A five-year surveillance study of vaccination schedules using viral-vectored vaccines against infectious laryngotracheitis in a high-density layer region / Cinco anos de monitoramento de esquemas vacinais com vacinas virais vetorizadas contra laringotraqueíte infecciosa em uma região altamente povoada por galinhas poedeiras

The effectiveness of vectored recombinant vaccines to control infectious laryngotracheitis (ILT) in chickens from a region (State of Minas Gerais, Brazil) with ~10 million layers was evaluated under field conditions from 2014-2018. During this period, only recombinant turkey herpesvirus (rHVT) or fowl poxvirus (rFPV) vaccines that express antigens of infectious laryngotracheitis virus (Gallid herpesvirus-1; GaHV-1) were used. Layer chickens (n=1,283), from eight different egg-producing companies, were individually sampled and examined (active surveillance), and in instances when government poultry health veterinarians were notified due to respiratory disease (passive surveillance). Clinical, macroscopic, and histopathology examinations were performed to diagnose ILT as well as molecular techniques for the detection and characterization of the GaHV-1 DNA from the trachea and trigeminal ganglia (TG). The layer hens sampled and examined belonged to flocks and farms that used different vaccination protocols (non-vaccinated, single dose vaccination, and prime/ boost vaccination). This is the first long-term field study of the effectiveness of ILT vectored vaccines in a high-density multiple age layer hen region. Using various diagnostic methods, the occurrence of GaHV-1 infection and ILT clinical disease in layer hens vaccinated with vectored recombinant vaccines in one quarantined region of Brazil were investigated. The number of ILTV positive chickens by PCR and ILT clinical disease cases was lower in farms when all chickens were vaccinated with at least one vaccine. However, the difference in the detection rates of GaHV-1 infection was significant only when compared farms with prime/ boost and farms using single dose of HTV-LT.

A efetividade das vacinas recombinantes vetorizadas para o controle da laringotraqueíte infecciosa (LTI) nas aves de uma região (Minas Gerais, Brasil) com aproximadamente 10 milhões de poedeiras foi avaliada em condições de campo, no período de 2014 a 2018. Durante este período, somente as vacinas recombinantes "turkey herpesvirus" (rHVT) ou "fowl poxvirus" (rFPV), que expressam antígenos do vírus da laringotraqueíte (Gallid herpesvirus-1; GaHV-1) foram utilizadas. Galinhas poedeiras (n=1.283), de oito diferentes granjas produtoras de ovos, foram individualmente amostradas e examinadas por monitoramento ativo e, na ocorrência de notificação de doença respiratória aos veterinários do serviço oficial, por monitoramento passivo. Exames clínicos, macroscópicos e histopatológicos foram realizados para o diagnóstico de LTI, bem como técnicas moleculares para a detecção e caracterização do DNA de GaHV-1 da traqueia e gânglio trigêmeo. As galinhas poedeiras pertenciam a lotes e granjas que usavam diferentes protocolos de vacinação (não vacinadas, uma dose ou tipo de vacina e duas doses ou tipos de vacina). Este é o primeiro longo estudo a campo sobre a efetividade das vacinas vetorizadas em uma região com população elevada de poedeiras de múltiplas idades. Utilizando vários métodos de diagnóstico, a ocorrência da infecção por GaHV-1 e a LTI clínica em poedeiras de uma região interditada do Brasil foi investigada. O número de galinhas positivas para o vírus GaHV-1 e para casos clínicos de LTI nas granjas foi menor quando todas as aves estavam vacinadas com, pelo menos, um tipo ou dose de vacina. Entretanto, a diferença na taxa de detecção da infecção por GaHV-1 foi significativa somente quando a comparação foi realizada entre granjas com aves vacinadas com duas doses e aves de granjas vacinadas com uma única dose de HVT-LT.

Anatomopathological, ultrastructural, immunohistochemical and molecular characterization of infectious laryngotracheitis outbreaks in poultry farms in Egypt (2018–2020)

Infectious laryngotracheitis (ILT) is a severe respiratory disease, which causes high morbidity and mortality in affected birds. In our study, ILT were reported in 42 farms from nine governates over two years (2018–2020) that showed clinical signs of ILT including dyspnea, blood expectoration of, excessive lacrimation, rattling, conjunctivitis. The disease affected different chicken breeds and age groups despite vaccination with licensed and commonly used vaccines. Samples of larynx, trachea, lungs and air sacs were examined and collected for histopathological, ultrastructural, immunohistochemical examination and molecular detection. Gross examination of laryngeal and tracheal lumen revealed different types of exudate varied from catarrhal to fibrinonecotric, also pneumonia and airsacculitis were detected. Histopathological examination showed different alternation in larynx, trachea, lung and air sac as characteristic syncytial cells containing intranuclear inclusion body hanged in fibrinoheterphilic exudate that precent in laryngeal, tracheal, bronchial and parabronchial lumen and air sacs. Tracheal lesion scoring system was used to categorize the severity of lesion in different governates. Tracheal lesion score showed that 6.02%, 26.5%, 43.3% of the birds exhibited mild, moderate, and severe changes, respectively, while 24.18% of the birds exhibited very severe changes. Furthermore, severe cases were related to the Qalyubia , Fayoum then Sharkia Governorate. Moreover, immunohistochemistry was used to detect viral particles in syncytial cells, inflammatory cells beside epithelium of trachea and lung. Transmission electron microscopy enabled the detection of virus particles and demonstrated that heterophils could be infected. PCR targeting a region in the thymidine kinase gene and glycoprotein gJ gene confirmed the presence of infectious laryngotracheitis ILT virus-specific DNA. In conclusion, anatomopathological, immunohistochemical, molecular and ultrastructural findings showed increased of ILTV severity in Egypt. Larynx, trachea, lungs and air sac should be collected and examined that aid in diagnosis. Importance of good biosecurity level to be considered.(AU)

Anatomopathological, ultrastructural, immunohistochemical and molecular characterization of infectious laryngotracheitis outbreaks in poultry farms in Egypt (2018–2020)

