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Chicken anemia virus (CAV) is a widespread and economically significant pathogen in the poultry industry. In this study 110 samples were collected from various poultry farms in selected Egyptian provinces during 2021-2022 and were tested against CAV by Polymerase Chain Reaction (PCR), revealing 22 positive samples with 20% incidence rate. Full sequence analysis of five selected CAV strains revealed genetic variations in VP1, VP2, and VP3 genes. Phylogenetic analysis grouped the Egyptian strains with reference viruses, mainly in group II, while vaccines like Del-Rose were categorized in group III. Recombination events were detected between an Egyptian strain (genotype II) and the Del-Rose vaccine strain (genotype III), indicating potential recombination between live vaccine strains and field isolates. To evaluate pathogenicity, one Egyptian isolate (F883-2022 CAV) and Del-Rose vaccine were tested in Specific Pathogen Free (SPF) chicks. Chicks in the positive group displayed clinical symptoms, including weakness and stunted growth, with postmortem findings consistent with CAV infection. The vaccine group showed milder symptoms and less severe postmortem changes. This study provides important insights into the genetic diversity of CAV in selected Egyptian poultry farms showing recombination event between field strain and vaccine strains, highlighting the need for advanced vaccination programs, especially for broilers.
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Since the detection of antigenically atypical very virulent Infectious bursal disease viruses (vvIBDV) in Egypt in 1999, the country has been experiencing recurrent outbreaks with high mortality rates and typical gross lesions associated with typical vvIBDV. However, a significant change occurred in 2023, marked by a notable increase in reported subclinical IBDV cases. To evaluate the field situation, samples from 21 farms in 2023 and 18 farms from 2021 and 2022, all of which had experienced IBD outbreaks based on clinical diagnosis, were collected, and subjected to VP2-HVR sequencing. Phylogenetic analysis revealed that all samples collected in 2021 and 2022 clustered with classical virulent strains and vvIBDV. In 2023, one sample clustered with the Egyptian vvIBDV, another with classical virulent IBDV, and the rest with the novel variant IBDV (nVarIBDV) circulating in China. The alignment of deduced amino acid sequences for VP2 showed that all Egyptian classic virulent strains were identical to the Winterfield or Lukert strains, while vvIBDV strains exhibited two out of the three typical residues found in Egyptian vvIBDV, namely Y220F and G254S, but not A321T. Meanwhile, all Egyptian variant strains exhibited typical residues found in nVarIBDV. However, all Egyptian variants showed a mutation at position 321 (321V), which represents the most exposed part of the capsid and is known to have a massive impact on IBDV antigenicity, except for one sample that had 318G instead. This report highlights the emergence of a new variant IBDV in Egypt, clustered with the Chinese new variants, spreading subclinically in broiler farms across a wide geographic area.RESEARCH HIGHLIGHTS New variant IBDV which emerged in Egypt clustered with Chinese nVarIBDV.nVarIBDV spread subclinically across a wide geographic area.Mutation at 321 represents capsid's most exposed part, a defining feature.Antigenically modified vvIBDV still circulating in Egypt with typical lesions.
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Infecções por Birnaviridae , Galinhas , Vírus da Doença Infecciosa da Bursa , Filogenia , Doenças das Aves Domésticas , Vírus da Doença Infecciosa da Bursa/genética , Vírus da Doença Infecciosa da Bursa/patogenicidade , Vírus da Doença Infecciosa da Bursa/isolamento & purificação , Animais , Egito/epidemiologia , Infecções por Birnaviridae/veterinária , Infecções por Birnaviridae/virologia , Infecções por Birnaviridae/epidemiologia , Doenças das Aves Domésticas/virologia , Doenças das Aves Domésticas/epidemiologia , Galinhas/virologia , Surtos de Doenças/veterinária , Sequência de Aminoácidos , Vacinas Virais/imunologia , Vacinação/veterinária , Proteínas Estruturais Virais/genética , Virulência , Variação GenéticaRESUMO
Wild migratory birds have the capability to spread avian influenza virus (AIV) over long distances as well as transmit the virus to domestic birds. In this study, swab and tissue samples were obtained from 190 migratory birds during close surveillance in Egypt in response to the recent outbreaks of the highly pathogenic avian influenza (HPAI) H5N1 virus. The collected samples were tested for a variety of AIV subtypes (H5N1, H9N2, H5N8, and H6N2) as well as other pathogens such as NDV, IBV, ILT, IBDV, and WNV. Among all of the tested samples, the HPAI H5N1 virus was found in six samples; the other samples were found to be negative for all of the tested pathogens. The Egyptian HPAI H5N1 strains shared genetic traits with the HPAI H5N1 strains that are currently being reported in Europe, North America, Asia, and Africa in 2021-2022. Whole genome sequencing revealed markers associated with mammalian adaption and virulence traits among different gene segments, similar to those found in HPAI H5N1 strains detected in Europe and Africa. The detection of the HPAI H5N1 strain of clade 2.3.4.4b in wild birds in Egypt underlines the risk of the introduction of this strain into the local poultry population. Hence, there is reason to be vigilant and continue epidemiological and molecular monitoring of the AIV in close proximity to the domestic-wild bird interface.
