F10) demonstrou reconhecer uma região menos conservada entre as estirpes de VBIG e pode ser útil em estudos comparativos entre isolados de VBIG.

Mortality by filarial nematodes in passeriformes of species Oryzoborus maximiliani in Brazil

Descreve-se a ocorrência de filariose em passeriformes da espécie Oryzoborus maximiliani (bicudo) mantidos em cativeiro. Os sinais clínicos incluiram insuficiência respiratória e prostração, evoluindo para decúbito lateral e morte. Todos os indivíduos capturados adoeceram e morreram em poucos dias. As lesões mais significativas foram encontradas nos pulmões, que estavam acinzentados na região adjacente ao saco aéreo abdominal. Impressões do pulmão observadas ao microscópio em 100 aumentos permitiram a visualização de grande número de formas alongadas típicas de nematódeo. Considerando suas dimensões e os relatos da literatura consultada, especulou-se a possibilidade de filariose. As condições de estresse de captura e cativeiro podem ser determinantes do quadro agudo observado, o que permite sugerir esta suspeita em casos semelhantes. Considera-se importante, entretanto, a possibilidade de a manifestação clínica na forma crônica ou assintomática poder ser mais comum que a aguda.