Bacteriological Quality of Table Eggs in Moroccan Formal and Informal Sector.

Eggs constitute an important part of the Moroccan diet. However, contaminated eggs can cause a serious public health problem if consumed undercooked, uncooked, or used in unpasteurized egg foodstuffs. This study was carried out to evaluate the microbial contents of eggs according to their sales sector in Morocco. For that, a total of 1770 eggs were collected from January to September 2021 from formal markets (refrigerated eggs from large shopping centers) and informal markets (eggs at ambient temperature from ambulatory sellers, street vendors, kiosks, and neighborhood stores) and transferred to the Avian Pathology Unit at Hassan II Agronomic and Veterinary Institute. The eggshells and their contents were tested separately; swabs of eggshells were used to inoculate Mac-Conkey agar, while the egg contents were cultured on Mac-Conkey and Mannitol salt agar, then standard microbiological tests were performed to identify the isolated organisms. The results showed that informal eggs were more contaminated (87%) than formal eggs (48) (p < 0.05). The bacteria isolated from the eggshells (informal and formal) were Enterobacter agglomerans (59% and 21%), Klebsiella spp. (24% and 4%), Enterobacter cloacae (17% and 8%), E. coli (9% and 1%), Serratia spp. (9% and1%), Pseudomonas aeruginosa (9% and 1%), Shigella spp. (5% and 0%), Salmonella enteritidis (0% and 2%), Proteus spp. (4% and 0%), Enterobacter sakazakii (2% and 0%), Rahnella aquatilis (1% and 0%), and Staphylococcus aureus (0% and 1%). For the egg-contents, the detected bacteria (informal and formal) were Enterobacter agglomerans (14% and 28%), Klebsiella spp. (7% and 6%), Staphylococcus aureus (6% and 1%), Enterobacter cloacae (4% and 4%), E. coli (4%, 1%), Shigella spp. (4%, 0%), Acinetobacter baumannii (3% and 1%), Salmonella enteritidis (2% and 0%), Serratia spp. (1% and 6%), Proteus spp. (1% and 3%), and Enterobacter sakazakii (1% and 0%). We conclude that eggs might be contaminated with several bacteria and can constitute a public health threat in Morocco.

Detection of Sutterella spp. in Broiler Liver and Breast.

Sutterella sp. is a gram-negative, microaerophilic bacterium that is particularly resistant to bile acids. It has recently been associated with several human pathologies such as inflammatory bowel disease, asthma, diabetes, and autism. Indeed, susceptibility patterns to ciprofloxacin and erythromycin, combined with resistance to metronidazole, indicate that Sutterella wadsworthensis patterns are closer to those of Campylobacter. The objective of this study is to identify, for the first time, Sutterella spp. in the liver and breast of broiler chickens by quantitative real-time PCR (qPCR). Liver, breast, and cecal content samples were taken from 25 birds and frozen at -20°C until analyzed. The main results showed that Sutterella sp. is part of the cecal microbiota of 48% of the birds and present in the liver and breast of, respectively 20 and 40% of the chicks with a variable Cq. We, therefore, conclude that Sutterella sp. exists in poultry and poultry meat and that foodstuffs of poultry origin might be considered as a potential source of contamination for humans.

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.

First Report of Isolation of Gallibacterium anatis from Layer Chickens in Morocco with Decrease in Laying Performance.

Gallibacterium is a genus of the family of Pasteurellaceae. It is well known as a commensal inhabitant of the respiratory and reproductive tract of healthy chickens. But in the last years, Gallibacterium anatis is increasingly reported in field cases with a decrease in laying performance due to infections of the reproductive tract. The aim of the present study was to investigate the implication of G. anatis infection in layer flocks facing a decrease in laying performance in Morocco. Birds were received from five different laying hen farms in two regions in Morocco showing a drop of egg production. Necropsy revealed 46.1 % (24/52) of sampled birds showed variable lesions in ovaries, salpinx, and trachea. In fact, 24 birds were affected by salpingitis, 18 by oophoritis, and 11 birds by atrophy of ovaries. Furthermore, tracheitis was observed in 24 birds. Bacteriological investigation was done from different organs, and G. anatis was found in ovaries (n = 20), trachea (n = 17), and cloaca (n = 3). Identification was based on growth morphology, Gram staining, and biochemical properties. Additionally, polymerase chain reaction test using specific primers for the genus identification was carried out. All isolates showed bands of 925 bp specific for G. anatis expressing the virulent toxin GtxA. Antibiotic resistance testing was performed and revealed that isolates were sensitive to enrofloxacin, florfenicol, and gentamycin but resistant to ampicillin, erythromycin, oxytetracycline, and sulfamethoxazole-trimethoprim. The present study is the first report of G. anatis in Morocco, demonstrating the need for further epidemiologic investigations as well as in regard to antibiotic resistance development.