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Identification of avian infectious bronchitis virus (IBV) genotypes is essential for controlling infectious bronchitis (IB) disease, because vaccines that differ from the circulating strains might not provide efficient cross-protection. In Egypt, IBV strain typing is a difficult process, due to the widespread distribution of four genotype lineages (GI-13, GI-23, GI-1, and GI-16), which may contribute to IBV vaccination failure. In this study, we developed a multiplex real-time quantitative reverse transcription polymerase chain reaction (mRT-qPCR) assay that targets highly conserved areas of the S1 gene in order to detect classical (G1) and Egyptian variant II (G23) strains in allantoic fluids and clinical samples. The viral genotyping technique was assessed using commercially available vaccines as well as local strains, and 16 field isolates were tested to investigate its clinical applicability. The assay was found to be specific for the detection of classical and VAR II strains and did not detect the VAR I strain or other avian pathogens such as Newcastle disease virus, avian influenza virus (H9N2 and H5N8), or infectious bursal disease virus. The results also showed that 28 out of 41 samples tested positive for IBV utilizing rt-qRT-PCR targeting the N gene and that 26 out of the 28 positive samples were genotyped by mRT-qPCR targeting the S1 gene, whereas the remaining two samples that were not genotyped were VAR 1 (4/91) and VAR I (793/B). Interestingly, the testing could identify combined infections in one sample, indicating a mixed infection with both genotypes. The real-time RT-PCR assay could detect viral RNA at concentrations as low as 102 EID50 /ml for both classical and variant II. This assay is rapid, specific, and sensitive. It appears to be a valuable tool for regular disease monitoring that can be used to differentiate as well as identify viruses.
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Infecções por Coronavirus , Vírus da Bronquite Infecciosa , Vírus da Influenza A Subtipo H9N2 , Doenças das Aves Domésticas , Animais , Vírus da Bronquite Infecciosa/genética , Vírus da Influenza A Subtipo H9N2/genética , Transcrição Reversa , Doenças das Aves Domésticas/diagnóstico , Galinhas , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/veterinária , Reação em Cadeia da Polimerase em Tempo Real/métodosRESUMO
The highly pathogenic avian influenza (HPAI) H5N8 virus was first detected in Egypt in late 2016. Since then, the virus has spread rapidly among different poultry sectors, becoming the dominant HPAI H5 subtype reported in Egypt. Different genotypes of the HPAI H5N8 virus were reported in Egypt; however, the geographic patterns and molecular evolution of the Egyptian HPAI H5N8 viruses are still unclear. Here, extensive epidemiological surveillance was conducted, including more than half a million samples collected from different poultry sectors (farms/backyards/live bird markets) from all governorates in Egypt during 2019-2021. In addition, genetic characterization and evolutionary analyses were performed using 47 selected positive H5N8 isolates obtained during the same period. The result of the conducted surveillance showed that HPAI H5N8 viruses of clade 2.3.4.4b continue to circulate in different locations in Egypt, with an obvious seasonal pattern, and no further detection of the HPAI H5N1 virus of clade 2.2.1.2 was observed in the poultry population during 2019-2021. In addition, phylogenetic and Bayesian analyses revealed that two major genotypes (G5 and G6) of HPAI H5N8 viruses were continually expanding among the poultry sectors in Egypt. Notably, molecular dating analysis suggested that the Egyptian HPAI H5N8 virus is the potential ancestral viruses of the European H5N8 viruses of 2020-2021. In summary, the data of this study highlight the current epidemiology, diversity, and evolution of HPAI H5N8 viruses in Egypt and call for continuous monitoring of the genetic features of the avian influenza viruses in Egypt.
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Virus da Influenza A Subtipo H5N1 , Vírus da Influenza A , Influenza Aviária , Influenza Humana , Animais , Teorema de Bayes , Egito/epidemiologia , Humanos , Vírus da Influenza A/genética , Influenza Aviária/epidemiologia , Epidemiologia Molecular , Filogenia , Aves DomésticasRESUMO
Highly pathogenic avian influenza (HPAI) viruses continue to circulate worldwide, causing numerous outbreaks among bird species and severe public health concerns. H5N1 and H5N8 are the two most fundamental HPAI subtypes detected in birds in the last two decades. The two viruses may compete with each other while sharing the same host population and, thus, suppress the spread of one of the viruses. In this study, we performed a statistical analysis to investigate the temporal correlation of the HPAI H5N1 and HPAI H5N8 subtypes using globally reported data in 2015-2020. This was joined with an in-depth analysis using data generated via our national surveillance program in Egypt. A total of 6412 outbreaks were reported worldwide during this period, with 39% (2529) as H5N1 and 61% (3883) as H5N8. In Egypt, 65% of positive cases were found in backyards, while only 12% were found in farms and 23% in live bird markets. Overall, our findings depict a trade-off between the number of positive H5N1 and H5N8 samples around early 2017, which is suggestive of the potential replacement between the two subtypes. Further research is still required to elucidate the underpinning mechanisms of this competitive dynamic. This, in turn, will implicate the design of effective strategies for disease control.
