Evaluating the effect of Spirulina platensis on the immune response of broiler chickens to various vaccines and virulent Newcastle disease virus challenge.

This study investigated the efficacy of co-administration of Spirulina platensis (SP) with vaccines on the immune response to Avian influenza (AI), Infectious bronchitis (IB), and Newcastle disease (ND), along with I/M challenging by virulent ND virus (vNDV) genotype VII. 126 one-day-old broiler chicks were allocated into six groups (21 birds/group with three replicates): G1: negative control; G2: positive control; G3: vaccinated, non-SP-supplemented; G4: vaccinated, SP-supplemented (0.1%); G5: vaccinated, SP-supplemented (0.3%); and G6: vaccinated, SP-supplemented (0.5%). G2-6 were challenged with a velogenic NDV genotype VII virus. Dietary SP administration prevented the ND-induced mortality compared to G2 (52.4%) and G3 (14.3%), in addition to alleviating the clinical disease. G3-6 showed significant improvement in body weight loss% and FCR during two weeks post vNDV challenge (pc), and the overall FCR (2.64 ± 0.28, 1.56 ± 0.03, 1.60 ± 0.05, 1.53 ± 0.04, and 1.54 ± 0.03 for G2, 3, 4, 5 and 6, respectively) (P<0.05). On the challenging day, the ND-HI titer (log2) of G3 (5.44 ± 0.24) was numerically higher than G6 (4.20 ± 0.55) and lower than G4 (6.10 ± 0.34) and G5 (6.00 ± 0.28). On the 10th day pc, ND-HI titer in G4-6 was numerically lower in a dose-dependent manner than that of G3, suggesting an antiviral efficacy of SP. G4-6 had lower viral shedding titer than G2 and G3 (P<0.05). In G3-6, viral shedding was reduced by 15, 27, 24, and 33.6%, respectively. In addition, the histopathological lesions in the trachea, lung, and spleen were severe in G2, moderately reduced in G3, and more relieved in G4-6. At three weeks after vaccination, the HI antibody titer of AIH5 was significantly higher after SP administration, especially at the 0.3% level, compared to the vaccine alone (P<0.05), demonstrating an immune-stimulating effect. In conclusion, dietary administration of SP, particularly a dose of 0.3%, for vaccinated chickens against NDV exerted an antiinflammatory and antiviral effects by preventing deaths, alleviating clinical disease and weight loss, and decreasing viral shedding post heterologous NDV challenge.

Pathogenicity of H5N8 avian influenza virus in chickens and in duck breeds and the role of MX1 and IFN-α in infection outcome and transmission to contact birds.

This study examined the pathogenicity, immunogenicity, and transmission potential of the H5N8 HPAI clade 2.3.4.4b virus in three breeds of ducks and in broiler chickens. Chickens, Muscovy, Pekin, and Mallard ducks (n = 10) received a dose of 6 log10 EID50 of HPAIV H5N8 directly. Nine contact chickens were introduced to each group on the day of infection. All infected chickens died, with MDT of 7.6 days. Muscovy and Pekin ducks died by 11.1% and 10%, respectively, with MDTs of 7 and 6 days. No Mallards died but showed more severe clinical disease than Pekin ducks. Mallards had the highest MX1 gene expression in the lung and spleen and IFN-α in the spleen. MX1 expression levels were lower in the spleen and lung of Pekin ducks, in the spleen of chickens and in the lung of Muscovy ducks than in noninfected controls. However, viral shedding was higher in ducks than in chickens and was highest in Mallards. 66.7% of chickens placed in contact with infected chickens died and 77.8% of in-contact chickens to infected three duck breeds died. In conclusion, there was a diversity in sensitivity and immunogenicity for HPAIV H5N8 among duck breeds, resulting in diverse infection outcomes and transmissibility to contacts. This study provides duck/chicken interface models for HPAIV transmission to poultry.

Examination of the protective efficacy of two avian influenza H5 vaccines against clade 2.3.4.4b H5N8 highly pathogenic avian influenza virus in commercial broilers.

