Partial Fusion (F) Gene Analysis of Newcastle Disease Virus Detected in Pakistan during 2021-2022

Newcastle disease (ND) virus is a leading threat to commercial and domestic poultry in Pakistan. The virus infects and constitutes irreversible impairment to the nervous system, damages the respiratory system, and marks severe gastrointestinal lesions leading to heavy mortality in short-living birds and substantial losses in layers and breeders. The continuous emergence and evolution of the virus made it inclined to evade the humoral response and indirectly the circumvention of artificial active immunization. Newcastle disease is caused by the orthoavula genus of the paramyxoviridae family and has shown high genetic diversity even in their genotypes while information regarding enzootic trends of the virus is scanty in Pakistan. A total of 40 tracheal samples of NDV were collected from different commercial broiler farms and 11 isolates of NDV were identified. In the current study, we determined the genetic diversity of the Newcastle disease virus based on the partial sequencing of the fusion protein gene available in the NCBI database. Genetic analysis showed that seven isolates belonged to class I genotype VII and four belonged to class II genotype II. Interestingly, two isolates had epidemiological connections with vaccine-like class II genotype II. Our findings, concerning the recent outbreaks of class I genotype VII and class II genotype II of NDV in vaccinated commercial flocks, suggest possible potential partial mutations in the fusion protein gene. Genetic diversity and formation of the new cleavage site in an important neutralizing protein of wild strain are linked with the potency of artificial active immunization and a major cause of vaccine failure.

Global Epidemic of Ebola Virus Disease and the Importation Risk into China: An Assessment Based on the Risk Matrix Method

Objective To analyze the global epidemic status of the Ebola virus disease(EVD) and assess the importation risk into China.Methods Data from World Health Organization reports were used. We described the global epidemic status of EVD from 1976–2021, and assessed and ranked the importation risk of EVD from the diseaseoutbreaking countries into China using the risk matrix and Borda count methods, respectively.Results From 1976–2021, EVD mainly occurred in western and central Africa, with the highest cumulative number of cases(14,124 cases) in Sierra Leone, and the highest cumulative fatality rate(85%) in the Congo. Outbreaks of EVD have occurred in the Democratic Republic of the Congo and Guinea since 2018. The importation risk into China varies across countries with outbreaks of disease.The Democratic Republic of the Congo had an extremely high risk(23 Borda points), followed by Guinea and Liberia. Countries with a moderate importation risk were Nigeria, Uganda, Congo, Sierra Leone,Mali, and Gabon, while countries with a low importation risk included Sudan, Senegal, and Co te d’Ivoire.Conclusion China is under the risk of EVD importation with the globalization and severe epidemic status of EVD. Key attention need to be paid to the Democratic Republic of the Congo, Guinea, and Liberia. Therefore, it is necessary to prevent and prepare in advance for importation risk in China.

Characterisation and separation of infectious bursal disease virus-like particles using aqueous two-phase systems

Infectious bursal disease(IBD)causes considerable economic losses in the commercial poultry industry worldwide.The principal way to control IBD virus(IBDV),the causative agent of IBD,is still through vaccination programs.Virus-like particles(VLPs)are recognised as a safe and potent recombinant vaccine platform.This research work explores the characterisation and separation of infectious bursal disease virus-like particles(IBD-VLPs)from crude feedstock.Various characteristics were studied with highperformance size-exclusion chromatography(HP-SEC),sodium dodecyl sulphate–polyacrylamide gel electrophoresis(SDS-PAGE)and transmission electron microscopy(TEM)analyses.Subsequently,the separation of IBD-VLPs using polyethylene glycol(PEG)/sodium citrate-based aqueous two-phase systems(ATPSs)was conducted and optimised.Moreover,a scale-up study of the best ATPS constituted of 15%PEG 6000,11%sodium citrate and 10%crude feedstock was performed to compare the separation performance of IBD-VLPs with and without centrifugation-assisted.The results indicated that the optimised ATPS with centrifugation-assisted for both 5 g and 50 g systems showed good recovery of IBDVLPs of>97%in the interphase between the PEG-rich top and salt-rich bottom phases.These optimised systems also showed high removal efficiencies of impurities of>95%.The results demonstrated that aqueous two-phase extraction could be a promising technology for efficient VLPs separation.

