The modulatory effect of carvacrol on viral shedding titer and acute phase response in broiler chickens experimentally infected with infectious bronchitis virus.

Infectious bronchitis virus (IBV) is one of the major respiratory diseases of broiler causing huge economic losses. The inability to control IBV using different vaccination programs owing to the high mutation rate and recombination ability of the RNA genome generates IBV variants. This study was designed to give a specific perspective of carvacrol effect on early immune response, viral shedding titer, oxidative stress, serum biochemical parameters and clinical consequences in broilers experimentally infected by IBV. One hundred and twenty-one-day old commercial broiler chicks were equally divided into 4 groups. First group was considered as control. Second group was given carvacrol, third group was infected with IBV and fourth group was given carvacrol and infected with IBV. Infection with variant IBV induced significant upregulation of chicken interferon-inducible transmembrane protein 3 (chIFITM3) gene in trachea, elevations in serum levels of Alpha-1 acid glycoprotein (α1-AGP) and Interleukin 6 (IL-6), total leucocytic count (TLC), heterophil/lymphocyte (H/L) ratio and oxidative stress in lung and kidney tissues. Beside, histopathological changes in trachea, lung and kidney induced by IBV, elevation of kidney function tests was detected. The pretreatment with carvacrol significantly reduced viral shedding titer, chIFITM3 gene expression, IL-6 and α1-AGP levels, leucocytic response and H/L ratio with minimization of clinical signs intensity. Also, carvacrol relieved oxidative stress, ameliorated the increased uric acid level and histopathological alterations in kidney and lung caused by viral infection.

Correction to: Oral inoculation of ultraviolet-irradiated Eimeria species oocysts protects chickens against coccidiosis.

The author name Salama Abohamra in the original published version of this article should have been Salama Abohamra Sayed Shany.

Molecular diagnosis of Eimeria stiedae in hepatic tissue of experimentally infected rabbits.

The early detection of Eimeria stiedae in the hepatic tissue of experimentally infected rabbits was investigated using molecular assay. Forty 6-week-old male New Zealand rabbits were divided into two groups. Group A (30 animals) was infected with 2.5 × 10(4) sporulated oocysts of E. stiedae per animal on Day 0 and Group B (10 animals) was used as the uninfected controls. Three animals from Group A and one from Group B were sacrificed at 0, 3, 6, 9, 12, 15, 18, 21, 24 and 27 days post infection (PI). Gross and microscopic post-mortem findings were recorded. Polymerase chain reaction (PCR) of the E. stiedae internal transcribed spacer 1 genomic region was conducted on blood, liver tissue, and feces from the Group A experimentally infected animals. Macroscopically, the liver showed irregular yellowish white nodules pathognomonic to E. stiedae infection beginning on Day 15 PI. Hepatomegaly and ascites were obvious from Day 21-24 PI. The presence of different E. stiedae schizonts and gametocytes in the histopathological sections of the biliary epithelium were evident on Day 15 PI. The E. stiedae PCR was first positive in liver tissues on Day 12 and in fecal samples on Day 18 PI, but the blood samples were negative. In conclusion, the PCR can be used for early diagnosis and control of E. stiedae schizonts before shedding of the oocysts in feces.

Efficacy of Two Vaccination Strategies against Infectious Bronchitis in Laying Hens.