Assuntos
Galinhas/virologia , Surtos de Doenças/veterinária , Monitoramento Epidemiológico/veterinária , Virus da Influenza A Subtipo H5N1/genética , Vírus da Influenza A Subtipo H5N8/genética , Influenza Aviária/epidemiologia , Influenza Aviária/virologia , Animais , Animais Selvagens/virologia , Surtos de Doenças/prevenção & controle , Egito/epidemiologia , Virus da Influenza A Subtipo H5N1/classificação , Virus da Influenza A Subtipo H5N1/patogenicidade , Vírus da Influenza A Subtipo H5N8/patogenicidade , Influenza Aviária/prevenção & controle , Filogenia , Doenças das Aves Domésticas/epidemiologia , Doenças das Aves Domésticas/virologiaRESUMO
Duck hepatitis virus (DHV) has always been considered one of the threats endangering duck farming in Egypt since the 1960s. In the current study, suspected DHV samples (n = 30) were obtained from commercial Pekin, Mulard (hybrid), and Muscovy duck farms and backyards in Beheira, Alexandria, Gharbia, Kafr El-Sheikh, and Giza provinces between 2012 and 2017. Diseased 3-21-day-old ducklings showed a clinical history of high mortality rates and nervous signs. Samples were screened by RT-PCR targeting the 5'UTR region and VP1 gene. The PCR-confirmed samples (n = 7) were isolated via allantoic route inoculation onto 9-day-old specific-pathogen-free embryonated chicken eggs. Embryos showed stunting, subcutaneous hemorrhages, and liver necrotic greenish-yellow foci. Duck hepatitis A virus-1 (DHAV-1) isolates were genetically analyzed in comparison to other field and vaccine strains. Phylogenetic analyses of the full-length VP1 gene sequences revealed that the obtained DHAV-1 field isolates clustered into genetic group 4 alongside other Egyptian strains isolated during the same period (95.9-99.72% similarity). Amino acid substitutions in the carboxyl-terminal of VP1 (I180T, G184E, D193N, and M213I) were identified in two strains. Also, deletion mutation at I189 was detected in three DHAV-1 strains. Additionally, the two amino acid residues E205 and N235 were common among the isolated strains and other virulent DHAV-1 strains. Two DHAV-1 isolates originated from Pekin source were selected for conducting the comparative pathogenicity testing based on detected point mutations at C-terminus of VP1. We evaluated the pathogenicity of these isolates by investigating clinical signs, mortality rates, and gross pathological and microscopic lesions. The study revealed that experimentally infected Pekin and Muscovy ducklings showed similar clinical signs including squatting down, lateral recumbency, and spasmodic kicking. Muscovy showed milder pathological changes in the liver compared to Pekin ducklings. Histopathological findings supported the gross pathological lesions detected in both breeds. In conclusion, these data provide updated information on the genetic diversity and pathotyping of Egyptian DHAV-1 strains. To the best of our knowledge, this is the first report of comparative pathogenicity of recent DHAV-1 strains in Pekin and Muscovy ducklings in Egypt and the Middle East region.
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This study was conducted to characterize the immunological parameters of chickens vaccinated with two formulated inactivated vaccines, water in oil (WO) and water in oil in water (WOW), prepared from velogenic Newcastle disease virus (vNDV) genotype VIIj isolated from outbreak among vaccinated chickens. Six groups (G1-G6) of commercial broiler chickens were established (n = 20). The G1-G3 were received homologous (WO and WOW) and heterologous (LaSota) inactivated vaccines, respectively. The G4 was vaccinated with live heterologous (LaSota) vaccine, while G5 and G6 were kept as control positive and control negative non-vaccinated groups. The antibody titers were measured against vNDV and LaSota antigens using hemagglutination inhibition (HI) test, the cytokine gene expressions of IFNγ, IL1ß, IL4, IL6, IL8, and IL18 were quantified using real-time RT-PCR, and the virus shedding was titrated on chicken embryo fibroblast cells after challenging by vNDV. The classical clinical signs and 100% mortality were observed only in G5 after vNDV challenging. The highest HI titers were detected in G1, G2, and G3 using NDV/168 antigen with no significant differences among them. These groups showed higher HI titer than G4 (2-4log2). Cytokine gene expression of IFNγ, IL1, IL6, IL8, and IL18 were significantly downregulated in vaccinated chickens with upregulation of IL4 than non-vaccinated challenge group. Viral shedding titers were significantly (0.0001, p ≤ 0.001) reduced in all samples form vaccinated chickens. In conclusion, the prepared vaccines produced highly efficient immunological responses and could be used for controlling the NDV infection.