The highly pathogenic avian influenza (HPAI) H5N8 virus of clade 2.3.4.4 was detected in 2017 in Egypt, which is one of the few countries using vaccination as a control strategy in poultry farms. This study was conducted to evaluate the efficacy of the commercial recombinant turkey herpes virus-H5 (rHVT-H5) vaccine (clade 2.2), alone or in combination with commercial inactivated reverse genetically engineered H5N1 vaccine (rgH5N1) (clade 2.2), in preventing the genetically distinct HPAI H5N8 virus of clade 2.3.4.4b in commercial broiler chickens. Four experimental groups of chickens were used as follows: G1, non-vaccinated and non-challenged; G2, non-vaccinated and challenged; G3, vaccinated with rHVT-H5; and G4, prime-boost vaccinated with rHVT-H5/rgH5N1. For challenge with the Egyptian HPAI H5N8 (2.3.4.4b) virus, the groups were divided into two subgroups (A and B); chickens in subgroups A were challenged at the age of 28 days, whereas those in subgroups B were challenged at the age of 35 days. Results showed that a protective efficacy (survival rate) of 40%-50% was obtained in the vaccinated subgroups A. By delaying challenge for 1 week (subgroups B), a single rHVT-H5 vaccination provided 80% protection, whereas prime-boost vaccination induced full protection and reduced viral shedding very efficiently (1/10 birds and only detected on the 3rd day post challenge) against HPAI H5N8 virus (2.3.4.4b). Moreover, body weight loss improved from 31.39% and 43.65% in G3A and G4A, respectively, to 16.34% and 7.7% in G3B and G4B, respectively. The HI titers obtained in G3A and G4A on the challenge day (28th d) using H5N8 antigen were 3 and 3.75 log2 (p > 0.05), respectively, whereas those in G3B and G4B on the challenge day (35th d) were 6.25 and 6 log2 (p > 0.05), respectively, which increased post-challenge in all vaccinated subgroups. Therefore, the dual use of vectored rHVT-H5 and inactivated rgH5N1 vaccines in the vaccination schedule in poultry farms is the most efficient tool for preventing the disease (mortality and viral shedding) caused by the genetically distinct virus (clade 2.3.3.4b HPAI H5N8) in combination with strict biosecurity and sanitary measures.

Effect of essential oils on the immune response to some viral vaccines in broiler chickens, with special reference to Newcastle disease virus.

This trial assessed the efficacy of a commercial essential oil (EO) product on the immune response to vaccination against Newcastle disease (ND) and subsequent challenge with virulent ND virus genotype VII (vNDv genotype VII) by using the following experimental groups of broiler chickens (Each group had 21 birds with 3 replicates in each, n = 7): NC (negative control), PC (positive control), VC (vaccinated), and VTC (vaccinated and treated with EOs). Moreover, in a trial to study the effect of EOs on vNDv genotype VII in vivo as a preventive or therapeutic measure, 2 additional ND-vaccinated groups were used (PRV: medicated 1 D before vNDv challenge for 5 D; and TTT: medicated 2 D after vNDv challenge for 5 D). In addition, the immune-modulatory effect of EOs on the avian influenza (AI), infectious bronchitis (IB), and infectious bursal disease (IBD) vaccines was assessed through the serological response. The use of EOs along with administration of ND vaccines (VTC) revealed a lower mortality rate (42.86%), clinical signs, and postmortem lesion score (11) than ND vaccines alone (VC) (52.28% mortality and score 15), in addition to lower hemagglutination inhibition (P < 0.05) (6.5 ± 0.46) and viral shedding (10 log 2.28 ± 0.24) titres 1 wk after challenge in comparison with VC (8.63 ± 0.65 and 10 log 3.29 ± 0.72, respectively). Nevertheless, the EOs mixture (VTC) (1952 ± 28.82) did not significantly (P > 0.05) improve growth performance compared with the nontreated birds (NC and VC) (1970 ± 19.56 and 1904 ± 38.66). EOs showed an antiviral effect on vNDv in vivo (in chickens) as a preventive measure (PRV) as well as some therapeutic effect (TTT) through decreasing the viral shedding titres (loNC0), mortality rate, and severity of clinical signs and postmortem lesions, in addition to serum malondialdhyde level. Regarding the other viruses, the EOs mixture did not improve the immune response to the AI and IB vaccines but significantly (P < 0.05) increased the ELISA antibody titre for IBD virus at the 28th D of age (2,108 ± 341.05). The studied EOs mixture showed an immune-stimulating response to ND and IBD vaccines, antiviral effect against ND virus, especially if administered before the challenge; however, it did not have a growth-promoting effect.

Incidence, Pathotyping, and Antibiotic Susceptibility of Avian Pathogenic Escherichia coli among Diseased Broiler Chicks.