Vaccination remains the leading control method against infectious bronchitis (IB) in poultry despite the frequently observed IB outbreaks in vaccinated flocks. Here, two vaccination regimes were evaluated against challenge with the Massachusetts (Mass) infectious bronchitis virus (IBV) strain that was linked to egg production defects in Western Canada. One vaccination strategy included live attenuated IB vaccines only, and the other used both inactivated and live attenuated IB vaccines. The two immunization programs involved priming with a monovalent live attenuated IB vaccine (Mass serotype) at day-old, followed by intervals of bivalent live attenuated IB vaccines containing the Mass and Connecticut (Conn) serotypes given to the pullets at 2-, 5-, 9-, and 14-week-old. Inactivated IB vaccine (Mass serotype) was administrated to only one group of the vaccinated birds at 14-week-old. At the peak of lay, the hens were challenged with the Mass IBV isolate (15AB-01) via the oculo-nasal route. The efficacy of the vaccines was assessed following the challenge by observing clinical signs, egg production, egg quality parameters, seroconversion, and systemic T-cell subsets (CD4+ and CD8+ cells). Moreover, the viral genome loads in the oropharyngeal (OP) and cloacal (CL) swabs were quantified at predetermined time points. At 14 days post-infection (dpi), all the hens were euthanized, and different tissues were collected for genome load quantification and histopathological examination. Post-challenge, both vaccination regimes showed protection against clinical signs and exhibited significantly higher albumen parameters, higher anti-IBV serum antibodies, and significantly lower levels of IBV genome loads in OP swabs (at 3 and 7 dpi) and trachea and cecal tonsils compared to the mock-vaccinated challenged group. However, only the birds that received live attenuated plus inactivated IB vaccines had significantly lower IBV genome loads in CL swabs at 7 dpi, as well as decreased histopathological lesion scores and IBV genome loads in magnum compared to the mock-vaccinated challenged group, suggesting a slightly better performance for using live attenuated and inactivated IB vaccines in combination. Overall, the present findings show no significant difference in protection between the two vaccination regimes against the Mass IBV challenge in laying hens.

Effect of aromatic oils on the expression of some virulence-associated and antimicrobial resistance genes of Escherichia coli isolated from broilers.

Objectives: This study aimed to prove the effects of Escherichia coli isolates isolated from diseased broilers to form biofilms, describe their antimicrobial sensetivity, and determine the effect of allicin and cinnamon essential oils on the expression of some genes (fimH, int1, and luxS) through quantitative polymerase chain reaction (q-PCR). Materials and Methods: 140 samples were obtained from diseased broilers in Beni-Suef Governorate, Egypt. These samples were examined by conventional bacteriology methods to detect the causative agent. The antimicrobial susceptibility of the isolated bacteria was assessed using the disc diffusion method, The ability of yeast extract-casamino acids Congo Red Agar to generate phenotypic biofilms was next tested. The presence of resistance and virulence genes in some multidrug resistant isolates was genotypically investigated. The antibacterial effects of allicin and cinnamon oil were evaluated against the growth of multidrug-resistant E. coli. Finally, q-PCR was utilized to assess changes in some genes' expression. Results: Escherichia coli was isolated from 61 samples (43.6%). An antimicrobial susceptibility test revealed that multidrug-resistance (MDR) (could resist more than three antimicrobial classes) E. coli prevalence was 100%. 40.8% of isolates phenotypically produce biofilms. The detection of resistance and virulence genes by PCR showed that all tested isolates carry aadB, fimH, int1, qnrS, and luxS genes, while only 40% harbor iss genes. q-PCR showed that after treatment with allicin and cinnamon oils, gene expression went down. Conclusion: This investigation highlights that E. coli showed resistance against most of the tested antimicrobials; all isolates were MDR. The study showed wide dissemination of virulence and resistance genes among E. coli. Allicin and cinnamon oils have antimicrobial activities and could be used as alternatives to synthetic antimicrobial agents.

Genotyping and In Silico Analysis of Delmarva (DMV/1639) Infectious Bronchitis Virus (IBV) Spike 1 (S1) Glycoprotein.