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Emulsões/administração & dosagem , Doença de Newcastle/prevenção & controle , Vírus da Doença de Newcastle/imunologia , Doenças das Aves Domésticas/prevenção & controle , Vacinas Virais/imunologia , Animais , Anticorpos Antivirais/sangue , Galinhas/imunologia , Galinhas/virologia , Citocinas/imunologia , Composição de Medicamentos , Genótipo , Doença de Newcastle/imunologia , Vírus da Doença de Newcastle/genética , Óleos , Doenças das Aves Domésticas/virologia , Vacinação , Vacinas de Produtos Inativados/administração & dosagem , Vacinas de Produtos Inativados/imunologia , Vacinas Virais/administração & dosagem , Eliminação de Partículas Virais , ÁguaRESUMO
Infectious bursal disease virus (IBDV), the agent of an immunosuppressive and sometimes lethal disease in chickens, is causing recurrent outbreaks in broiler chickens in Egypt. In particular, an antigenically modified isolate of very virulent IBDV (vvIBDV) called 99323 was detected in Egypt nearly twenty years ago; this isolate was shown to be experimentally controlled by an antigenically classical live vaccine. However, acute IBD is still reported, even in vaccinated flocks, and little is known about the genetic and antigenic properties of viruses currently circulating in Egypt. In the present study, ten samples collected in Egyptian broiler farms in 2015 as well as five samples collected in 2001 were analyzed. Genetic analyses of partial VP2 sequences revealed that 8 isolates clustered with vvIBDV strains, and 5 with tissue culture adapted and vaccine strains. Similar results were observed for partial VP1 sequences with the exception of isolate 160019, for which VP2 clustered with the vaccine strain Bursine while VP1 clustered with vvIBDV, suggesting reassortment. For isolates genetically related to vvIBDV, antigenic profiling revealed two patterns: while some isolates exhibited typical European vvIBDV reactivity with lack of binding of mAbs 5, other revealed extensive antigenic modifications, with lack of binding of mAbs 3, 5, 6, 8 and 9, similar to isolate 99323. These different patterns were associated with a single amino acid mutation at position 321 of VP2 that is located within peak PHI. Full genome sequencing was performed for three isolates, among which two were representative of the two antigenic patterns observed for vvIBDV as well as the reassortant isolate 160019. This study highlights the co-circulation of both antigenically typical and modified vvIBDV during the last fifteen years in Egypt.
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Infecções por Birnaviridae/veterinária , Galinhas/virologia , Vírus da Doença Infecciosa da Bursa/patogenicidade , Doenças das Aves Domésticas/virologia , Proteínas Estruturais Virais/genética , Substituição de Aminoácidos , Animais , Infecções por Birnaviridae/virologia , Surtos de Doenças/veterinária , Egito/epidemiologia , Vírus da Doença Infecciosa da Bursa/classificação , Vírus da Doença Infecciosa da Bursa/genética , Vírus da Doença Infecciosa da Bursa/imunologia , Filogenia , Análise de Sequência de RNA , Proteínas Estruturais Virais/imunologia , Virulência , Sequenciamento Completo do GenomaRESUMO
Highly pathogenic avian influenza (HPAI) H5N1 and H5N8 have become endemic among domestic poultry in Egypt since 2006 and 2016, respectively. In parallel, the low pathogenic avian influenza H9N2 virus has been endemic since 2010. Despite the continuous circulation of these subtypes for several years, no natural reassortant has been detected so far among the domestic poultry population in Egypt. In this study, the HPAI (H5N2) virus was isolated from a commercial duck farm, giving evidence of the emergence of the first natural reassortment event in domestic poultry in Egypt. The virus was derived as a result of genetic reassortment between avian influenza viruses of H5N8 and H9N2 subtypes circulating in Egypt. The exchange of the neuraminidase segment and high number of acquired mutations might be associated with an alteration in the biological propensities of this virus.