Study on Control Techniques of Virus Diseases in Facility Cherry

The diseases of sweet cherry caused by viruses and viroids have occurred in the main cherry producing areas of China and are increasing year by year, which has become an important factor affecting the yield and quality of sweet cherry. For this reason, this paper elaborated the prevention and countermeasures of virus diseases from six aspects, including soil treatment, cultivation of disease-free seedlings, prevention of disease and pest damage, reasonable pruning, water management, and treatment of virus diseases, in order to provide technical guidance for the disease resistance, high quality and high yield of facility cherry, as well as information reference for the further improvement of the technical system of virus disease prevention and cultivation of facility cherry.

Hepatitis B virus infection in patients with Wilson disease:A large retrospective study

BACKGROUND Wilson disease(WD)is the most common genetic metabolic liver disease.Some studies have shown that comorbidities may have important effects on WD.Data on hepatitis B virus(HBV)infection in patients with WD are limited.AIM To investigate the prevalence and clinical impact of HBV infection in patients with WD.METHODS The clinical data of patients with WD were analyzed retrospectively,and the data of patients with concurrent WD and HBV infection were compared with those of patients with isolated WD.RESULTS Among a total of 915 WD patients recruited,the total prevalence of current and previous HBV infection was 2.1%[95%confidence interval(CI):1.2%-3.0%]and 9.2%(95%CI:7.3%-11.1%),respectively.The main finding of this study was the identification of 19 patients with concurrent WD and chronic hepatitis B(CHB)infection.The diagnosis of WD was missed in all but two patients with CHB infection.The mean delay in the diagnosis of WD in patients with concurrent WD and CHB infection was 32.5 mo,which was significantly longer than that in patients with isolated WD(10.5 mo).The rates of severe liver disease and mortality in patients with concurrent WD and CHB infection were significantly higher than those in patients with isolated WD(63.1%vs 19.3%,P=0.000 and 36.8%vs 4.1%,P<0.001,respectively).Binary logistic regression analysis revealed a significantly higher risk of severe liver disease at the diagnosis of WD in patients with current HBV infection[odds ratio(OR)=7.748;95%CI:2.890-20.774;P=0.000)]or previous HBV infection(OR=5.525;95%CI:3.159-8.739;P=0.000)than in patients with isolated WD.CONCLUSION The total prevalence of current HBV infection in patients with WD was 2.1%.The diagnosis of WD in CHB patients is usually missed.HBV infection is an independent risk factor for severe liver disease in WD patients.The diagnosis of WD should be ruled out in some patients with CHB infection.

Ebola Virus Disease: General Characteristics, Thoughts, and Perspectives

In December 2013, a new round of Ebola virus disease （EVD） first occurred in a remote countryside of Guinea and then spread in Guinea, Liberia, Sierra Leone, and Nigeria of West Africa. EVD, caused by Ebolavirus and previously known as Ebola hemorrhagic fever, is an acute infectious disease with fatality rates up to 90%. As of August 22, 2014, the number of suspected and confirmed cases was 2615, causing 1427 deaths[I]. On August 8, 2014, World Health Organization announced the current outbreak in West Africa as an international public health emergency. The global epidemic tendency remains ambiguous to date. In recent years, China closely collaborates with West Africa in labor, business, overseas education, and also sends aid medical team there. Thus, the risk of importing the disease cannot be ignored. We conduct this literature review of epidemiology, pathogen, prophylaxis, and treatment to provide evidence for controlling the risk and carrying out effective interventions.