Infectious laryngotracheitis (ILT) is a severe respiratory disease, which causes high morbidity and mortality in affected birds. In our study, ILT were reported in 42 farms from nine governates over two years (2018–2020) that showed clinical signs of ILT including dyspnea, blood expectoration of, excessive lacrimation, rattling, conjunctivitis. The disease affected different chicken breeds and age groups despite vaccination with licensed and commonly used vaccines. Samples of larynx, trachea, lungs and air sacs were examined and collected for histopathological, ultrastructural, immunohistochemical examination and molecular detection. Gross examination of laryngeal and tracheal lumen revealed different types of exudate varied from catarrhal to fibrinonecotric, also pneumonia and airsacculitis were detected. Histopathological examination showed different alternation in larynx, trachea, lung and air sac as characteristic syncytial cells containing intranuclear inclusion body hanged in fibrinoheterphilic exudate that precent in laryngeal, tracheal, bronchial and parabronchial lumen and air sacs. Tracheal lesion scoring system was used to categorize the severity of lesion in different governates. Tracheal lesion score showed that 6.02%, 26.5%, 43.3% of the birds exhibited mild, moderate, and severe changes, respectively, while 24.18% of the birds exhibited very severe changes. Furthermore, severe cases were related to the Qalyubia , Fayoum then Sharkia Governorate. Moreover, immunohistochemistry was used to detect viral particles in syncytial cells, inflammatory cells beside epithelium of trachea and lung. Transmission electron microscopy enabled the detection of virus particles and demonstrated that heterophils could be infected. PCR targeting a region in the thymidine kinase gene and glycoprotein gJ gene confirmed the presence of infectious laryngotracheitis ILT virus-specific DNA. In conclusion, anatomopathological, immunohistochemical, molecular and ultrastructural findings showed increased of ILTV severity in Egypt. Larynx, trachea, lungs and air sac should be collected and examined that aid in diagnosis. Importance of good biosecurity level to be considered.

Evaluation of actions of the official veterinary service to mitigate outbreaks of infectious laryngotracheitis and improve biosecurity on laying hen farms

ABSTRACT: Infectious laryngotracheitis (ILT), caused by an Alphaherpesvirus (Gallid herpesvirus-1; GaHV-1), has been noticed in the region of the Terras Altas da Mantiqueira, Minas Gerais. From 2010 to 2018, the Serviço Veterinário Oficial (SVO) of the Instituto Mineiro Agropecuário (IMA), implemented measures to prevent spread of the virus to other regions and control the disease in the area. Due to the close proximity and consequent epidemiological link among farms, the region was considered a unique epidemiological unit. To check the efficiency of the ILT control measures, we carried out: (1) a seroepidemiological survey, (2) questionnaires for evaluating biosecurity measures; and (3) an evaluation of the influence of farm population density on the occurrence of ILT. In 2016, 2017, and 2018, ILT was investigated using epidemiological and clinicopathological methods, along with GaHV-1 molecular detection. Serological survey was carried out on 24 farms in the quarantined region and on 13 farms from other regions of the state. In 2010 and 2018, questionnaires were applied to collect data and determine indicators of biosecurity practices in all farms of the quarantined area. The differences were then assessed (Wilcoxons p 0.05). The results indicated positive serology throughout the region, although only on four farms (16.6%) the chickens have clinical signs, macroscopic and histological lesions of ILT. The prevalence of viral infection increased from 2016 (27%) to 2017 (50%) and was higher in farms with a high stock density (p=0.033). No disease, virus or antibodies were detected in the farms outside of the quarantined area. Although the biosecurity indicators had improved on all farms in the quarantined area (p 0.05), the virus was active and circulating in the region. The contingency measures have contained the outbreak, but biosecurity practices are paramount in the control of new outbreaks. Official control will be maintained in the region, including surveillance of new cases and biosecurity procedures to mitigate the risk of the virus reaching other regions.

RESUMO: Laringotraqueíte infecciosa (LTI), causada por um alfaherpesvírus (herpesvírus Gallid-1; GaHV-1), foi observada na região das Terras Altas da Mantiqueira, Minas Gerais. De 2010 a 2018, o Serviço Veterinário Oficial (SVO) do Instituto Mineiro Agropecuário (IMA) implementou medidas para impedir a disseminação do vírus para outras regiões do estado e controlar a doença na região interditada. Devido à proximidade e consequente vínculo epidemiológico entre as granjas, a região foi considerada uma unidade epidemiológica única. Para verificar a eficiência das medidas de controle de LTI, foram realizados: (1) pesquisa soroepidemiológica, (2) questionários para avaliar medidas de biosseguridade; e (3) avaliação da influência da densidade populacional da granja na ocorrência de LTI. Em 2016, 2017 e 2018, a LTI foi investigada usando métodos epidemiológicos e clínico-patológicos, com a detecção molecular de GaHV-1. O levantamento sorológico foi realizado em 24 granjas da região interditada e em 13 granjas de outras regiões do estado. Em 2010 e 2018, foram aplicados questionários para coletar dados e determinar indicadores de medidas de biosseguridade em todas as granjas da área interditada. As diferenças foram avaliadas (p 0,05 de Wilcoxon). Os resultados indicaram sorologia positiva em toda a região, embora apenas em quatro granjas (16,6%) as galinhas apresentaram sinais clínicos, lesões macroscópicas e histológicas da LTI. A prevalência de infecção viral aumentou de 2016 (27%) para 2017 (50%) e foi maior em fazendas com alta densidade de alojamento (p=0,033). Presença da doença, vírus ou anticorpos foram detectados nas granjas fora da área interditada. Embora os indicadores de biosseguridade tenham melhorado em todas as fazendas da área interditada (p 0,05), o vírus está ativo e circulava na região. As medidas de contingência contiveram o surto, mas as práticas de biosseguridade são fundamentais para o controle de novos surtos. O controle oficial será mantido na região, incluindo a vigilância de novos casos e procedimentos de biosseguridade para mitigar o risco de transmissão do vírus para outras regiões.