A total of 54 broiler flocks during the first two weeks of life was used to investigate the incidence of avian pathogenic E. coli in Egypt; 28 isolates (51.85%) were revealed by colony morphology and biochemical identification which then investigated for their serogroups and only 18/28 isolates were serotyped. The most prevalent serotypes were O115, O142, O158, O55, O125, O114, O27, O20, and O15. By application of polymerase chain reaction (PCR), 83.3% (15/18) of the serotyped isolates were confirmed to be E. coli, and 93.3% (14/15), 46.6% (7/15), and 20% (3/15) of isolates harbored the iss, iutA, and fimH genes, respectively. Virulence testing of the selected 13 APEC isolates on the specific-pathogen-free (SPF) chicks revealed them to be highly virulent (15.4%), moderately virulent (23.1%), and avirulent (61.5%); however, all isolates (100%) were extremely virulent towards SPF embryonated chicken eggs. Antibiotic resistance (100% of isolates (n = 13)) was observed for ampicillin, amoxycillin-clavulanic acid, and tetracyclines, colistin (92.31%; 12/13), doxycycline and spiramycin (84.62%; 11/13), florfenicol (69.23%; 9/13), cefotaxime (61.54%; 8/13), and ciprofloxacin (53.85%; 7/13). The highest percentage of sensitivity (53.85% of isolates; 7/13) was recorded for ofloxacin and enrofloxacin followed by gentamycin (46.15%; 6/13). The results suggest that the diagnosis of APEC with PCR is rapid and more accurate than traditional methods for E. coli identification; moreover, the presence or absence of iss, iutA, and/or fimH genes is not an indicator of in vivo pathogenicity of APEC. Thus, further studies, including a wider range of virulence genes and gene sequencing, are required. In addition, serotyping has no effect on the virulence of APEC.

Isolation, conventional and molecular characterization of Salmonella spp. from newly hatched broiler chicks.

Salmonella is an important pathogen for poultry production as well as for human due to zoonotic importance. It has more than 2600 identified serovars despite of this identification and classification of Salmonella isolates into different serovars is critical for study of incidence and surveillance. This study investigates the epidemiology and molecular characterization of Salmonella isolates in broiler chicks during 1st week of life. A total of (n = 1000) samples including liver, intestine, yolk sac, spleen and heart blood were collected from El-Gharbia, El-Behera, Kafr-Elshikh, Alexandria, Marsamatroh Provinces in Egypt and tested through bacteriological, biochemical, serological and molecular examinations. Incidence of Salmonella was demonstrated on 75 positive samples from 1000 samples and the predominance of Salmonella that isolated from internal organs of newly hatched chicks was highest from yolk sacs (10%), liver and intestines (9%) followed by the spleen (7.5%) then heart blood (2%). Serotyping of the isolated strains using slide agglutination test revealed that 24 isolates belonging to S. enteritidis (1,9,12 g.m 1,7), while, 14 isolates belonging to S. virchow (6,7 r 1,2), in addition to, 12 isolates belonging to S. typhimurium (1,4,5,12.i.1,2) and 8 isolates belonging to S. kentucky (6,8.I,z). Enterobacterial Repetitive Intergenic Consensus (ERIC) PCR revealed that two S. enteriditis isolates were identical and one isolate differ by 40%, while two S. typhimurium isolates were identical by 80% and one isolate was similar by 20% to the other two isolates, in addition, two S. virchow isolates were identical by 80% and the two S. kentucky isolates were different.

Muscovy ducks infected with velogenic Newcastle disease virus (genotype VIId) act as carriers to infect in-contact chickens.

This work was designed to study the dynamics of transmission of Newcastle disease virus (NDV), genotype VIId, from Muscovy ducks (Cariana moscata) infected either by intramuscular (IM) or intranasal (IN) inoculation, to in-contact broiler chickens (Gallus gallus). IM-infected Muscovy ducks (G1d) exhibited only 5% mortality, and the concentration of virus shed from the cloaca was greater and for longer period than virus shed from the trachea. In contrast, IN-infected ducks (G2d) exhibited no mortality, and virus shedding from the trachea was higher than that from the cloaca starting from 4 days post infection (dpi) and continued up to 16 dpi, while in IM-infected ducks (G1d), tracheal shedding stopped at 11 dpi. Chickens in contact with IM-infected and IN-infected ducks, G1c and G2c, respectively, not only developed severe clinical symptoms and death (80% and 20% mortality, respectively), but also shed the virus at higher concentrations than infected ducks. G1c chickens had higher viral shedding titers in both the trachea and cloaca than G2c chickens until 11 dpi. All broiler chickens infected by IM route (G3c) died, while the IN route of infection resulted in lower mortality (70%) in G4c. Generally, all IM-infected birds produced an earlier and higher level of NDV hemagglutination inhibition (HI) antibody titer, along with higher rates and shorter periods of viral shedding than those infected by the intranasal route. Our conclusion is that Muscovy ducks are efficient carriers of NDV-genotype VIId and transmit the virus to contact chickens.

Comparative Evaluation of HVT-IBD Vector, Immune Complex, and Live IBD Vaccines against vvIBDV in Commercial Broiler Chickens with High Maternally Derived Antibodies.