Genetic diversity and evolution of infectious bronchitis virus (IBV) are mainly impacted by mutations in the spike 1 (S1) gene. This study focused on whole genome sequencing of an IBV isolate (IBV/Ck/Can/2558004), which represents strains highly prevalent in Canadian commercial poultry, especially concerning features related to its S1 gene and protein sequences. Based on the phylogeny of the S1 gene, IBV/Ck/Can/2558004 belongs to the GI-17 lineage. According to S1 gene and protein pairwise alignment, IBV/Ck/Can/2558004 had 99.44-99.63% and 98.88-99.25% nucleotide (nt) and deduced amino acid (aa) identities, respectively, with five Canadian Delmarva (DMV/1639) IBVs isolated in 2019, and it also shared 96.63-97.69% and 94.78-97.20% nt and aa similarities with US DMV/1639 IBVs isolated in 2011 and 2019, respectively. Further homology analysis of aa sequences showed the existence of some aa substitutions in the hypervariable regions (HVRs) of the S1 protein of IBV/Ck/Can/2558004 compared to US DMV/1639 isolates; most of these variant aa residues have been subjected to positive selection pressure. Predictive analysis of potential N-glycosylation and phosphorylation motifs showed either loss or acquisition in the S1 glycoprotein of IBV/Ck/Can/2558004 compared to S1 of US DMV/1639 IBV. Furthermore, bioinformatic analysis showed some of the aa changes within the S1 protein of IBV/Ck/Can/2558004 have been predicted to impact the function and structure of the S1 protein, potentially leading to a lower binding affinity of the S1 protein to its relevant ligand (sialic acid). In conclusion, these findings revealed that the DMV/1639 IBV isolates are under continuous evolution among Canadian poultry.

Genotype VII.1.1-Based Newcastle Disease Virus Vaccines Afford Better Protection against Field Isolates in Commercial Broiler Chickens.

This study evaluated the efficacy of live and inactivated conventional GII LaSota and recombinant GVII Newcastle disease vaccines in commercial broilers. The experimental groups (G2-G7) were vaccinated on day 7 and day 21 of age with live vaccines from the same vaccine type "GII LaSota, GVII vaccine (A), GVII vaccine (B)" via eye drop; however, G3, G5, and G7 received a single dose from inactivated counterpart vaccines subcutaneously on day 7 of age. Vaccine efficacy was evaluated based on elicited humoral immunity, clinical protection, and reduction in virus shedding after challenge with virulent GVII 1.1. strain. Results demonstrated that live and inactivated recombinant GVII vaccine based on VG/GA strain backbone elicited superior protection parameters (100% protection). Although the conventional GII LaSota live and inactivated vaccination regime protected 93.3% of vaccinated birds, the virus shedding continued until 10 DPC. The post-vaccination serological monitoring was consistent with protection results. The study concludes that conventional GII ND vaccines alone are probably insufficient due to the current epidemiology of the GVII 1.1 NDV strains. Our findings further support that protection induced by recombinant GVII 1.1. ND vaccines are superior. Interestingly, the efficacy of recombinant ND vaccines seemed to be influenced by the backbone virus since the VG/GA backbone-based vaccine provided better protection and reduced virus shedding.

Respiratory disease due to mixed viral infections in poultry flocks in Egypt between 2017 and 2018: Upsurge of highly pathogenic avian influenza virus subtype H5N8 since 2018.

For several years, poultry production in Egypt has been suffering from co-circulation of multiple respiratory viruses including highly pathogenic avian influenza virus (HPAIV) H5N1 (clade 2.2.1.2) and low pathogenic H9N2 (clade G1-B). Incursion of HPAIV H5N8 (clade 2.3.4.4b) to Egypt in November 2016 via wild birds followed by spread into commercial poultry flocks further complicated the situation. Current analyses focussed on 39 poultry farms suffering from respiratory manifestation and high mortality in six Egyptian governorates during 2017-2018. Real-time RT-PCR (RT-qPCR) substantiated the co-presence of at least two respiratory virus species in more than 80% of the investigated flocks. The percentage of HPAIV H5N1-positive holdings was fairly stable in 2017 (12.8%) and 2018 (10.2%), while the percentage of HPAIV H5N8-positive holdings increased from 23% in 2017 to 66.6% during 2018. The proportion of H9N2-positive samples was constantly high (2017:100% and 2018:63%), and H9N2 co-circulated with HPAIV H5N8 in 22 out of 39 (56.8%) flocks. Analyses of 26 H5, 18 H9 and 4 N2 new sequences confirmed continuous genetic diversification. In silico analysis revealed numerous amino acid substitutions in the HA and NA proteins suggestive of increased adaptation to mammalian hosts and putative antigenic variation. For sensitive detection of H9N2 viruses by RT-qPCR, an update of primers and probe sequences was crucial. Reasons for the relative increase of HPAIV H5N8 infections versus H5N1 remained unclear, but lack of suitable vaccines against clade 2.3.4.4b cannot be excluded. A reconsideration of surveillance and control measures should include updating of diagnostic tools and vaccination strategies.