Evaluation of actions of the official veterinary service to mitigate outbreaks of infectious laryngotracheitis and improve biosecurity on laying hen farms / Avaliação de ações do serviço veterinário oficial para bloqueio de foco de laringotraqueite infecciosa e melhoria da biosseguridade em granjas de postura

Infectious laryngotracheitis (ILT), caused by an Alphaherpesvirus (Gallid herpesvirus-1; GaHV-1), has been noticed in the region of the Terras Altas da Mantiqueira, Minas Gerais. From 2010 to 2018, the "Serviço Veterinário Oficial" (SVO) of the "Instituto Mineiro Agropecuário" (IMA), implemented measures to prevent spread of the virus to other regions and control the disease in the area. Due to the close proximity and consequent epidemiological link among farms, the region was considered a unique epidemiological unit. To check the efficiency of the ILT control measures, we carried out: (1) a seroepidemiological survey, (2) questionnaires for evaluating biosecurity measures; and (3) an evaluation of the influence of farm population density on the occurrence of ILT. In 2016, 2017, and 2018, ILT was investigated using epidemiological and clinicopathological methods, along with GaHV-1 molecular detection. Serological survey was carried out on 24 farms in the quarantined region and on 13 farms from other regions of the state. In 2010 and 2018, questionnaires were applied to collect data and determine indicators of biosecurity practices in all farms of the quarantined area. The differences were then assessed (Wilcoxon's p<0.05). The results indicated positive serology throughout the region, although only on four farms (16.6%) the chickens have clinical signs, macroscopic and histological lesions of ILT. The prevalence of viral infection increased from 2016 (27%) to 2017 (50%) and was higher in farms with a high stock density (p=0.033). No disease, virus or antibodies were detected in the farms outside of the quarantined area. Although the biosecurity indicators had improved on all farms in the quarantined area (p<0.05), the virus was active and circulating in the region. The contingency measures have contained the outbreak, but biosecurity practices are paramount in the control of new outbreaks. Official control will be maintained in the region, including surveillance of new cases and biosecurity procedures to mitigate the risk of the virus reaching other regions.(AU)

Laringotraqueíte infecciosa (LTI), causada por um alfaherpesvírus (herpesvírus Gallid-1; GaHV-1), foi observada na região das Terras Altas da Mantiqueira, Minas Gerais. De 2010 a 2018, o Serviço Veterinário Oficial (SVO) do Instituto Mineiro Agropecuário (IMA) implementou medidas para impedir a disseminação do vírus para outras regiões do estado e controlar a doença na região interditada. Devido à proximidade e consequente vínculo epidemiológico entre as granjas, a região foi considerada uma unidade epidemiológica única. Para verificar a eficiência das medidas de controle de LTI, foram realizados: (1) pesquisa soroepidemiológica, (2) questionários para avaliar medidas de biosseguridade; e (3) avaliação da influência da densidade populacional da granja na ocorrência de LTI. Em 2016, 2017 e 2018, a LTI foi investigada usando métodos epidemiológicos e clínico-patológicos, com a detecção molecular de GaHV-1. O levantamento sorológico foi realizado em 24 granjas da região interditada e em 13 granjas de outras regiões do estado. Em 2010 e 2018, foram aplicados questionários para coletar dados e determinar indicadores de medidas de biosseguridade em todas as granjas da área interditada. As diferenças foram avaliadas (p<0,05 de Wilcoxon). Os resultados indicaram sorologia positiva em toda a região, embora apenas em quatro granjas (16,6%) as galinhas apresentaram sinais clínicos, lesões macroscópicas e histológicas da LTI. A prevalência de infecção viral aumentou de 2016 (27%) para 2017 (50%) e foi maior em fazendas com alta densidade de alojamento (p=0,033). Presença da doença, vírus ou anticorpos foram detectados nas granjas fora da área interditada. Embora os indicadores de biosseguridade tenham melhorado em todas as fazendas da área interditada (p<0,05), o vírus está ativo e circulava na região. As medidas de contingência contiveram o surto, mas as práticas de biosseguridade são fundamentais para o controle de novos surtos. O controle oficial será mantido na região, incluindo a vigilância de novos casos e procedimentos de biosseguridade para mitigar o risco de transmissão do vírus para outras regiões.(AU)

Evaluation of actions of the official veterinary service to mitigate outbreaks of infectious laryngotracheitis and improve biosecurity on laying hen farms / Avaliação de ações do serviço veterinário oficial para bloqueio de foco de laringotraqueite infecciosa e melhoria da biosseguridade em granjas de postura

Infectious laryngotracheitis (ILT), caused by an Alphaherpesvirus (Gallid herpesvirus-1; GaHV-1), has been noticed in the region of the Terras Altas da Mantiqueira, Minas Gerais. From 2010 to 2018, the "Serviço Veterinário Oficial" (SVO) of the "Instituto Mineiro Agropecuário" (IMA), implemented measures to prevent spread of the virus to other regions and control the disease in the area. Due to the close proximity and consequent epidemiological link among farms, the region was considered a unique epidemiological unit. To check the efficiency of the ILT control measures, we carried out: (1) a seroepidemiological survey, (2) questionnaires for evaluating biosecurity measures; and (3) an evaluation of the influence of farm population density on the occurrence of ILT. In 2016, 2017, and 2018, ILT was investigated using epidemiological and clinicopathological methods, along with GaHV-1 molecular detection. Serological survey was carried out on 24 farms in the quarantined region and on 13 farms from other regions of the state. In 2010 and 2018, questionnaires were applied to collect data and determine indicators of biosecurity practices in all farms of the quarantined area. The differences were then assessed (Wilcoxon's p<0.05). The results indicated positive serology throughout the region, although only on four farms (16.6%) the chickens have clinical signs, macroscopic and histological lesions of ILT. The prevalence of viral infection increased from 2016 (27%) to 2017 (50%) and was higher in farms with a high stock density (p=0.033). No disease, virus or antibodies were detected in the farms outside of the quarantined area. Although the biosecurity indicators had improved on all farms in the quarantined area (p<0.05), the virus was active and circulating in the region. The contingency measures have contained the outbreak, but biosecurity practices are paramount in the control of new outbreaks. Official control will be maintained in the region, including surveillance of new cases and biosecurity procedures to mitigate the risk of the virus reaching other regions.(AU)

