Genotype IX Newcastle disease virus isolated from wild birds is attenuated by hemagglutinin-neuraminidase mutation

Thermostable Newcastle disease virus (NDV) vaccines have been used widely to protect village chickens against Newcastle disease, due to their decreased dependence on cold chain for transport and storage. However, the genetic basis underlying the NDV thermostability is poorly understood. In this study, we generated chimeric viruses by exchanging viral genes between the thermostable TS09-C strain and thermolabile LaSota strain using reverse genetics technology. Evaluations of these chimeric NDVs demonstrated that the thermostability of NDV was dependent on the origin of HN protein. Chimeras bearing the HN protein derived from thermostable virus exhibited a thermostable phenotype, and vice versa. Both hemagglutinin and neuraminidase activities of viruses bearing the TS09-C HN protein were more thermostable than those containing LaSota HN protein. Furthermore, the newly developed thermostable virus rLS-T-HN, encoding the TS09-C HN protein in LaSota backbone, induced significantly higher antibody response than the TS09-C virus, and conferred complete protection against virulent NDV challenge. Taken together, the data suggest that the HN protein of NDV is a crucial determinant of thermostability, and the HN gene from a thermostable NDV could be engineered into a thermolabile NDV vaccine strain for developing novel thermostable NDV vaccine.

This study aimed at determining the possible exposure of wild birds to avian influenza (AI), infectious bronchitis (IB) and Newcastle disease (ND) viruses. Apparently healthy species of free flying wild birds were captured using locally-made baited traps set at strategic watering and feeding locations and in poultry farms. Few species of captive wild birds in households and live bird markets (LBMs) were also sampled. Sera from blood samples collected were analyzed for antibodies to AI, IB and ND viruses using enzyme linked immunorsorbent assay (ELISA). Out of the 209 sera analysed, Bubulcus ibis was 24%, 70% and 27% while Psittacus erithacus was 7%, 21% and 7% positive for antibodies to AI, IB and ND viruses, respectively. Branta canadensis , was 35% and 64% positive for antibodies to AI and IB viruses. Balearica regulorum and Numidia maleagris were 100% and 9% positive to AI virus antibodies. Free flying birds were 19 (15%), 57 (45%) and 27 (21%) positive while captive wild birds were 11%, 20% and 14% positive to AI, IB and ND viruses antibodies, respectively. The results of this study confirm that wild birds were exposed to AI, IB and ND viruses. There was co-exposure of some wild bird species to AI, IB and ND viruses. These birds could possibly serve as carriers and disseminators of AI, IB and ND to poultry. Therefore, control measures against these important poultry diseases should include incursion of wild birds. Keywords: Avian influenza, Infectious bronchitis, Newcastle disease, Nigeria, Wild birds

Different infectious bursal disease virus (IBDV) live vaccines (intermediate, intermediate plus) were compared for their immunosuppressive abilities in specific-pathogen-free (SPF) layer-type chickens or commercial broilers. The Newcastle disease virus (NDV) vaccination model was applied to determine not only IBDV-induced immunosuppression but also bilateral effects between IBDV and NDV. None of the IBDV vaccines abrogated NDV vaccine-induced protection. All NDV-vaccinated SPF layers and broilers were protected against NDV challenge independent of circulating NDV antibody levels. Sustained suppression of NDV antibody development was observed in SPF layers, which had received the intermediate plus IBDV vaccine. We observed a temporary suppression of NDV antibody development in broilers vaccinated with one of the intermediate, as well as the intermediate plus, IBDV vaccines. Different genetic backgrounds, ages, and residual maternal antibodies might have influenced the pathogenesis of IBDV in the different types of chickens. Temporary suppression of NDV antibody response in broilers was only seen if the NDV vaccine was administered before and not, as it was speculated previously, at the time the peak of IBDV-induced bursa lesions was detected. For the first time, we have demonstrated that the NDV vaccine had an interfering effect with the pathogenesis of the intermediate as well as the intermediate plus IBDV vaccine. NDV vaccination enhanced the incidence of IBDV bursa lesions and IBDV antibody development. This observation indicates that this bilateral effect of an IBDV and NDV vaccination should be considered in the field and could have consequences for the performance of broiler flocks.

