Avian Influenza a H9N2 Viruses in Morocco, 2018-2019.

Low pathogenic H9N2 avian influenza (LPAI H9N2) is considered one of the most important diseases found in poultry (broiler, laying hens, breeding chickens, and turkeys). This infection causes considerable economic losses. The objective of this work was to monitor and assess the presence of avian influenza virus (AIV) H9N2 in eight different regions of Morocco using real-time RT-PCR, and to assess the phylogenetic and molecular evolution of the H9N2 viruses between 2016 and 2019. Field samples were collected from 108 farms suspected of being infected with LPAI H9N2 virus. Samples were analyzed using H9N2-specific real-time RT-PCR. Highly positive samples were subjected to virus isolation and seven isolates were fully sequenced. Low pathogenic H9N2 avian influenza virus was introduced in Morocco in 2016. We show that in 2018-2019, the virus was still present irrespective of vaccination status. Phylogenetic and molecular analyses showed mutations related to virulence, although our viruses were related to 2016 Moroccan viruses and grouped in the G1 lineage. Specific amino acid substitutions were identified in Moroccan H9N2 viruses that are believed to lead to increased resistance to antiviral drugs.

Protective Efficacy Evaluation of Four Inactivated Commercial Vaccines Against Low Pathogenic Avian Influenza H9N2 Virus Under Experimental Conditions in Broiler Chickens.

Avian influenza vaccines are commonly used in the poultry industry. The objective of this study was to compare, under experimental conditions, the protective efficacy of four imported commercial inactivated H9N2 vaccines (A, B, C, and D) in broiler chickens. A total of 150 one-day-old chicks were divided into six groups: four experimental groups, each containing 30 chicks, received one of the vaccines (A, B, C, or D) delivered in a 0.3-ml dose subcutaneously at 1 day of age, whereas the control, Group T, was not vaccinated but challenged and Group E was kept unvaccinated and unchallenged. At 21 days postvaccination, Groups A, B, C, D, and T were challenged with 107 embryo infective dose 50% of A/Chicken/Morocco/01/2016 (H9N2). All chicks were observed daily for clinical signs during the 12 days postchallenge (dpc). At 5 and 12 dpc, chicks were euthanatized for necropsy examination. Blood samples were collected weekly for serologic analysis and oropharyngeal swabs were collected for virus detection by real-time RT-PCR. Respiratory signs started at 48 hr pc and maximum severity was observed on 9 dpc. Chiefly, the birds vaccinated with vaccine B showed significantly more respiratory signs than did their counterparts. Serologic analysis revealed that the sera of Groups A and D birds showed a decrease in antibody (Ab) levels up to day 26; then a slight increase of Ab level was observed until day 31, while Group B and C birds showed a stabilization of the titers from day 21 until the end of the experiment. The viral shedding rate was significantly lower in Groups C and A (40%-50% of the birds shed virus for <7 days) compared with other challenged groups (60%-75% of the birds shed virus for ≥9 days). This experiment illustrated that vaccination applied on the first day in the hatchery with the four vaccines tested did not provide an acceptable protection against H9N2 in comparison with the controls that did not receive any vaccine. However, at first glance, we might favor vaccines A and C for their ability to reduce and shorten viral shedding as compared with vaccines B and D.

Evaluación de la eficacia protectora de cuatro vacunas comerciales inactivadas contra el virus de la influenza aviar H9N2 de baja patogenicidad bajo condiciones experimentales en pollos de engorde. Las vacunas contra la influenza aviar se utilizan comúnmente en la industria avícola. El objetivo de este estudio fue comparar, en condiciones experimentales, la eficacia protectora de cuatro vacunas H9N2 inactivadas comerciales importadas (A, B, C y D) en pollos de engorde. Un total de 150 pollitos de un día se dividieron en seis grupos: cuatro grupos experimentales, cada uno con 30 pollitos, recibieron una de las vacunas (A, B, C o D) administradas en una dosis de 0.3 ml por vía subcutánea al día. de edad, mientras que el control, Grupo T, que no fue vacunado y desafiado y el Grupo E que se mantuvo sin vacunar y sin desafiar. A los 21 días después de la vacunación, los Grupos A, B, C, D y T fueron desafiados con 107 dosis infecciosas de embriones al 50% del virus A/Chicken/Marruecos/01/2016 (H9N2). Todos los pollos fueron observados diariamente para detectar signos clínicos durante los 12 días posteriores al desafío (dpc). A los cinco y 12 días después del desafío, los polluelos fueron sacrificados humanitariamente para un examen de necropsia. Se recolectaron muestras de sangre semanalmente para análisis serológicos y se recolectaron hisopos orofaríngeos para la detección de virus mediante RT-PCR en tiempo real. Los signos respiratorios comenzaron a las 48 horas después del desafío y la severidad máxima se observó a los nueve días después del desafío. Principalmente, las aves vacunadas con la vacuna B mostraron significativamente más signos respiratorios que sus contrapartes. El análisis serológico reveló que los sueros de las aves de los Grupos A y D mostraron una disminución en los niveles de anticuerpos (Ab) hasta el día 26; luego se observó un ligero aumento del nivel de anticuerpos hasta el día 31, mientras que las aves de los Grupos B y C mostraron una estabilización de los títulos desde el día 21 hasta el final del experimento. La tasa de excreción viral fue significativamente menor en los Grupos C y A (40% -50% de las aves excretaron el virus durante <7 días) en comparación con otros grupos desafiados (60% -75% de las aves excretaron el virus durante ≥9 días). Este experimento ilustró que la vacunación aplicada el primer día en la incubadora con las cuatro vacunas probadas no proporcionó una protección aceptable contra el virus H9N2 en comparación con los controles que no recibieron ninguna vacuna. Sin embargo, a primera vista, podríamos favorecer las vacunas A y C por su capacidad para reducir y acortar la diseminación viral en comparación con las vacunas B y D.