Laringotraqueíte infecciosa (LTI), causada por um alfaherpesvírus (herpesvírus Gallid-1; GaHV-1), foi observada na região das Terras Altas da Mantiqueira, Minas Gerais. De 2010 a 2018, o Serviço Veterinário Oficial (SVO) do Instituto Mineiro Agropecuário (IMA) implementou medidas para impedir a disseminação do vírus para outras regiões do estado e controlar a doença na região interditada. Devido à proximidade e consequente vínculo epidemiológico entre as granjas, a região foi considerada uma unidade epidemiológica única. Para verificar a eficiência das medidas de controle de LTI, foram realizados: (1) pesquisa soroepidemiológica, (2) questionários para avaliar medidas de biosseguridade; e (3) avaliação da influência da densidade populacional da granja na ocorrência de LTI. Em 2016, 2017 e 2018, a LTI foi investigada usando métodos epidemiológicos e clínico-patológicos, com a detecção molecular de GaHV-1. O levantamento sorológico foi realizado em 24 granjas da região interditada e em 13 granjas de outras regiões do estado. Em 2010 e 2018, foram aplicados questionários para coletar dados e determinar indicadores de medidas de biosseguridade em todas as granjas da área interditada. As diferenças foram avaliadas (p<0,05 de Wilcoxon). Os resultados indicaram sorologia positiva em toda a região, embora apenas em quatro granjas (16,6%) as galinhas apresentaram sinais clínicos, lesões macroscópicas e histológicas da LTI. A prevalência de infecção viral aumentou de 2016 (27%) para 2017 (50%) e foi maior em fazendas com alta densidade de alojamento (p=0,033). Presença da doença, vírus ou anticorpos foram detectados nas granjas fora da área interditada. Embora os indicadores de biosseguridade tenham melhorado em todas as fazendas da área interditada (p<0,05), o vírus está ativo e circulava na região. As medidas de contingência contiveram o surto, mas as práticas de biosseguridade são fundamentais para o controle de novos surtos. O controle oficial será mantido na região, incluindo a vigilância de novos casos e procedimentos de biosseguridade para mitigar o risco de transmissão do vírus para outras regiões.(AU)

Epidemiology of avian infectious laryngotracheitis with special focus to south america: an update

Avian Infectious laryngotracheitis (AILT) is a respiratory tract disease of great importance because it causes significant economic losses in the poultry industry around the world. It is caused by a Gallid herpesvirus type 1, a member of the genus Iltovirus. The target system for Avian Infectious Laryngotracheitis virus (AILTV) infections is the respiratory system, and the main organ in which the virus remains latent is the trigeminal ganglia. However, the virus has demonstrated tropism for other organs besides the respiratory tract. The main transmission routes are ocular and respiratory. Infected birds with clinical symptoms are main sources of transmission, but birds with latent infections, litter, and contaminated fomites may also transmit the virus. Clinical signs usually appear 6-12 days after natural exposure and may be moderate or severe. The causative agent of this disease can be propagated in chorioallantoic membrane (CAM) of developing chicken embryos and replicate in mature chicken kidney cells, as well as in a variety of epithelial chick embryo cells, such as kidneys, liver and lungs. There are several procedures for the diagnosis of ILT such as the observation of clinical signs, the detection of gross and histopathological lesions, and the use of molecular techniques, including RFLP, polymerase chain reaction (PCR), real-time PCR, and loop-mediated isothermal amplification. Vaccination with different types of vaccine provides a good expectation on disease control, such as vaccines produced in chicken-embryo-origin (CEO), tissue-culture-origin (TCO), and recombinant vaccines. However, in endemic areas, biosecurity measures and best management practices are important for the control of the disease. It is distributed worldwide and, in South America, it has been reported in Brazil, Peru, Ecuador, Bolivia, and Argentina causing great economic losses.

Epidemiology of avian infectious laryngotracheitis with special focus to south america: an update

Avian Infectious laryngotracheitis (AILT) is a respiratory tract disease of great importance because it causes significant economic losses in the poultry industry around the world. It is caused by a Gallid herpesvirus type 1, a member of the genus Iltovirus. The target system for Avian Infectious Laryngotracheitis virus (AILTV) infections is the respiratory system, and the main organ in which the virus remains latent is the trigeminal ganglia. However, the virus has demonstrated tropism for other organs besides the respiratory tract. The main transmission routes are ocular and respiratory. Infected birds with clinical symptoms are main sources of transmission, but birds with latent infections, litter, and contaminated fomites may also transmit the virus. Clinical signs usually appear 6-12 days after natural exposure and may be moderate or severe. The causative agent of this disease can be propagated in chorioallantoic membrane (CAM) of developing chicken embryos and replicate in mature chicken kidney cells, as well as in a variety of epithelial chick embryo cells, such as kidneys, liver and lungs. There are several procedures for the diagnosis of ILT such as the observation of clinical signs, the detection of gross and histopathological lesions, and the use of molecular techniques, including RFLP, polymerase chain reaction (PCR), real-time PCR, and loop-mediated isothermal amplification. Vaccination with different types of vaccine provides a good expectation on disease control, such as vaccines produced in chicken-embryo-origin (CEO), tissue-culture-origin (TCO), and recombinant vaccines. However, in endemic areas, biosecurity measures and best management practices are important for the control of the disease. It is distributed worldwide and, in South America, it has been reported in Brazil, Peru, Ecuador, Bolivia, and Argentina causing great economic losses.(AU)