Primer informe de aislamiento de Gallibacterium anatis de gallinas de postura en Marruecos con disminución en la eficiencia de la postura. Gallibacterium es un género de la familia Pasteurellaceae. Es bien conocido como un habitante comensal del tracto respiratorio y reproductivo de pollos sanos. Pero en los últimos años, han aumentado los reportes de Gallibacterium anatis en casos de campo con una disminución en la eficiencia de la postura debido a infecciones del tracto reproductivo. El objetivo del presente estudio fue investigar la implicación de la infección por G. anatis en parvadas de gallinas de postura que presentaban una disminución en la eficiencia de la postura en Marruecos. Se recibieron aves de cinco granjas diferentes de gallinas postura en dos regiones de Marruecos, mostrando una caída en la producción de huevos. La necropsia reveló que 46.1% (24/52) de las aves muestreadas mostraron lesiones variables en ovarios, oviducto y tráquea. De hecho, 24 aves mostraron salpingitis, 18 ooforitis y 11 aves atrofia de ovarios. Además, se observó traqueítis en 24 aves. Se realizó la investigación bacteriológica de diferentes órganos, y se encontró G. anatis en ovarios (n = 20), tráquea (n = 17) y cloaca (n = 3). La identificación se basó en la morfología del crecimiento, en la tinción de Gram y en las propiedades bioquímicas. Además, se realizó una prueba de reacción en cadena de la polimerasa utilizando iniciadores específicos para la identificación del género. Todos los aislamientos mostraron bandas de 925 pares de bases específicas para G. anatis que expresaban la toxina virulenta GtxA. Se realizaron pruebas de resistencia a los antibióticos y revelaron que los aislamientos eran sensibles a enrofloxacina, florfenicol y gentamicina, pero resistentes a ampicilina, eritromicina, oxitetraciclina y sulfametoxazol-trimetoprima. El presente estudio es el primer informe de G. anatis en Marruecos, que demuestra la necesidad de más investigaciones epidemiológicas, así como en relación con el desarrollo de resistencia a los antibióticos.

Efficacy of Massachusetts and 793B Vaccines Against Infectious Bronchitis Moroccan-Italy 02 Virus in Specific-Pathogen-Free Chickens and Commercial Broilers.

The ability of commercial vaccines H120 and 4/91 to protect against Moroccan-Italy 02 infectious bronchitis virus (Mor-It02) was investigated in specific-pathogen-free (SPF) chickens and commercial broiler chickens. Commercial broiler chicks (Experiment 1) were vaccinated at the hatchery with H120 vaccine at Day 1, and challenged at Day 21 with 104 50% egg-infective dose (EID50) of Mor-It02. All chicks were observed daily for clinical signs attributable to Mor-It02 infection during the 10 days postchallenge (pc). At 5 and 10 days pc, chicks were humanely sacrificed for necropsy examination, and tissues were collected for histopathology evaluation. To better understand the findings on commercial broilers, day-old SPF chicks were divided into five groups in a second experiment: Group Mass/4-91, vaccinated with H120 and 4/91 respectively at Days 1 and 15 of age; Group Mass/Mass, vaccinated by H120 at Days 1 and 15; Group Mass, vaccinated with H120 at Day 1; Group NV, kept unvaccinated; and Group NC, kept as a negative control (unchallenged). At Day 24 of age, Groups Mass/4-91, Mass/Mass, Mass, and NV were challenged with 104 EID50 of Mor-It02. In both experiments, blood samples were collected at different periods for serologic analyses. Oropharyngeal swabs were collected for virus detection by reverse-transcription PCR. In Experiments 1 and 2, respiratory signs started as early as 24 hr pc and maximum severity was observed on Days 3 and 4 pc. The viral shedding rate was significantly lower in Group Mass/4-91 compared to other challenged groups. Serologic analysis in both experiments showed that the sera of challenged group exhibited significantly higher antibody titers than sera collected before challenge. Histopathologic investigations in SPF birds showed deciliation and hyperplasia in Group NV and less-pronounced lesions in Groups Mass/Mass and Mass. In commercial broilers vaccinated with H120 alone, hyperplasia and deciliation were observed in 90% of the tracheas. These experiments illustrated that Mor-It02 is pathogenic for chickens and a combination of live H120 and 4/91 vaccines given respectively at Day 1 and Day 15 of age confer a good protection against Mor-It02.

First outbreaks and phylogenetic analyses of avian influenza H9N2 viruses isolated from poultry flocks in Morocco.

BACKGROUND: H9N2 avian influenza viruses continue to spread in poultry and wild birds worldwide. Morocco just faced its first H9N2 influenza virus outbreaks early 2016 affecting different types of poultry production. After its introduction, the virus spread very rapidly throughout the country. METHODS: Samples were collected from 11 chicken flocks with high morbidity and mortality rates. Four viruses were successfully isolated from broiler chickens and one from broiler breeders and fully sequenced. RESULTS: Phylogenetic and molecular markers analyses showed the Moroccan viruses belonged to the G1 lineage and likely originated from the Middle East. As known for H9N2 viruses, the Moroccanisolates possess several genetic markers that enhance virulence in poultry and transmission to humans. CONCLUSION: The present study demonstrated that under field conditions H9N2 could have a devastating effect on egg production and mortalities and highlighted a lack of surveillance data on the pathogen in the region.