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Patos/virologia , Vírus da Influenza A Subtipo H5N2/isolamento & purificação , Influenza Aviária/virologia , Vírus Reordenados/isolamento & purificação , Animais , Egito , Vírus da Influenza A Subtipo H5N2/classificação , Vírus da Influenza A Subtipo H5N2/genética , Vírus Reordenados/classificação , Vírus Reordenados/genéticaRESUMO
Immunosuppressive viral diseases have a great economic importance in the poultry industry due to the increased susceptibility to secondary infections. Chicken anaemia virus (CAV) is one of the major immunosuppressive diseases in chickens. In addition, low pathogenic avian influenza (LPAI) of subtype H9N2 and infectious bronchitis (IB) viruses are among the most frequently reported respiratory viral diseases in poultry worldwide. In the present study, specific pathogen free chickens were used to understand the impact of CAV on secondary infection with LPAI-H9N2 or IB viruses. Clinical outcomes, viral shedding dynamics, and cytokine levels wereassessed. The results exhibit that chickens previously infected with CAV produceconsiderablyhigher titresof LPAI-H9N2 or IB viruses in the oropharyngeal swabs (P < 0.05), tracheas and kidneys. In addition, the immunologic effect of CAV provokedthe development of clinical signs of LPAI-H9N2 and IB virus infections. Moreover, results suggested that pre-infection with CAV directly correlated with elevated levels of IL-6 and IFNγ. These findings underline the importance of CAV pre-infection on LPAI-H9N2 or IB infection in chickens, and indicate that co-circulation of CAV can contribute to the spread and evolution of LPAI H9N2 and IB viruses.
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Infecções por Circoviridae/veterinária , Coinfecção/veterinária , Infecções por Coronavirus/veterinária , Influenza Aviária/imunologia , Doenças das Aves Domésticas/virologia , Animais , Vírus da Anemia da Galinha/imunologia , Galinhas/virologia , Infecções por Circoviridae/imunologia , Coinfecção/imunologia , Coinfecção/virologia , Infecções por Coronavirus/imunologia , Citocinas/sangue , Vírus da Bronquite Infecciosa/imunologia , Vírus da Influenza A Subtipo H9N2 , Doenças das Aves Domésticas/imunologia , Organismos Livres de Patógenos Específicos , Eliminação de Partículas ViraisRESUMO
AIM: The aim of this work was to study the full F gene sequence of Newcastle disease virus (NDV) in regard to pathotyping and genotyping and to study the evolution of this NDV in Egypt. MATERIALS AND METHODS: The present study was conducted using samples from seven suspected NDV flocks of vaccinated chickens during 2012-2016 from six governorates in Egypt. The NDV was successfully isolated from pathological specimens through inoculation in specific pathogen-free embryonated chicken eggs. RESULTS: Pathogenicity of the NDV isolates has been estimated through intracerebral pathogenicity index and ranged from 1.66 to 1.73 which indicates the velogenic type of NDV isolates. Pathotyping and genotyping of these isolates were done through sequencing of full-length F gene. Results indicated that the seven NDV isolates showed characteristic cleavage site motif (112RRQKRF117) for the velogenic strains of NDV. Phylogenetic analysis of the F gene clustered these isolates within Group I of genotype VIId within Israeli strains NDV/IS/2015, NDV-Ch/SD883, and most of the Middle East strains. Six of seven sequenced isolates have six potential N-linked glycosylation sites. The neutralization epitope on the five antigenic sites of fusion is conserved in all Egyptian strains of this study except NDV-KFR-B7-2012 which has a substitution at D 170 N in epitope A4. In all our strains, 10 cysteine residues are recorded, except one loss of cysteine at residue 370 in both NDV-EG-35-2014 and NDV-GHB-328F-2016. CONCLUSION: All viruses in this study have 52 amino acid substitutions within fusion gene in compared with Lasota strain that reveals importance for its antigenic and structural function. The present work highlights the important need to sequence F gene of NDV genotype VIId to investigate the evolution of this NDV in Egypt.
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Chicken anemia virus (CAV) is one of the commercially important diseases of poultry worldwide. In Egypt, CAV has been reported to be a potential threat to the commercial poultry sectors. Hence, this study was aimed at isolation and full genomic analysis of CAVs circulating in chicken populations in different geographical location in Egypt. A total of 42 samples were collected from broiler chicken flocks in 9 governorates in Egypt from 12 to 42 days of age. The mortality rate observed among chickens was ranging from 3% to 22%. Nineteen out of 42 farms were found positive for the CAV genome by polymerase chain reaction (PCR). Full genome sequencing was conducted for 18 positive samples. Genetic analysis revealed a high similarity of >99% in 11 viruses with the vaccine strain Del-Ros; while the other seven samples shared close similarity to CAV field strains isolated from China, Taiwan, and Brazil. The data also indicated Q139 and Q144 amino acids substitutions among the VP1 of Egyptian field strains, which are known to be important in virus replication and spread. Phylogenetic analysis of the sequenced viruses (nâ¯=â¯18) based on either the full gene nucleotide sequence or VP1 coding sequence, suggested the circulation of four distinct genotypes in Egypt designated as group A, B, C and D. Moreover, evidence of recombination was detected among four Egyptian CAVs located within group A. The findings of this study succeeded to elucidate the epidemiological and genetic features of CAVs circulating in Egypt, and underscores the important of CAVs surveillance.