Vigilância epidemiológica dos vírus da influenza aviária, da doença de Newcastle e da laringotraqueíte infecciosa das galinhas em propriedades avícolas

A sanidade avícola é necessária para garantir a posição de destaque mundial da avicultura brasileira. O objetivo deste trabalho foi realizar a vigilância epidemiológica do vírus da IA (VIA) e do vírus da DNC (VDNC) em 2017 e 2018, além de avaliar a circulação de estirpes do vírus de laringotraqueíte infecciosa das galinhas (VLTI) nas granjas de postura comercial em duas regiões quarentemadas (Bastos e Guatapará) entre 2010 e 2018. Diferentes programas vacinais contra a LTI foram implementados na região, com a substituição de vacinas vivas atenuadas por recombinantes em 2012 (Bastos) e 2013 (Guatapará). Para a vigilância de VIA e VDNC foram coletados pools de suabes (traqueais e cloacais, n=220) de aves de subsistências localizadas no entorno de propriedades avícolas de reprodutoras. Essas amostras foram testadas pela reação de transcrição reversa-PCR em tempo real (RRT-PCR). O estudo longitudinal de VLTI coletou amostras de pools de suabes orofaríngeos de aves localizadas em Bastos (n=364) e Guatapará (n=214) para a detecção de VLTI por PCR. O sequenciamento de DNA dos genes ICP4, TK e genomas completos das amostras positivas foram realizados usando sequenciamento Sanger e sequenciamento de última geração (NGS). As bibliotecas de DNA foram preparadas com o kit Nextera DNA Flex Library Prep e sequenciadas com o sequenciador MiSeq. Análise filogenética e as identidades genéticas foram inferidas com as sequências obtidas. Nosso estudo não detectou VIA e do VDNC em nenhuma das amostras no período avaliado. O VLTI foi detectado em 11,85% e 12,6% das amostras testadas de Bastos em 2013 e 2018. A região de Guatapará apresentou a maior taxa de detecção (60,5%) em 2010. A taxa de detecção de amostras testadas da região de Guatapará variou de 12,2% e 21,7% após 2013. Análise filogenética do gene ICP4 agrupou as sequências das amostras de Bastos com vacinas de CEO e outras sequências de Bastos detectadas em anos anteriores, com 99% da identidade genética entre estas sequências. As análises filogenéticas dos genes ICP4, TK e do genoma completo agruparam as amostras de Guatapará separadamente das amostras vacinais. A sequência completa do genoma de uma amostra obtida de aves sintomáticas sem vacinação na região de Guatapará em 2010 (IB8098) teve 152.985 nucleotídeos, com 4755 leituras e 98,2% de cobertura. A análise filogenética agrupou a amostra IB8098 com outras estirpes virulentas, como da Rússia (MF405079). As identidades genéticas apresentaram 99,9%, quando comparadas à estirpe russa e 99,6% em relação às vacinais (CEO e TCO). Nosso estudo mostrou o VLTI ainda está presente nas regiões de Bastos e Guatapará, apesar das medidas de controle e vacinação vigentes. Os programas de vacinação nas regiões quarentenadas diminuíram a circulação do vírus com um leve aumento da detecção do VLTI com a utilização apenas das vacinas recombinantes. Esse é o primeiro relato da obtenção do genoma completo de VLTI no Brasil e sua caracterização genômica desde o início da circulação do vírus na região há pelo menos dez anos. O monitoramento contínuo da área é essencial para auxiliar as ações da defesa agropecuária, avaliar as medidas de vacinação implementadas contribuindo para a melhoria da sanidade avícola.

Poultry health is important to ensure the world's leading position in Brazilian poultry production. The present study performed the epidemiological surveillance of avian influenza virus (AIV) and Newcastle disease virus (NDV) in 2017 and 2018; the circulation of virulent avian infection laryngotracheitis virus (ILTV) strains was also evaluated in commercial layer farms from two quarantined regions (Bastos and Guatapara) between 2010 and 2018. Different vaccine programs against ILT were established in the area, and a replacement of live vaccines by recombinant vaccines was done in 2012 (Bastos) and 2013 (Guatapara). For the AIV and NDV surveillance, swab pool samples (trachea and cloacal, n=220) were collected from backyard birds located closed to breeder farms. Real-time reverse-transcription polymerase chain reaction (RRT-PCR) tested these samples. The longitudinal study collected oropharyngeal swab pool samples from commercial layer chickens located in Bastos (n=364) and Guatapara (n=214) for virus ILTV detection by PCR. DNA sequencing from the ICP4, TK genes and complete genomes was performed in positive samples using Sanger sequencing and next-generation sequencing (NGS). DNA libraries were prepared with the Nextera DNA Flex Library Prep kit, sequenced with the MiSeq sequencer for NGS. Phylogenetic analysis and genetic identities were inferred using the obtained sequences. AIV and NDV were not detected in any sample during the study. ILTV was detected in 11.85% and 12.6% of tested samples from Bastos in 2013 and 2018. Guatapara region had the highest detection rate (60.5%) in 2010. The detection rate from tested samples of the Guatapara region ranged from 12.2% and 21.7% after 2013. Phylogenetic analysis based on ICP4 gene grouped sequences from Bastos samples with CEO vaccines and other sequences detected previously in Bastos, with 99% of nucleotide identity among them. Phylogenetic analysis based on ICP4, TK, and complete genomes grouped sequences from Guatapara separately from vaccine strains. The complete genome sequence of a sample (IB8098), collected from layer chickens displaying clinical signs without vaccination in the Guatapara region in 2010, had 152,985 nucleotides, with 4,755 reads and 98,2% coverage. The phylogenetic analysis grouped the IB8098 sample with other virulent ILTV strains, such as from Russia (MF405079). Nucleotide identities were 99.9% compared to the Russian, and 99.6% to vaccine strains (CEO and TCO). Our study showed the ILTV is still present in the Bastos and Guatapara regions, despite the control and vaccination measures. The vaccination programs in the quarantine regions decreased the virus circulation, slightly increasing when only recombinant vaccines were applied. That is the first report of complete genome ILTV sequences in Brazil and its genomic characterization, after its circulation in the region for at least ten years. Continuous monitoring in poultry flocks is essential to help the poultry health measures, evaluate the vaccination programs placed in the area contributing to the Brazilian poultry health plans improvement.