Simple SummaryNewcastle disease virus (NDV) is a highly contagious viral disease affecting a wide range of avian species. The disease can be particularly virulent in chickens, resulting in high mortality and morbidity. In this study, we characterized velogenic NDV sub-genotype VII.1.1 from wild birds and assessed its pathogenicity in susceptible chickens. One hundred wild birds from the vicinity of poultry farms with a history of NDV infection were examined clinically. Pooled samples from the spleen, lung, and brain were screened using real-time reverse transcriptase polymerase chain reaction (RRT-PCR) and reverse transcriptase polymerase chain reaction (RT-PCR) to detect the NDV F gene fragment, and phylogenetic analysis was carried out for identification of the genetic relatedness of the virus. Chickens were infected with the strains identified, and the major histopathological changes were assessed. Interestingly, NDV was detected in 44% of cattle egret samples and 26% of house sparrow samples by RRT-PCR, while RT-PCR detected NDV in 36% of cattle egrets examined and 20% of house sparrow samples. Phylogenetic analysis revealed close identity, of 99.7–98.5% (0.3–1.5% pairwise distance), between the isolates used in our study and other Egyptian class II, sub-genotype VII.1.1 NDV strains. Histopathological examination identified marked histopathological changes that are consistent with NDV. These findings provide interesting data in relation to the detection of NDV sub-genotype VII.1.1 in wild birds and reveal the major advantages of the combined use of molecular and histopathological methods in the detection and characterization of the virus. More research is needed to determine the characteristics of this contagious disease in the Egyptian environment.Newcastle disease (ND) is considered to be one of the most economically significant avian viral diseases. It has a worldwide distribution and a continuous diversity of genotypes. Despite its limited zoonotic potential, Newcastle disease virus (NDV) outbreaks in Egypt occur frequently and result in serious economic losses in the poultry industry. In this study, we investigated and characterized NDV in wild cattle egrets and house sparrows. Fifty cattle egrets and fifty house sparrows were collected from the vicinity of chicken farms in Kafrelsheikh Governorate, Egypt, which has a history of NDV infection. Lung, spleen, and brain tissue samples were pooled from each bird and screened for NDV by real-time reverse transcriptase polymerase chain reaction (RRT-PCR) and reverse transcriptase polymerase chain reaction (RT-PCR) to amplify the 370 bp NDV F gene fragment. NDV was detected by RRT-PCR in 22 of 50 (44%) cattle egrets and 13 of 50 (26%) house sparrows, while the conventional RT-PCR detected NDV in 18 of 50 (36%) cattle egrets and 10 of 50 (20%) of house sparrows. Phylogenic analysis revealed that the NDV strains identified in the present study are closely related to other Egyptian class II, sub-genotype VII.1.1 NDV strains from GenBank, having 99.7–98.5% identity. The pathogenicity of the wild-bird-origin NDV sub-genotype VII.1.1 NDV strains were assessed by experimental inoculation of identified strains (KFS-Motobas-2, KFS-Elhamoul-1, and KFS-Elhamoul-3) in 28-day-old specific-pathogen-free (SPF) Cobb chickens. The clinical signs and post-mortem changes of velogenic NDV genotype VII (GVII) were observed in inoculated chickens 3 to 7 days post-inoculation, with 67.5–70% mortality rates. NDV was detected in all NDV-inoculated chickens by RRT-PCR and RT-PCR at 3, 7, and 10 days post-inoculation. The histopathological findings of the experimentally infected chickens showed marked pulmonary congestion and pneumonia associated with complete bronchial stenosis. The spleen showed histocytic cell proliferation with marked lymphoid depletion, while the brain had malacia and diffuse gliosis. These findings provide interesting data about the characterization of NDV in wild birds from Egypt and add to our understanding of their possible role in the transmission dynamics of the disease in Egypt. Further research is needed to explore the role of other species of wild birds in the epidemiology of this disease and to compare the strains circulating in wild birds with those found in poultry.

This study was carried out to investigate the immunomodulating activity of β-glucanwhich extracted from the cell wall of Saccharomyces cerevisiae.β-glucan of 225µg/ml wassupplemented in drinking water of broiler chicken vaccinated with Newcastle disease virus(NDV) vaccine. The parameters of the immunomodulating activity employed were the bodyweight, Phagocytic activity and determination of antibody titer to NDV vaccine. Chicks oneday old (120), were divided into four equal groups, G1 treated with β-glucan for six weeksand vaccinated with NDV vaccine; G2 treated with β-glucan for three weeks and vaccinatedwith NDV vaccine; G3 not given β-glucan but vaccinated with NDV vaccine (control group);while G4 was not treated with β-glucan and not vaccinated with NDV vaccine (secondcontrol). The results of the body weight indicated that; there were significant differences (P<0.05) between treated groups (G1 and G2) compared to the control groups (G3 and G4) at21,28 and 35 day of age. The results of phagocytic activity showed that treated group hadsignificantly (P< 0.05) higher clearance of carbon particles from blood circulation than did thecontrol groups; and the antibody titer to NDV showed significant differences (P<0.05)between treated and non treated groups at 14d and 28d. The data presented in this studycontribute for the first time in Iraq; that β –glucan given via drinking water to chicks from dayone for 35 days improves the immune responses and body weight.