Isolation and characterization of highly pathogenic avian influenza virus subtype H5N1 from donkeys.

BACKGROUND: The highly pathogenic H5N1 is a major avian pathogen that crosses species barriers and seriously affects humans as well as some mammals. It mutates in an intensified manner and is considered a potential candidate for the possible next pandemic with all the catastrophic consequences. METHODS: Nasal swabs were collected from donkeys suffered from respiratory distress. The virus was isolated from the pooled nasal swabs in specific pathogen free embryonated chicken eggs (SPF-ECE). Reverse transcriptase polymerase chain reaction (RT-PCR) and sequencing of both haemagglutingin and neuraminidase were performed. H5 seroconversion was screened using haemagglutination inhibition (HI) assay on 105 donkey serum samples. RESULTS: We demonstrated that H5N1 jumped from poultry to another mammalian host; donkeys. Phylogenetic analysis showed that the virus clustered within the lineage of H5N1 from Egypt, closely related to 2009 isolates. It harboured few genetic changes compared to the closely related viruses from avian and humans. The neuraminidase lacks oseltamivir resistant mutations. Interestingly, HI screening for antibodies to H5 haemagglutinins in donkeys revealed high exposure rate. CONCLUSIONS: These findings extend the host range of the H5N1 influenza virus, possess implications for influenza virus epidemiology and highlight the need for the systematic surveillance of H5N1 in animals in the vicinity of backyard poultry units especially in endemic areas.

Virulence and resistance determinants in Pseudomonas aeruginosa isolated from pericarditis in diseased broiler chickens in Egypt.

OBJECTIVE: This study was performed to probe the antimicrobial resistance and virulence genes profiling in Pseudomonas aeruginosa recovered from the cases of pericarditis in broiler chickens. MATERIALS AND METHODS: The samples (n = 250) collected from the cases of pericarditis in broiler chickens were bacteriologically examined. Antimicrobial susceptibility was tested by disc diffusion technique. The isolates were genotypically studied for the presence of antimicrobial resistance and virulence gene traits. Finally, the nucleotide sequence of representative resistance gene (mexR gene) and virulence genes (toxA and lasI genes) was analyzed. RESULTS: P. aeruginosa was isolated from 45 samples (18%). Antimicrobial susceptibility testing revealed multidrug resistance in most of the recovered P. aeruginosa isolates, whereas colistin and imipenem were the furthermost in vitro-sensitive antibiotics. Antimicrobial resistance genes, such as bla CTX, fox, and mexR, were prevalent in 100%, 80%, and 100% of the isolates, respectively. PCR confirmed virulence genes such as toxA, exoY, lasB, and lasI in 100%, 60%, 80%, and 80% of the isolates, respectively. Nucleotide sequence analysis of representative resistance gene (mexR gene) and virulence genes (toxA and lasI genes) revealed a high correlation between P. aeruginosa recovered from pericarditis in broiler chickens in the present study with PAO1 (reference strain) and with other sequences published on the GenBank representing different localities worldwide. CONCLUSION: It could be concluded that P. aeruginosa recovered from pericarditis in broiler chickens in the current study is highly virulent bacteria, resisting most of the therapeutic agents which not only bear hazards for poultry industry but also represent a public health concern.