Occurrence of infectious Laryngotracheitis Virus (ILTV) in 2009-2013 in the State of São Paulo - Brazil

Infectious laryngotracheitis is a very important respiratory disease because it causes significant economic losses in the poultry industry. The target of ILTV infections is the respiratory system, and the main organ in which the virus remains latent is the trigeminal ganglia. However, the virus has demonstrated tropism for other organs as well. The present study was conducted to determine the presence of Infectious Laryngotracheitis Virus (ILTV) in the state of São Paulo. Samples submitted to LABOR- USP during the last four years (2009-2013) analyzed by a nested/PCR technique. Out of the 682 samples from layers tested for LTIV, 12.46 % were positive, and derived from in both traditional (trachea and trigeminal ganglion) and untraditional (cecal tonsils, digestive tract and kidneys) organs utilized for ILTV diagnosis. The present work showed that ILTV is circulating in commercial layer flocks in São Paulo State, and that the LTIV is present in other organs in addition to the respiratory tract and trigeminal ganglion; however, it was not determined if the circulating virus is a vaccinal or field strain.

Occurrence of infectious Laryngotracheitis Virus (ILTV) in 2009-2013 in the State of São Paulo - Brazil

Infectious laryngotracheitis is a very important respiratory disease because it causes significant economic losses in the poultry industry. The target of ILTV infections is the respiratory system, and the main organ in which the virus remains latent is the trigeminal ganglia. However, the virus has demonstrated tropism for other organs as well. The present study was conducted to determine the presence of Infectious Laryngotracheitis Virus (ILTV) in the state of São Paulo. Samples submitted to LABOR- USP during the last four years (2009-2013) analyzed by a nested/PCR technique. Out of the 682 samples from layers tested for LTIV, 12.46 % were positive, and derived from in both traditional (trachea and trigeminal ganglion) and untraditional (cecal tonsils, digestive tract and kidneys) organs utilized for ILTV diagnosis. The present work showed that ILTV is circulating in commercial layer flocks in São Paulo State, and that the LTIV is present in other organs in addition to the respiratory tract and trigeminal ganglion; however, it was not determined if the circulating virus is a vaccinal or field strain.(AU)

Laringotraqueíte infecciosa das aves e diagnóstico diferencial de outras doenças respiratórias

A laringotraqueíte infecciosa (LTI)é uma infecção virale de distribuição cosmopolita que acomete especialmente o trato respiratório superior e a conjuntiva das aves comerciais 21,33.O vírus da laringotraqueíte pertence ao gênero Iltovirus,família Herpesviridae e subfamília Herpesvirinae. O vírus é taxonomicamente identificado como Gallid herpesvírus 1 (GaHV-1).

Laringotraqueíte infecciosa das aves e diagnóstico diferencial de outras doenças respiratórias

A laringotraqueíte infecciosa (LTI)é uma infecção virale de distribuição cosmopolita que acomete especialmente o trato respiratório superior e a conjuntiva das aves comerciais 21,33.O vírus da laringotraqueíte pertence ao gênero Iltovirus,família Herpesviridae e subfamília Herpesvirinae. O vírus é taxonomicamente identificado como Gallid herpesvírus 1 (GaHV-1).

Pathological, immunohistochemical, and molecular findings in commercial laying hens and in backyard chickens naturally infected with the infectious laryngotracheitis virus

Seventy-eight chickens from a very high poultry density (approximately eight million) region and twelve backyard chickens from neighboring areas were analyzed by histopathology and additional techniques for the presence of the infectious laryngotracheitis virus. The virus distribution was determined in different tissues using immunohistochemistry (IHC) and polymerase chain reaction (PCR). The disease was histopathologically diagnosed in 41.0% (32/78) of the commercial layers. Lesions were mainly characterized by syncytial cells with eosinophilic intranuclear inclusion body formed from the hyperplastic epithelium of the upper respiratory tract, primary and secondary bronchi, and conjunctiva. IHC showed 70% (21/30) positive signal in the larynx/trachea and, 53.8% (14/26) in the lungs, either in epithelial cells or syncytia. In the turbinates and paranasal sinuses, 29.6% (8/27) of samples showed positive signal. PCR detected the following gallid herpesvirus 1-positive percentages: conjunctiva 63.2% (31/49), lungs 57.6% (30/52), turbinates and paranasal sinuses 56% (28/50), and larynx/trachea 50% (39/78). IHC showed to be a useful additional tool for definitive ILT diagnosis, especially during the subacute phase of the disease when syncytial cells with intranuclear inclusion bodies are no longer observed. PCR using specific primers from ICP4 gene, generating a product of 237 base pairs, was sensitive for ILT diagnosis, and very useful for rapid detection of GaHV-1 in chickens. Fixed tissues allowing histopatological examination and detection of GaHV-1 by PCR, are a good option in areas where farms are located several hundred kilometers away from a diagnostic center, reducing problems with conservation of fresh samples and the risk of virus spread.