Infectious bursal disease (IBD) causes increased mortality and severe immunosuppression in commercial chickens. Currently, vaccination mainly used to control IBD. In this study, Group A (n = 30) received the HVT-IBD vector vaccine (Vaxxitek®) s/c and Group B (n = 30) received the immune complex vaccine (Bursa-Plex®) s/c at 1 day of age. Group C (n = 30) received a single dose of intermediate plus vaccine (228E) through the eye-drop route at 14 days of age. Group D (n = 30) was vaccinated twice with the intermediate vaccine (D78) at 12 and 22 days of age by eye-drop. Group E (n = 30) had the same treatment as group D along with the IBD killed vaccine (Nobilis G®) at 5 days of age. The PC (n = 20) and NC (n = 20) groups were non IBD vaccinated birds either challenged or not with vvIBDV, respectively; 20 chicks from each group were challenged with vvIBDV at 4 weeks of age. Based on clinical signs, postmortem gross lesions, histopathological changes, mortality rate, feed conversion rate, serology, bursal and spleen indices, the HVT-IBD vector vaccine administered was found to be safer and provided better protection against the vvIBDV challenge. The use of a killed IBD vaccine at an earlier age in broilers strengthened the protection induced by double doses of intermediate vaccines in broilers with high maternally derived antibodies against the vvIBDV challenge.

Co-infections, genetic, and antigenic relatedness of avian influenza H5N8 and H5N1 viruses in domestic and wild birds in Egypt.

A total of 50 poultry farms of commercial broilers (N = 39) and commercial layers (N = 11) suffered from respiratory problems and mortality during the period from January 2016 to December 2017 were investigated. Also, samples were collected from quail (N = 4), Bluebird (Sialis, N = 1), and Greenfinch (Chloris chloris, N = 1) for analysis. Respiratory viral pathogens were screened by PCR and positive samples were subjected to virus isolation and genetic identification. Antigenic relatedness of isolated avian influenza (AI) H5 subtype was evaluated using cross-hemagglutination inhibition. Results revealed that the incidence of single virus infections in commercial broilers was 64.1% (25/39), with the highest incidence for ND (33.3%) and H9N2 (20.5%), followed by H5N1 (7.7%) and H5N8 (2.7). Meanwhile, H9N2/ND mixed infection was the most observed case (7.7%). Other mixed infections H5N1/ND, H5N1/H9N2/ND, H5N1/H9N2/ND/IB, H9N2/IB, and H9N2/ILT were also observed (2.6% each). In commercial layers, H5N1 and ILT were the only detected single infections (18.1% each). Mixed H9N2/ND was the most predominant infection in layers (27.3%). Other mixed infections of H9N2/IB, H5N1/H5N8/H9N2, and H9N2/ND/IB were observed in 3 separate farms (9.1% each). The H5N8 virus was detected in one quail farm and 2 out of 3 wild bird's samples. Partial HA gene sequence analysis showed the clustering of the selected AI H5N8 within the 2.3.4.4 clade, while H5N1 clustered with the clade 2.2.1.2. Interestingly, the H5N8 isolated from chickens possessed 6 amino acids substitutions at HA1 compared to those isolated from wild birds with low antigenic relatedness to AI H5N1 clades 2.2.1 or 2.2.1.2. In conclusion, mixed viral infections were observed in both broiler and layer chickens in Egypt. The detected triple H5N1, H9N2, and H5N8 influenza co-infection raises the concern of potential AI epidemic strain emergence. The low genetic and antigenic relatedness between AI H5N1 and H5N8 viruses suggest the need for modification of vaccination strategies of avian influenza in Egypt along with strict biosecurity measures.

The Simultaneous Administration of a Probiotic or Prebiotic with Live Salmonella Vaccine Improves Growth Performance and Reduces Fecal Shedding of the Bacterium in Salmonella-Challenged Broilers.

Salmonellosis is one of the most important bacterial diseases in poultry, causing heavy economic losses, increased mortality and reduced production. The aim of this study was the comparative efficacy of a commercial probiotic and/or prebiotic with a live attenuated Salmonella Enteritidis (SE) vaccine on the protection of broiler chickens from SE challenge. The efficacy of probiotic or prebiotic products, as well as a live Salmonella Enteritidis (SE) vaccine at the 7th day of age, administered via drinking water, were evaluated for clinical protection and effects on growth performance of broiler chickens experimentally challenged with SE at the 28th day of age. The use of probiotic or prebiotic simultaneously with the live Salmonella vaccine can diminish the negative effect of live vaccine growth performance, reducing mortality rate, fecal shedding, and re-isolation of SE from liver, spleen, heart and cecum. The use of probiotic or prebiotic simultaneously with the application of the live Salmonella vaccine is a good practice to diminish the negative effect of the harmful bacteria and improve the growth performance of broilers. Thus, further studies may be carried out with layers and breeders.