Genotyping and reassortment analysis of highly pathogenic avian influenza viruses H5N8 and H5N2 from Egypt reveals successive annual replacement of genotypes.

Highly pathogenic (HP) H5N1, clade 2.2.1, and low pathogenic avian influenza (LPAI) H9N2 viruses, G1-B lineage, are endemic in poultry in Egypt and have co-circulated for almost a decade. Surprisingly, no inter-subtypic reassortment events have been reported from the field during that time. After the introduction of HPAIV H5N8, clade 2.3.4.4b, in Egyptian poultry in 2016, suddenly HP H5N2 reassortants with H9N2 viruses emerged. The current analyses focussed on studying 32 duck flocks, 4 broiler chicken flocks, and 1 turkey flock, suffering from respiratory manifestations with moderate to high mortality reared in two Egyptian governorates during 2019. Real-time RT-PCR substantiated the presence of HP H5N8 in 21 of the 37 investigated flocks with mixed infection of H9N2 in two of them. HP H5N1 was not detected. Full hemagglutinin (HA) sequencing of 10 samples with full-genome sequencing of three of them revealed presence of a single genotype. Very few substituting mutations in the HA protein were detected versus previous Egyptian HA sequences of that clade. Interestingly, amino acid substitutions in the Matrix (M2) and the Neuraminidase (NA) proteins associated with conferring both Amantadine and Oseltamivir resistance were present. Systematic reassortment analysis of all publicly available Egyptian whole genome sequences of HP H5N8 (n = 23), reassortant HP H5N2 (n = 2) and LP H9N2 (n = 53) viruses revealed presence of at least seven different genotypes of HPAI H5Nx viruses of clade 2.3.4.4b in Egypt since 2016. For H9N2 viruses, at least three genotypes were distinguishable. Heat mapping and tanglegram analyses suggested that several internal gene segments in both HP H5Nx and H9N2 viruses originated from avian influenza viruses circulating in wild bird species in Egypt. Based on the limited set of whole genome sequences available, annual replacement patterns of HP H5Nx genotypes emerged and suggested selective advantages of certain genotypes since 2016.

Efficacy of Live Attenuated Vaccine and Commercially Available Lectin Against Avian Pathogenic E. coli Infection in Broiler Chickens.

In this study, the protective efficacy of an E. coli live attenuated vaccine was compared to the preventive administration of lectin preparation before the challenge. Two hundred broiler chicks were divided into eight equal groups. The first group was used as a negative control group. Three groups were vaccinated at day 1 with the avian colibacillosis live vaccine of which one group served as a vaccinated nonchallenged group. Another two groups were treated with lectin product (0.5 mL/L drinking water) for three days before the challenge. The last two groups served as challenge control for either E. coli O78 or O125 strains. The challenge was conducted at three weeks of age with either homologous O78 or heterologous O125 E. coli strains, using 0.5 mL/bird of each avian pathogenic E. coli (APEC) strain (~108 colony forming units "CFU"/mL)/subcutaneously. The bodyweight and feed conversion ratios (FCR) were calculated for four weeks. Clinical signs and gross and histopathological lesions were scored at two and seven days post inoculation (dpi). The heart and liver of euthanized chickens at 2 dpi were removed aseptically and homogenized to evaluate pathogenic E. coli colonization. Results showed that live avian colibacillosis vaccine reduced mortalities and APEC colonization in the homologous challenge group but not in the heterologous challenge group. Lectin-treated groups showed 20% and 16% mortality after challenge with E. coli O78 and O125, respectively, and both groups showed performance parameters, clinical signs, and histopathological lesion scores comparable to the negative control group, with variable E. coli colonization of heart and liver. The study demonstrated the efficacy of live attenuated avian colibacillosis vaccine against homologous but not heterologous APEC challenge in broiler chickens. The lectin-containing products can be used as a preventive medication to reduce the clinical impacts of colibacillosis regardless of the challenge strain. Standardization of the evaluation parameters for APEC vaccines is recommended.