Pathological, immunohistochemical, and molecular findings in commercial laying hens and in backyard chickens naturally infected with the infectious laryngotracheitis virus

Seventy-eight chickens from a very high poultry density (approximately eight million) region and twelve backyard chickens from neighboring areas were analyzed by histopathology and additional techniques for the presence of the infectious laryngotracheitis virus. The virus distribution was determined in different tissues using immunohistochemistry (IHC) and polymerase chain reaction (PCR). The disease was histopathologically diagnosed in 41.0% (32/78) of the commercial layers. Lesions were mainly characterized by syncytial cells with eosinophilic intranuclear inclusion body formed from the hyperplastic epithelium of the upper respiratory tract, primary and secondary bronchi, and conjunctiva. IHC showed 70% (21/30) positive signal in the larynx/trachea and, 53.8% (14/26) in the lungs, either in epithelial cells or syncytia. In the turbinates and paranasal sinuses, 29.6% (8/27) of samples showed positive signal. PCR detected the following gallid herpesvirus 1-positive percentages: conjunctiva 63.2% (31/49), lungs 57.6% (30/52), turbinates and paranasal sinuses 56% (28/50), and larynx/trachea 50% (39/78). IHC showed to be a useful additional tool for definitive ILT diagnosis, especially during the subacute phase of the disease when syncytial cells with intranuclear inclusion bodies are no longer observed. PCR using specific primers from ICP4 gene, generating a product of 237 base pairs, was sensitive for ILT diagnosis, and very useful for rapid detection of GaHV-1 in chickens. Fixed tissues allowing histopatological examination and detection of GaHV-1 by PCR, are a good option in areas where farms are located several hundred kilometers away from a diagnostic center, reducing problems with conservation of fresh samples and the risk of virus spread.

Pathological, immunohistochemical, and molecular findings in commercial laying hens and in backyard chickens naturally infected with the infectious laryngotracheitis virus

Seventy-eight chickens from a very high poultry density (approximately eight million) region and twelve backyard chickens from neighboring areas were analyzed by histopathology and additional techniques for the presence of the infectious laryngotracheitis virus. The virus distribution was determined in different tissues using immunohistochemistry (IHC) and polymerase chain reaction (PCR). The disease was histopathologically diagnosed in 41.0% (32/78) of the commercial layers. Lesions were mainly characterized by syncytial cells with eosinophilic intranuclear inclusion body formed from the hyperplastic epithelium of the upper respiratory tract, primary and secondary bronchi, and conjunctiva. IHC showed 70% (21/30) positive signal in the larynx/trachea and, 53.8% (14/26) in the lungs, either in epithelial cells or syncytia. In the turbinates and paranasal sinuses, 29.6% (8/27) of samples showed positive signal. PCR detected the following gallid herpesvirus 1-positive percentages: conjunctiva 63.2% (31/49), lungs 57.6% (30/52), turbinates and paranasal sinuses 56% (28/50), and larynx/trachea 50% (39/78). IHC showed to be a useful additional tool for definitive ILT diagnosis, especially during the subacute phase of the disease when syncytial cells with intranuclear inclusion bodies are no longer observed. PCR using specific primers from ICP4 gene, generating a product of 237 base pairs, was sensitive for ILT diagnosis, and very useful for rapid detection of GaHV-1 in chickens. Fixed tissues allowing histopatological examination and detection of GaHV-1 by PCR, are a good option in areas where farms are located several hundred kilometers away from a diagnostic center, reducing problems with conservation of fresh samples and the risk of virus spread.

Pathological, immunohistochemical, and molecular findings in commercial laying hens and in backyard chickens naturally infected with the infectious laryngotracheitis virus

Seventy-eight chickens from a very high poultry density (approximately eight million) region and twelve backyard chickens from neighboring areas were analyzed by histopathology and additional techniques for the presence of the infectious laryngotracheitis virus. The virus distribution was determined in different tissues using immunohistochemistry (IHC) and polymerase chain reaction (PCR). The disease was histopathologically diagnosed in 41.0% (32/78) of the commercial layers. Lesions were mainly characterized by syncytial cells with eosinophilic intranuclear inclusion body formed from the hyperplastic epithelium of the upper respiratory tract, primary and secondary bronchi, and conjunctiva. IHC showed 70% (21/30) positive signal in the larynx/trachea and, 53.8% (14/26) in the lungs, either in epithelial cells or syncytia. In the turbinates and paranasal sinuses, 29.6% (8/27) of samples showed positive signal. PCR detected the following gallid herpesvirus 1-positive percentages: conjunctiva 63.2% (31/49), lungs 57.6% (30/52), turbinates and paranasal sinuses 56% (28/50), and larynx/trachea 50% (39/78). IHC showed to be a useful additional tool for definitive ILT diagnosis, especially during the subacute phase of the disease when syncytial cells with intranuclear inclusion bodies are no longer observed. PCR using specific primers from ICP4 gene, generating a product of 237 base pairs, was sensitive for ILT diagnosis, and very useful for rapid detection of GaHV-1 in chickens. Fixed tissues allowing histopatological examination and detection of GaHV-1 by PCR, are a good option in areas where farms are located several hundred kilometers away from a diagnostic center, reducing problems with conservation of fresh samples and the risk of virus spread.

Pathological, immunohistochemical, and molecular findings in commercial laying hens and in backyard chickens naturally infected with the infectious laryngotracheitis virus

Seventy-eight chickens from a very high poultry density (approximately eight million) region and twelve backyard chickens from neighboring areas were analyzed by histopathology and additional techniques for the presence of the infectious laryngotracheitis virus. The virus distribution was determined in different tissues using immunohistochemistry (IHC) and polymerase chain reaction (PCR). The disease was histopathologically diagnosed in 41.0% (32/78) of the commercial layers. Lesions were mainly characterized by syncytial cells with eosinophilic intranuclear inclusion body formed from the hyperplastic epithelium of the upper respiratory tract, primary and secondary bronchi, and conjunctiva. IHC showed 70% (21/30) positive signal in the larynx/trachea and, 53.8% (14/26) in the lungs, either in epithelial cells or syncytia. In the turbinates and paranasal sinuses, 29.6% (8/27) of samples showed positive signal. PCR detected the following gallid herpesvirus 1-positive percentages: conjunctiva 63.2% (31/49), lungs 57.6% (30/52), turbinates and paranasal sinuses 56% (28/50), and larynx/trachea 50% (39/78). IHC showed to be a useful additional tool for definitive ILT diagnosis, especially during the subacute phase of the disease when syncytial cells with intranuclear inclusion bodies are no longer observed. PCR using specific primers from ICP4 gene, generating a product of 237 base pairs, was sensitive for ILT diagnosis, and very useful for rapid detection of GaHV-1 in chickens. Fixed tissues allowing histopatological examination and detection of GaHV-1 by PCR, are a good option in areas where farms are located several hundred kilometers away from a diagnostic center, reducing problems with conservation of fresh samples and the risk of virus spread.