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Vírus da Anemia da Galinha/classificação , Vírus da Anemia da Galinha/isolamento & purificação , Infecções por Circoviridae/veterinária , Variação Genética , Doenças das Aves Domésticas/virologia , Recombinação Genética , Substituição de Aminoácidos , Animais , Vírus da Anemia da Galinha/genética , Galinhas , Infecções por Circoviridae/epidemiologia , Infecções por Circoviridae/mortalidade , Infecções por Circoviridae/virologia , Egito/epidemiologia , Evolução Molecular , Genoma Viral , Genótipo , Filogeografia , Doenças das Aves Domésticas/epidemiologia , Doenças das Aves Domésticas/mortalidade , Análise de Sequência de DNARESUMO
We isolated highly pathogenic avian influenza virus (H5N8) of clade 2.3.4.4 from the common coot (Fulica atra) in Egypt, documenting its introduction into Africa through migratory birds. This virus has a close genetic relationship with subtype H5N8 viruses circulating in Europe. Enhanced surveillance to detect newly emerging viruses is warranted.
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Migração Animal/fisiologia , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A Subtipo H5N8/genética , Influenza Aviária/epidemiologia , Vírus Reordenados/genética , Sequência de Aminoácidos , Animais , Animais Selvagens , Aves , Egito/epidemiologia , Expressão Gênica , Glicoproteínas de Hemaglutininação de Vírus da Influenza/química , Glicoproteínas de Hemaglutininação de Vírus da Influenza/metabolismo , Vírus da Influenza A Subtipo H5N8/classificação , Vírus da Influenza A Subtipo H5N8/isolamento & purificação , Vírus da Influenza A Subtipo H5N8/patogenicidade , Influenza Aviária/transmissão , Influenza Aviária/virologia , Modelos Moleculares , Mutação , Filogenia , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Vírus Reordenados/classificação , Vírus Reordenados/isolamento & purificação , Vírus Reordenados/patogenicidadeRESUMO
In Egypt, since 2006, descendants of the highly pathogenic avian influenza virus (HP AIV) H5N1 of clade 2.2 continue to cause sharp losses in poultry production and seriously threaten public health. Potentially zoonotic H9N2 viruses established an endemic status in poultry in Egypt as well and co-circulate with HP AIV H5N1 rising concerns of reassortments between H9N2 and H5N1 viruses along with an increase of mixed infections of poultry. Nucleotide sequences of whole genomes of 15 different isolates (H5N1: 7; H9N2: 8), and of the hemagglutinin (HA) and neuraminidase (NA) encoding segments of nine further clinical samples (H5N1: 2; H9N2: 7) from 2013 and 2014 were generated and analysed. The HA of H5N1 viruses clustered with clade 2.2.1 while the H9 HA formed three distinguishable subgroups within cluster B viruses. BEAST analysis revealed that H9N2 viruses are likely present in Egypt since 2009. Several previously undescribed substituting mutations putatively associated with host tropism and virulence modulation were detected in different proteins of the analysed H9N2 and H5N1 viruses. Reassortment between HP AIV H5N1 and H9N2 is anticipated in Egypt, and timely detection of such events is of public health concern. As a rapid tool for detection of such reassortants discriminative SYBR-Green reverse transcription real-time PCR assays (SG-RT-qPCR), targeting the internal genes of the Egyptian H5N1 and H9N2 viruses were developed for the rapid screening of viral RNAs from both virus isolates and clinical samples. However, in accordance to Sanger sequencing, no reassortants were found by SG-RT-qPCR. Nevertheless, the complex epidemiology of avian influenza in poultry in Egypt will require sustained close observation. Further development and continuing adaptation of rapid and cost-effective screening assays such as the SG-RT-qPCR protocol developed here are at the basis of efforts for improvement the currently critical situation.
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Evolução Molecular , Virus da Influenza A Subtipo H5N1/genética , Vírus da Influenza A Subtipo H9N2/genética , Influenza Aviária/virologia , Doenças das Aves Domésticas/virologia , Animais , Embrião de Galinha , Cães , Egito , Genes Virais , Virus da Influenza A Subtipo H5N1/patogenicidade , Vírus da Influenza A Subtipo H9N2/patogenicidade , Células Madin Darby de Rim Canino , Filogenia , Aves Domésticas/virologia , Seleção Genética , Proteínas Virais/genética , VirulênciaRESUMO
Vaccination is the main tool implemented in Egypt since 2007 to control H5N1 avian influenza. The present study aimed at comparing the effectiveness of three avian influenza vaccination regimes in commercial broiler chickens carrying high levels of maternally derived antibodies (MDAs). Day-old chicks were divided into four experimental groups. Group I received only the rHVT-H5 vaccine (recombinant turkey herpesvirus (HVT) which carries a H5 clade 2.2 insert) administered at D1. Group II received only the KV-H5 (an oil emulsion killed vaccine prepared from reassortant HPAI virus (A/duck/Anhui/1/06)) vaccine (inactivated reverse genetic H5N1 clade 2.3.4 virus) administered at D8. Group III received rHVT-H5 and KV-H5 as prime/boost. Group IV served as unvaccinated control. Weekly serological monitoring was conducted using the haemagglutination inhibition test. Two challenge experiments were conducted at D28 and D35 using HPAI H5N1 clade 2.2.1 virus. Birds were monitored daily 14 days post-challenge for morbidity and mortality, and oropharyngeal swabs were collected for virological monitoring. Initially, day-old chicks had high mean MDA titres (9 + 0.9 log2). The MDA half-life was >7 and <7 days, respectively, for unvaccinated and vaccinated birds. Group III showed the highest post-vaccination humoral immune response and seroconversion rate. The highest protection rate against morbidity (80-90%) and mortality (90-90%) was obtained in Group III after challenge at D28 and D35, respectively, as compared to Group I (70-70%) and (80-90%) and Group II (0-0%) and (30-30%). Groups I and III had lower number of shedder birds. The vaccination regime with prime/boost conferred the highest and earliest protection, and can hence be recommended for the broiler production sector in endemic and high HPAI H5N1 challenge areas.