Emerging infectious bronchitis virus (IBV) in Egypt: Evidence for an evolutionary advantage of a new S1 variant with a unique gene 3ab constellation.

Infectious bronchitis virus (IBV), a gamma-coronavirus, causes infectious bronchitis (IB), a major respiratory disease of chicken. Its high mutation rate in conjunction with recombination of the RNA genome constantly creates IBV variants that are difficult to control by currently available vaccines. In this study, we addressed the question whether small-scale holdings might harbor IBV variants that serve as a reservoir for newly emerging variants. Egyptian IBV isolate EGY/NR725/2016 (NR725/16) from a small-scale broiler farm was assigned to genotype I, clade 23 (S1:GI-23), based on partial S1 gene sequences and corroborated by full genome sequencing. Analysis of the S1 gene established three subclades for historical IBV strains (S1:GI-23.1, S1:GI-23.2.1 and S1:GI-23.2.2) and confirmed NR725/16 as being part of a separate fourth subclade (S1:GI-23.3). Samples from the years 2018 and 2019 revealed that the new subclade prevails in Egypt, carrying fixed mutations within the hypervariable regions (HVR) 1-3 of the S1 protein that affect two neutralization sensitive epitopes at sites 294F, 297S and 306Y (48.2) and 329R (62.1). In addition, recombination was recognized in isolate NR 725/16, with intra-subtype mixing for the entire genes 3ab and E and inter-subtype mixing for the entire gene 6b with a close match to QX like viruses of genotype GI-19. Further analysis of gene 3ab detected the homologous gene pool to NR725/16 in samples from 2013 (3ab:C) and closely related 3ab genotypes in IBV Egyptian isolates from 2016, 2018 and 2019. These data prove a flourishing exchange between poultry holdings with a common gene pool. The continued circulation of viruses harboring genes S1:GI-23.3 and 3ab:C indicates an evolutionary advantage of this combination possibly by combining antigenic escape with modulated pathogenicity to facilitate IBV spread in the vaccinated poultry population in Egypt.

Sequence diversity of the haemagglutinin open reading frame of recent highly pathogenic avian influenza H5N1 isolates from Egypt.

The sequences encoding the haemagglutinin (HA) of twelve H5N1 isolates obtained in 2006 and 2007 from different avian species in backyard holdings and poultry farms in Egypt revealed amino acid variations across the polypeptide and also in the polybasic cleavage motif of three of the isolates from backyard poultry with one, so far, unique mutation in an isolate from a chicken. The HAs of two isolates (A/goose/Egypt/R4/2007, A/chicken/Egypt/R3/2007) collected on the same day in the same village from two neighbouring houses were found to differ from each other. Five out of the seven nucleotide exchanges in these two isolates were translationally silent, and two resulted in amino acid substitutions: one in the polybasic cleavage motif and the other in the signal peptide. Circulation of different H5N1 strains possessing considerable variations in backyard poultry, particularly domestic waterfowl, draws attention to the evolution of H5N1 subtypes in Egypt.

Experimental co-infection of infectious bronchitis and low pathogenic avian influenza H9N2 viruses in commercial broiler chickens.