Pathological, immunohistochemical, and molecular findings in commercial laying hens and in backyard chickens naturally infected with the infectious laryngotracheitis virus

Seventy-eight chickens from a very high poultry density (approximately eight million) region and twelve backyard chickens from neighboring areas were analyzed by histopathology and additional techniques for the presence of the infectious laryngotracheitis virus. The virus distribution was determined in different tissues using immunohistochemistry (IHC) and polymerase chain reaction (PCR). The disease was histopathologically diagnosed in 41.0% (32/78) of the commercial layers. Lesions were mainly characterized by syncytial cells with eosinophilic intranuclear inclusion body formed from the hyperplastic epithelium of the upper respiratory tract, primary and secondary bronchi, and conjunctiva. IHC showed 70% (21/30) positive signal in the larynx/trachea and, 53.8% (14/26) in the lungs, either in epithelial cells or syncytia. In the turbinates and paranasal sinuses, 29.6% (8/27) of samples showed positive signal. PCR detected the following gallid herpesvirus 1-positive percentages: conjunctiva 63.2% (31/49), lungs 57.6% (30/52), turbinates and paranasal sinuses 56% (28/50), and larynx/trachea 50% (39/78). IHC showed to be a useful additional tool for definitive ILT diagnosis, especially during the subacute phase of the disease when syncytial cells with intranuclear inclusion bodies are no longer observed. PCR using specific primers from ICP4 gene, generating a product of 237 base pairs, was sensitive for ILT diagnosis, and very useful for rapid detection of GaHV-1 in chickens. Fixed tissues allowing histopatological examination and detection of GaHV-1 by PCR, are a good option in areas where farms are located several hundred kilometers away from a diagnostic center, reducing problems with conservation of fresh samples and the risk of virus spread.(AU)

Laringotraqueite Infecciosa das Aves: Análise das Ações do Serviço Veterinário Oficial para Controle de Foco em Granjas de Postura do Estado de Minas Gerais

Em Minas Gerais, a LTI em galinhas foi notificada pela primeira vez em novembro de 2010, na região denominada Terras Altas da Mantiqueira e desde então o controle da doença tem sido fiscalizado pelo Instituto Mineiro de Agropecuária. Para verificar a eficiência deste controle, o Serviço Veterinário Oficial realizou inquéritos soroepidemiológicos nos seguintes períodos: novembro de 2016, outubro de 2017 e maio de 2018. De forma simultânea, foram avaliadas as taxas de mortalidade e as medidas de biosseguridade implementadas nas granjas da região interditada. Utilizaram-se diferentes técnicas para identificar a doença (sinais clínicos, macroscopia e histopatologia) e técnicas moleculares para identificar a infecção. As granjas da região interditada que possuem maior capacidade de alojamento e consequentemente maior fluxo de veículos e de pessoas foram relacionadas à ocorrência de surtos de LTI, apesar de medidas mínimas de biosseguridade terem sido implementadas nestas granjas. A doença não foi diagnosticada nas granjas menores e mais isoladas, apesar dos resultados demonstrarem a atividade do vírus nestas explorações. As taxas de mortalidade tiveram valores acima do esperado, entretanto este aumento não foi atribuído exclusivamente à LTI. Amostras de aves alojadas em granjas externas à região do foco também foram analisadas para verificar a ocorrência da doença fora da área interditada. As medidas de restrição do trânsito de aves e de esterco impediram, até o momento, a disseminação do vírus ou da doença para granjas poedeiras de outras regiões do estado de MG, uma vez que os dados clínicos e os testes moleculares e sorológicos foram negativos para a doença e vírus, respectivamente. Por fim, os resultados deste estudo indicaram que o vírus está ativo na região de controle oficial, e que as medidas tomadas pelo SVO foram capazes de restringir o foco nesta região e melhorar a biosseguridade das granjas. Porém, o controle deve ser mantido por meio de uma vigilância rigorosa e exigências na melhoria na biosseguridade para mitigar o risco de ocorrência da laringotraqueite nessa e em outras regiões.

ILT in chickens was notified for the first time in November 2010, in the region named Terras Altas da Mantiqueira, Minas Gerais state, Brazil. Since then, the control of the disease has been monitored by the Instituto Mineiro de Agropecuária. To verify the effectiveness of this control, the Official Veterinary Service carried out epidemiological serum surveys in the following periods: November 2016, October 2017 and May 2018. Simultaneously, mortality rates and biosecurity measures on farms region were evaluated. Different techniques were used to identify the disease (clinical signs, macroscopy and histopathology) and molecular techniques to identify the infection. The farms in the quarantine region that have greater housing capacity and consequently greater flow of vehicles and people, were related to the occurrence of ILT outbreaks, although minimal biosecurity measures were implemented in these farms. The disease was not diagnosed in the smaller and more isolated farms, although the results demonstrated the existence of the virus in these farms. The mortality rates were higher than expected, however, this increase is not attributed exclusively to ILT. Samples of chickens housed on farms outside of the quarantine region were also examined to verify the occurrence of the disease outside to the quarantine area. Measures to restrict chickens and poultry litter traffic have so far prevented the spread of the virus or disease to laying farms in other regions of the state of Minas Gerais, since clinical and molecular and/or serological tests were negative for the disease and virus, respectively. Finally, the results of this study indicated that the virus is active in the official control region, and the measures taken by the SVO have been able to prevent outbreak outside of this region and improve the biossecurity of farms. However, control should be maintained through strict surveillance and demands for improved biosecurity to mitigate the risk of laryngotracheitis occurring in this and other regions