Pathogenesis of Avian Influenza Virus Subtype H9N2 in Turkeys and Evaluation of Inactivated Vaccine Efficacy.

Avian influenza H9N2 viruses circulate in all types of poultry species, including turkeys, and cause significant losses for the poultry industry in many parts of the word. The aim of this study was to assess the pathogenesis of the Moroccan avian influenza virus (AIV) H9N2 under experimental conditions in turkeys and the protection efficacy of an inactivated commercial vaccine against AIV H9N2. Unvaccinated turkeys showed marked depression sinusitis, respiratory distress characterized by bronchiolar and tracheal rales of moderate severity, and a mortality rate of 50%. Postmortem examinations of dead and euthanatized birds revealed the presence of fibrinous tracheitis and airsacculitis lesions. Vaccination reduced the mortality rate to 20%. Vaccinated birds recovered at day 10 postchallenge, and only 12.5% (1/8) and 37.5% of birds still displayed fibrinous and nonfibrinous airsacculitis lesions, respectively, at day 15 postinoculation. Viral shedding in cloacal and tracheal swabs was lower in vaccinated than in control birds. Although viral RNA was detected in the cloacal swabs of all unvaccinated turkeys at day 3 postinoculation, only 50% of the vaccinated turkeys were positive for virus detection. At day 11 postinoculation, no viral RNA was detected in oropharyngeal swabs of vaccinated turkeys, whereas 40% of the unvaccinated turkeys were still shedding virus.

Artículo regular­Patogenia del subtipo H9N2 del virus de la influenza aviar en pavos y evaluación de la eficacia de una vacuna inactivada. Los virus de la influenza aviar H9N2 circulan en todo tipo de especies de aves comerciales, incluidos los pavos, y causan pérdidas significativas para la industria avícola en muchas partes del mundo. El objetivo de este estudio fue evaluar la patogenia del virus de la influenza aviar de Marruecos (AIV) H9N2 bajo condiciones experimentales en pavos y la eficacia de protección de una vacuna comercial inactivada contra el virus de la influenza aviar H9N2. Los pavos no vacunados mostraron una marcada sinusitis, depresión, dificultad respiratoria caracterizada por estertores bronquiolares y traqueales de severidad moderada y una tasa de mortalidad del 50%. Los exámenes post mortem de aves muertas y sacrificadas revelaron la presencia de traqueítis fibrinosa y aerosaculitis. La vacunación redujo la tasa de mortalidad al 30%. Las aves vacunadas se recuperaron en el día 10 después del desafío, y solo el 12.5% (1/8) y el 37.5% de las aves todavía mostraban aerosaculitis fibrinosa y no fibrinosa, respectivamente, el día 15 después de la inoculación. La diseminación viral en los hisopos cloacales y traqueales fue menor en las aves vacunadas que en las aves control. Aunque se detectó ARN viral en los hisopados cloacales de todos los pavos no vacunados en el día tres después de la inoculación, solo el 50% de los pavos vacunados dieron positivo para la detección del virus. En el día 11 después de la inoculación, no se detectó ARN viral en hisopados orofaríngeos de pavos vacunados, mientras que el 40% de los pavos no vacunados todavía estaban diseminando virus.

Determination of Zinc in Camel Skin Using Laser-Induced Breakdown Spectroscopy.

Zinc plays a major role in skin integrity, which can be affected by dromedary camels' hard life conditions. Deficiencies in some trace elements especially in zinc can explain susceptibility of this species to skin diseases. Compared with ruminants, camel is already known for his relatively low zincemia. In order to assess dromedary camels' skin zinc content, the present study was carried out in several provinces located in the south of Morocco where camel skin diseases are commonly observed. Zinc content in dromedary camel skin was determined using for the first time laser-induced breakdown spectroscopy (LIBS), method considered so far quick and simple with few or no sample processing. Collected data showed that zinc skin contents ranged between 115 ± 60 (for external side) and 94 ± 82 ppm (for internal side) with significant variability according to animals and to skin layers (external side versus internal side). Zinc skin content decreased from external to internal layers. Such preliminary results could be used to compare skin zinc nutritional level in healthy camels with those affected by skin diseases.