In this study, commercial broilers were experimentally infected with single (classical IBV, variant IBV or AIV-H9N2) or mixed AIV-H9N2 with classical, variant or vaccine strains of IBV. Birds were monitored for clinical and pathological outcomes and virus shedding for 10days post infection (DPI). Clinical signs were limited to the respiratory tract in all challenged groups and varied from mild to moderate mouth breathing to severe respiratory signs with snorting sound and extended head. Mortalities were only recorded in mixed AIV-H9N2/variant IBV challenge group. AIV-H9N2 challenge caused tracheal petechial hemorrhage that progressed to tracheal congestion and caseation. In mixed AIV-H9N2/IBV vaccine challenge, severe tracheitis with bronchial cast formation was observed. In mixed AIV-H9N2/variant IBV challenge severe congestion of the tracheal mucosa and excessive exudates with a tendency to form tubular casts were observed. Kidney ureate deposition was only observed in variant IBV challenge group. Histopathologically, tracheal congestion, severe degeneration, and deciliation were noticed in all groups of mixed infection. Interestingly, hemorrhage and atrophy were observed in thymus gland of birds challenged with single AIV-H9N2 or mixed AIV-H9N2/IBV. There was no difference in the tracheal shedding level of variant IBV between single and mixed infected groups while classical IBV shedding increased in mixed infection group. Interestingly, the AIV-H9N2 showed constantly high shedding titers till 7DPI with variant or vaccine IBV co-infection. In conclusion, co-infection of IBV and AIV-H9N2 induced severe clinical outcome and high mortality. Also, IBV co-infection increased the shedding of AIV-H9N2 in experimentally infected birds.

Prevalence of avian respiratory viruses in broiler flocks in Egypt.

In this study, respiratory viral pathogens were screened using real-time RT-PCR in 86 broiler chicken flocks suffering from respiratory diseases problems in 4 Egyptian governorates between January 2012 and February 2014. The mortality rates in the investigated flocks ranged from 1 to 47%. Results showed that mixed infection represented 66.3% of the examined flocks. Mixed infectious bronchitis (IBV) and avian influenza (AI)-H9N2 viruses were the most common infection (41.7%). Lack of AI-H9N2 vaccination and high rates of mixed infections in which AI-H9N2 is involved indicate an early AI-H9N2 infection with a potential immunosuppressive effect that predisposes for other viral infections. High pathogenic AI-H5N1 and virulent Newcastle disease virus (vNDV) infections were also detected (26.7% and 8.1%, respectively). Interestingly, co-infection of AI-H9N2 with either AIV-H5N1 or vNDV rarely resulted in high mortality. Partial cell-mediated immunity against similar internal AI genes, as well as virus interference between AI and vNDV, could be an explanation for this. Highly prevalent IBV and AI-H9N2 were isolated and were molecularly characterized based on S1 gene hypervariable region 3 ( HVR3: ) and hemagglutinin gene (HA) sequences, respectively. IBV strains were related to the variant group of IBV with multiple mutations in HVR3. Though AI-H9N2 viruses showed low rate of evolution in comparison to recent strains, few amino acid substitutions indicative of antibody selection pressure were observed in the HA gene. In conclusion, mixed viral infections, especially with IBV and AI-H9N2 viruses, are the predominant etiology of respiratory disease problems in broiler chickens in Egypt. Further investigations of the role of AI, IBV, and ND viruses' co-infections and interference in terms of altering the severity of clinical signs and lesions and/or generating novel reassortants within each virus are needed.

Humoral antibody responses to different H5N1 and H5N2 vaccination regimes: implications for the development of autogenously based vaccines.

Whereas H5N1 vaccine and several H5N2 vaccines are commercially available and are used to control H5N1 outbreaks in some endemic countries, infections hit many vaccinated flocks. The following study was conducted to compare the efficacy of such vaccines and to assess their potential induction of antibodies against the haemagglutinin of local H5N1 isolate after single vaccination. The possible beneficiary effect of booster dose at different intervals was screened for both H5N1 vaccine as well as a selected H5N2 candidate. Differences in the serological immune response among native and cross breeds were also screened. No significant variations were detected between available commercial H5N1 and H5N2 vaccines after single vaccination. Two vaccination shots using H5N1 but not H5N2 vaccine were found to be superior to a single vaccination scheme, where chicks developed more conceivable antibody titers than in single vaccination program. There was considerable variation among chicken lines in the immune response to H5N1 vaccine: native breeds possessed the highest antibody titers as compared to other breeds.