This work was carried out to investigate the effectiveness of selection for high immune response against Newcastle Disease Virus (NDV) vaccine in local Egyptian chicken strain (Dokki4) for two generations. Two lines (selected and control) of Dokki-4 strain were used; in the selected line, chicks were selected according to its immune response against NDV vaccine. In the control line, chicks were randomly chosen. The population families were vaccinated against the common prevalent diseases according to the vaccination programs in the Sakha Animal Production Research Station, Kafr El-Sheikh. Results of selection for high immune response against NDV vaccine showed that; selection was efficient in native Egyptian chicken strain (Dokki-4) for increasing antibody titer levels in the high selected line when compared with the control line (P < 0.05) in two generations. Highly significant differences between generation and line were detected. Sex differences in immune response to NDV vaccine were significant (P < 0.05) for the overall averages a long all generations of selection. Female’s titers were higher than those of corresponding males of the same line and generation. Realized response per generation in the selected line was 16.76 and 60.18 for male and 18.54 and 69.50 for female in the first and second generations respectively. This study conducted that the cumulative response for high antibody titers responses against NDV vaccine was 76.94 for male and 88.04 for female after two generations of selection. There were significant differences in livability percentage between high and control lines for antibody response; where high line had higher livability than the control line. The high selected line showed a higher survival rate post challenge, with significant difference (P < 0.05) than the control group in the first week (92.86 % and 86.11 %) and the second weeks (87.69 % and 77.42 %) of observing vaccinated birds. It could be concluded that, genetic selection for high antibody titers against NDV vaccine in Dokki-4 strain improved immune response at different ages under investigation with increasing livability percentages and disease resistance of the birds through two successive generations of selection.

Newcastle Disease Virus (NDV) is an infectious disease that infect many kinds of wild and domesticated birds. Infection of NDV become a massive problem for poultry industry around the world especially in Indonesia. Vaccination is an effort to prevent the infection of NDV in poultry. NDV vaccine that used in Indonesia is a conventional life vaccine from LaSota and B1 strains. These type of vaccine is 21%-23% genetically distinct with the virus that spread in the environment. The antibody protection provided by the vaccine is not effective. Therefore, vaccination with new local NDV strain is needed to prevent the NDV infection in Indonesia. The previously study research reported that the local isolate of NDV from Kulon Progo, Indonesia has been isolated. Fusion (F) protein encoding gene that has been inserted into pBT7-N-His expression p lasmid which isolated from clone C-2a of E. coli, then it was expressed by the Cell-free protein expression system. The aim of this study was to confirm whether clone C-2a of E.coli carrying a recombinant plasmid pBT7-N-His-Fusion NDV and to express a recombinant F protein of NDV in-vitro from expression plasmid by cell-free protein expression system. This work started by detection of recombinant plasmid pBT7-N-His-Fusion NDV by DNA plasmid extraction followed by agarose gel electrophoresis. The recombinant F protein was in-vitro expressed by cell-free protein expression kit. The expressed F protein of NDV then was visualized by SDS-PAGE and Westernblott to analyse the expression of NDV recombinant F protein. It confirmed that clone C-2a of E. coli contained plasmid pBT7-N-His (4.001 bp) inserted by recombinant F protein of NDV gene (642 bp). The visualisation of expressed recombinant F protein by SDS-PAGE and Westernblott showed the NDV recombinant F protein was a specific protein fragment with molecular weight of 25,6 kDa..