Assuntos
Virus da Influenza A Subtipo H5N1/imunologia , Vacinas contra Influenza/administração & dosagem , Influenza Aviária/prevenção & controle , Doenças das Aves Domésticas/prevenção & controle , Vacinação , Animais , Galinhas , Egito , Testes de Inibição da Hemaglutinação , Vacinas contra Influenza/imunologia , Influenza Aviária/epidemiologia , Influenza Aviária/imunologia , Doenças das Aves Domésticas/epidemiologia , Doenças das Aves Domésticas/imunologia , Vacinas de Produtos Inativados/administração & dosagem , Vacinas de Produtos Inativados/imunologia , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/imunologiaRESUMO
Duck hepatitis virus (DHV) is an acute highly contagious disease of ducklings caused by three distinct serotypes of duck hepatitis A virus (DHAV), a member of the RNA family Picornaviridae, where serotype 1 is the most widespread serotype worldwide. To date, little if any is known about the prevalence and genetic characterisation of DHAV outside Asia. The current study describes surveillance on DHV in 46 commercial duck farms in Egypt with a history of high mortality in young ducklings from 3 to 15 day-old from 2012 to 2014. Clinical samples were examined by generic RT-PCR assays followed by partial sequence analysis of the 5'UTR, VP1 and 3D genes of the vaccine strain and 15 field viruses. The overall positive rate was 37% (n=17/46). All duck breeds (Pekin, Muscovy, Mallard and Green Winged) were susceptible to the disease with mortality ranged from 15% to 96.7%. Sequence and phylogenetic analyses indicated that the Egyptian strains cluster in the DHAV serotype 1 with Asian viruses and distinguishable from the vaccine strains. So far, this is the first report on the genetic characterisation of DHAV in Egypt. This study may be useful to better understand the epidemiology and evolution of DHAV.
Assuntos
Patos , Vírus da Hepatite do Pato/genética , Infecções por Picornaviridae/veterinária , Doenças das Aves Domésticas/virologia , Regiões 5' não Traduzidas/genética , Animais , Cruzamento , Proteínas do Capsídeo/química , Proteínas do Capsídeo/genética , Patos/classificação , Patos/virologia , Egito/epidemiologia , Vírus da Hepatite do Pato/classificação , Vírus da Hepatite do Pato/isolamento & purificação , Rim/patologia , Rim/virologia , Fígado/patologia , Fígado/virologia , Dados de Sequência Molecular , Filogenia , Infecções por Picornaviridae/epidemiologia , Infecções por Picornaviridae/virologia , Reação em Cadeia da Polimerase/veterinária , Doenças das Aves Domésticas/epidemiologia , Doenças das Aves Domésticas/mortalidade , Prevalência , Análise de Sequência/veterinária , Baço/patologia , Baço/virologiaRESUMO
The effectiveness of recombinant turkey herpesvirus avian influenza (A/swan/Hungary/4999/2006(H5N1)) clade 2.2 virus (rHVT-H5) vaccine was evaluated in two layer chicken breeds (White Bovans [WB] and Brown Shaver [BS]). One dose of rHVT-H5 vaccine was administered at day 1 and birds were monitored serologically (haemagglutination inhibition test) and virologically for 19 weeks. Maternally-derived antibody and post-vaccination H5 antibody titres were measured using the Chinese (A/Goose/Guangdong/1/96(H5N1)) HA and the Egyptian (A/chicken/Egypt/128s/2012(H5N1)) HA as antigens. The challenge was conducted at 19 weeks of age and on six experimental groups: Groups I (WB) and II (BS), both vaccinated and challenged; Groups III (WB) and IV (BS), both vaccinated but not challenged; Groups V and VI, unvaccinated specific pathogen free chickens, serving respectively as positive and negative controls. The challenge virus was the clade 2.2.1 highly pathogenic avian influenza H5N1 A/chicken/Egypt/128s/2012 at a dose of 10(6) median embryo infective dose. For both breeds, complete maternally-derived antibody waning occurred at the age of 4 weeks. The immune response to rHVT-H5 vaccination was detected from the sixth week. The seroconversion rates for both breeds reached 85.7 to 100% in the eighth week of age. Protection levels of 73.3%, 60% and 0% were respectively recorded in Groups I, II and V. No mortalities occurred in the unchallenged groups. Group I showed superior results for all measured post-challenge parameters. In conclusion, a single rHVT-H5 hatchery vaccination conferred a high level of protection for a relatively extended period. This vaccine could be an important tool for future A/H5N1 prevention/control in endemic countries. Further studies on persistence of immunity beyond 19 weeks, need for booster with inactivated vaccines, breed susceptibility and vaccinal response, and transmissibility are recommended.