Worldwide, wild birds are frequently suspected to be involved in the occurrence of outbreaks of different diseases in captive-bred birds although proofs are lacking and most of the dedicated studies are insufficiently conclusive to confirm or characterize the roles of wild birds in such outbreaks. The aim of this study was to assess and compare, for the most abundant peridomestic wild birds, the different exposure routes for avian influenza and Newcastle disease viruses in conservation breeding sites of Houbara bustards in the United Arab Emirates. To do so, we considered all of the potential pathways by which captive bustards could be exposed to avian influenza and Newcastle disease viruses by wild birds, and ran a comparative study of the likelihood of exposure via each of the pathways considered. We merged data from an ecological study dedicated to local wild bird communities with an analysis of the contacts between wild birds and captive bustards and with a prevalence survey of avian influenza and Newcastle disease viruses in wild bird populations. We also extracted data from an extensive review of the scientific literature and by the elicitation of expert opinion. Overall, this analysis highlighted those captive bustards had a high risk of being exposed to pathogens by wild birds. This risk was higher for Newcastle disease virus than avian influenza virus, and House sparrows represented the riskiest species for the transmission of both viruses through direct exposure from direct contact with an infectious bird that got inside the aviary and indirect exposure from consumption of water contaminated from the faeces of an infected bird that got inside the aviary for Newcastle disease virus and avian influenza virus, respectively. These results also reaffirm the need to implement biosecurity measures to limit contacts between wild and captive birds and highlight priority targets for a thoughtful and efficient sanitary management strategy.

Newcastle disease virus (NDV) infects a wide range of bird species worldwide and is of importance to the poultry industry. Although certain virus genotypes are clearly associated with wild bird species, the role of those species in the movement of viruses and the migratory routes they follow is still unclear. In this study, we performed a phylogenetic analysis of nineteen NDV sequences that were identified among 21,924 samples collected from wild and synanthropic birds from different regions of Ukraine from 2006 to 2015 and compared them with isolates from other continents. In synanthropic birds, NDV strains of genotype II, VI, VII, and XXI of class II were detected. The fusion gene sequences of these strains were similar to strains detected in birds from different geographical regions of Europe and Asia. However, it is noteworthy to mention the isolation of vaccine viruses from synanthropic birds, suggesting the possibility of their role in viral transmission from vaccinated poultry to wild birds, which may lead to the further spreading of vaccine viruses into other regions during wild bird migration. Moreover, here we present the first publicly available complete NDV F gene from a crow (genus Corvus). Additionally, our phylogenetic results indicated a possible connection of Ukrainian NDV isolates with genotype XXI strains circulating in Kazakhstan. Among strains from wild birds, NDVs of genotype 1 of class I and genotype I of class II were detected. The phylogenetic analysis highlighted the possible exchange of these NDV strains between wild waterfowl from the Azov-Black Sea region of Ukraine and waterfowl from different continents, including Europe, Asia, and Africa.

To treat patients with stage I-IV malignant tumors of digestive tract using autologous tumor cell vaccine and NDV (Newcastle disease virus) vaccine, and observe the survival period and curative effect. 335 patients with malignant tumors of digestive tract were treated with autologous tumor cell vaccine and NDV vaccine. The autologous tumor cell vaccine received were assigned for long-term survival observation. While these failed to obtain the autologous tumor tissue were given with NDV vaccine for a received short-term observation on curative effect. The colorectal cancer patients treated with autologous tumor cell vaccine were divided into two groups: the controlled group (subjected to resection alone) (n=257), the vaccine group (subjected to both resection and immunotherapy) (n=310). 25 patients treated with NDV immunotherapy were all at stage IV without having resection. In postoperation adjuvant therapy patients, the 5, 6 and 7-year survival rates were 66.51 %, 60.52 %, 56.50 % respectively; whereas in patients with resection alone, only 45.57 %, 44.76 % and 43.42 % respectively. The average survival period was 5.13 years (resection alone group 4.15 years), the median survival period was over 7 years (resection alone group 4.46 years). There were significant differences between the two groups. The patients treated with resection plus vaccine were measured delayed-type hypersensitivity (DTH) reactions after vaccination, (indurative scope >5 mm). The magnitude of DTH was related to the prognosis. The 5-year survival rate was 80 % for those with indurations greater than 5 mm, compared with 30 % for those with indurations less than 5 mm. The 1-year survival rate was 96 % for 25 patients treated with NDV immunotherapy. The total effective rate (CR+PR) was 24.00 % in NDV immunotherapy; complete remission (CR) in 1 case (4.00 %), partial remission (PR) in 5 cases (20.00 %), stabilizedin in 16 cases (64.00 %), progression (PD) in 1 case (4.00 %). After NDV vaccine immunotherapy, the number of NK cell increased and immune function imporved obviously. The autologous tumor cell vaccine and NDV vaccine can prolong the patients' life. NDV vaccine is notably effective for short-term with promotion of quality of life and can be used whenever necessary with good prospects.