Assuntos
Galinhas/imunologia , Herpesvirus Meleagrídeo 1/imunologia , Virus da Influenza A Subtipo H5N1/imunologia , Influenza Aviária/prevenção & controle , Doenças das Aves Domésticas/prevenção & controle , Vacinação/veterinária , Animais , Egito , Feminino , Vacinas contra Influenza/imunologia , Influenza Aviária/imunologia , Influenza Aviária/virologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/virologia , Organismos Livres de Patógenos Específicos , Vacinas de Produtos Inativados/imunologia , Vacinas SintéticasRESUMO
BACKGROUND: The endemic H5N1 high pathogenicity avian influenza virus (A/H5N1) in poultry in Egypt continues to cause heavy losses in poultry and poses a significant threat to human health. METHODS: Here we describe results of A/H5N1 surveillance in domestic poultry in 2009 and wild birds in 2009-2010. Tracheal and cloacal swabs were collected from domestic poultry from 22024 commercial farms, 1435 backyards and 944 live bird markets (LBMs) as well as from 1297 wild birds representing 28 different types of migratory birds. Viral RNA was extracted from a mix of tracheal and cloacal swabs media. Matrix gene of avian influenza type A virus was detected using specific real-time reverse-transcription polymerase chain reaction (RT-qPCR) and positive samples were tested by RT-qPCR for simultaneous detection of the H5 and N1 genes. RESULTS: In this surveillance, A/H5N1 was detected from 0.1% (n = 23/) of examined commercial poultry farms, 10.5% (n = 151) of backyard birds and 11.4% (n = 108) of LBMs but no wild bird tested positive for A/H5N1. The virus was detected from domestic poultry year-round with higher incidence in the warmer months of summer and spring particularly in backyard birds. Outbreaks were recorded mostly in Lower Egypt where 95.7% (n = 22), 68.9% (n = 104) and 52.8% (n = 57) of positive commercial farms, backyards and LBMs were detected, respectively. Higher prevalence (56%, n = 85) was reported in backyards that had mixed chickens and waterfowl together in the same vicinity and LBMs that had waterfowl (76%, n = 82). CONCLUSION: Our findings indicated broad circulation of the endemic A/H5N1 among poultry in 2009 in Egypt. In addition, the epidemiology of A/H5N1 has changed over time with outbreaks occurring in the warmer months of the year. Backyard waterfowl may play a role as a reservoir and/or source of A/H5N1 particularly in LBMs. The virus has been established in poultry in the Nile Delta where major metropolitan areas, dense human population and poultry stocks are concentrated. Continuous surveillance, tracing the source of live birds in the markets and integration of multifaceted strategies and global collaboration are needed to control the spread of the virus in Egypt.
Assuntos
Virus da Influenza A Subtipo H5N1/isolamento & purificação , Influenza Aviária/epidemiologia , Influenza Aviária/virologia , Animais , Aves , Cloaca/virologia , Surtos de Doenças , Egito/epidemiologia , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Virus da Influenza A Subtipo H5N1/genética , Neuraminidase/genética , Aves Domésticas , Prevalência , Reação em Cadeia da Polimerase em Tempo Real , Estações do Ano , Traqueia/virologia , Proteínas da Matriz Viral/genética , Proteínas Virais/genéticaRESUMO
This study describes the first isolation of H9N2 avian influenza virus (AIV) from commercial bobwhite quail (Colinus virginianus) in Egypt. Infected birds showed neither clinical signs nor mortality. Virus isolation and real-time reverse transcription polymerase chain reaction confirmed the presence of the H9N2 virus in cloacal swab samples collected at 35 days of age and the absence of other AIV subtypes, including H5 and H7. The hemagglutinin and neuraminidase genes of the isolated virus showed 99.1% and 98.2% nucleotide identity and 97.3% and 100% amino acid identity, respectively, to those of H9N2 viruses currently circulating in poultry in the Middle East. Phylogenetically, the Egyptian H9N2 virus was closely related to viruses of the G1-like lineage isolated from neighbouring countries, indicating possible epidemiological links.