Phylogenetic characterization and virulence of two Newcastle disease viruses isolated from wild birds in China

Protection of chickens against overt clinical disease and determination of viral shedding following vaccination with commercially available Newcastle disease virus vaccines upon challenge with highly virulent virus from the California 2002 exotic Newcastle disease outbreak

This paper describes two experiments. In each experiment, 1-day-old specific pathogen free chicks were divided into three groups. In Experiment 1—[avian metapneumo virus (aMPV) challenge]—one group served as unvaccinated controls; the second group was vaccinated with live aMPV (subtype B) vaccine only, and the third group received the aMPV vaccine in combination with live Newcastle disease virus (NDV) vaccine (VG/GA strain). Oropharyngeal swabs, tissues and blood samples were collected before and after challenge with a virulent subtype aMPV at 21 days post vaccination. Chicks were monitored for post-challenge clinical signs. Swabs and tissues were examined for the detection of challenge aMPV by virus isolation and by reverse-transcriptase polymerase-chain reaction. Sera were assayed for antibodies against aMPV and NDV. The single and combined vaccinated chicks were all protected against clinical signs and no challenge virus was isolated from either of the vaccinated-challenged groups. In Experiment 2 (NDV challenge), as in Experiment 1, chicks were divided into three groups where one group remained as unvaccinated control and the other two groups were vaccinated as above, except that the second group received live NDV vaccine only, instead of aMPV. At 21 days post vaccination, 15 chicks from each of the three groups were removed to a different site and challenged with a virulent NDV (Texas GB strain). Re-isolation of the challenge virus was not attempted. All chicks in both NDV-vaccinated challenged groups were protected against clinical signs and mortality. These results show that, based on parameters monitored for the respective challenge virus, simultaneous application of live aMPV and NDV vaccines did not affect the efficacy of either vaccine.

This study was conducted to evaluate the immune response against Newcastle disease (ND) virus vaccines (live attenuated and inactivated) in purebred Lohman Selected Leghorn (LSL), Fayoumi, male Fayomi × female LSL (FL crossbred), male LSL × female Fayomi (LF reciprocal crossbred) chickens. One-hundred-day-old chicks of each genetic type were assigned to five equal replicates. The log geometric means of the hemagglutination inhibition (HI) antibody titers were calculated. The FL crossbred chickens had a significantly higher HI antibody titer at day 26 of age when compared with the LSL chickens (P = 0.039). The Fayoumi and FL crossbred chickens had significantly higher HI antibody titers at day 45 of age (2.35 and 2.23, respectively) when compared with the LSL chickens (P = 0.031). In the same way, the purebred Fayoumi and FL crossbred chickens had significantly higher HI antibody titers at 60 and 75days of age (P = 0.009 and 0.041, respectively) when compared with the purebred LSL and LF crossbred chickens. The LSL chickens showed a significantly higher (P < 0.05) correlation estimate between HI titer to the ND vaccine on day 75 of age and body weight at week 12 of age. When challenged with the virulent ND virus, the hazard ratio (HR) for mortality rates in purebred LSL and LF crossbred chickens were significantly (HR = 3.52 and 2.07; P = 0.001 and 0.049, respectively) higher than the Fayoumi chickens. In conclusion, purebred Fayoumi and FL crossbred chickens showed superior antibody titers against live attenuated and inactivated ND virus vaccines. Hence, Fayoumi breed may be incorporated in the crossbreeding programs to improve the genetic resistance to ND.

Newcastle disease (ND), caused by the virulent Newcastle disease virus (NDV), is one of the most important viral diseases of birds globally, but little is currently known regarding enzootic trends of NDV in northeastern China, especially for class I viruses. Thus, we performed a surveillance study for NDV in northeastern China from 2013 to 2015. A total 755 samples from wild and domestic birds in wetlands and live bird markets (LBMs) were collected, and 10 isolates of NDV were identified. Genetic and phylogenetic analyses showed that five isolates from LBMs belong to class I subgenotype 1b, two (one from wild birds and one from LBMs) belong to the vaccine-like class II genotype II, and three (all from wild birds) belong to class II subgenotype Ib. Interestingly, the five class I isolates had epidemiological connections with viruses from southern, eastern, and southeastern China. Our findings, together with recent prevalence trends of class I and virulent class II NDV in China, suggest possible virus transmission between wild and domestic birds and the potential for an NDV epidemic in the future.