Effect of Astragalus polysaccharides on Erythrocyte Immune Adherence of Chickens Inoculated with Infectious Bursal Disease Virus

Two hundred and forty specific pathogen free leghorn chickens were randomly divided into four groups and reared in isolated pens. The tested chickens were negative to infectious bursal disease virus （IBDV） at 25 d old. Group 1 was treated with saline, whereas Groups 2, 3, and 4 were inoculated with 0.3 mL IBDV suspension intranasally the next day. Groups 3 and 4 were also administered with Astragalus polysaccharides （APS） intramuscularly twice daily at 5 or 10 mg kg-1 BW, respectively, until 31 d old. The erythrocyte-C3b receptor rosette rate （E-C3bRR） and the erythrocyte-C3b immune complex rosette rate （E-ICRR） were measured at 25, 29, 32, 35, and 38 d old. The results showed that IBDV significantly reduced E-C3bRR and E-ICRR when compared with the control group （P 〈 0.05）, while simultaneous administration of APS with 1BDV maintained E-C3bRR at similar levels to the control group （P 〉 0.05） and increased E-ICRR when compared with the control group and the group non-treated with APS （P 〈 0.05）. APS treatment reduced the morbidity and mortality of chickens inoculated with IBDV （P 〈 0.05）. The results suggest that APS may enhance the immune adherence of chickens erythrocytes by affecting the activity and/or the number of complement receptors on the erythrocyte membrane. These findings can be beneficial in providing an understanding of the basic mechanisms required for the rational application of APS in modern medicine.

An improved scheme for infectious bursal disease virus genotype classification based on both genome-segments A and B

Infectious bursal disease(IBD)is caused by infectious bursal disease virus(IBDV),which has a genome consisting of two segments of double-stranded linear RNA.IBDVs have been traditionally divided into four phenotypes based on their pathogenicity and antigenicity,including classic,variant,very virulent,and attenuated IBDV.With the emergences of divergent molecular characteristics of novel strains produced by continuous mutations and recombination,it is increasingly difficult to define new IBDV strains using the traditional descriptive classification method.The most common classification scheme for IBDV with segmented genome is based solely on segment A,while the significance of segment B has been largely neglected.In this study,an improved scheme for IBDV genotype classification based on the molecular characteristics of both VP2(a viral capsid protein encoded by segment A)and VP1(an RNA-dependent RNA polymerase protein encoded by segment B)was proposed for the first time.In this scheme,IBDV was classified into nine genogroups of A and five genogroups of B,respectively;the genogroup A2 was further divided into four lineages.The commonly used phenotypic classifications of classic,variant,very virulent,and attenuated IBDVs correspond to the A1 B1,A2 B1,A3 B2,and A8 B1 genotypes of the proposed classification scheme.The novel variant IBDVs including the strains identified in this study were classified as belonging to genotype A2 d B1.The flexibility and versatility of this improved classification scheme will allow the unambiguous identification of existing and emerging IBDV strains,which will greatly facilitate molecular epidemiology studies of IBDV.

Newcastle disease virus-based MERS-CoV candidate vaccine elicits high-level and lasting neutralizing antibodies in Bactrian camels

Middle East respiratory syndrome coronavirus （MERS-CoV）, a member of the Coronavifidae family, is the causative pathogen for MERS that is characterized by high fever, pneumonia, acute respiratory distress syndrome （ARDS）, as well as extrapul- monary manifestations. Currently, there are no approved treatment regimens or vaccines for MERS. Here~ we generated recombinant nonvirulent Newcastle disease virus （NDV） LaSota strain expressing MERS-CoV S protein （designated as rLa- MERS-S）, and evaluated its immunogenicity in mice and Bactrian camels. The results revealed that rLa-MERS-S showed similar growth properties to those of LaSota in embryonated chicken eggs, while animal immunization studies showed that rLa-MERS-S induced MERS-CoV neutralizing antibodies in mice and camels. Our findings suggest that recombinant rLa- MERS-S may be a potential MERS-CoV veterinary vaccine candidate for camels and other animals affected by MERS.

Activity of T Cells Stimulated by Hemagglutinin-neuraminidase of Newcastle Disease Virus in vivo

To investigate the stimulated activity of T cells and the anti-tumor properties of hemagglutinin-neuraminidase（HN） of Newcastle disease virus（NDV） strain Changchun（NDVcc）, the expression of HN gene in hepatoma cells（human HepG-2 and mouse H22 cells） infected with the recombinant adenovirus（Ad-HN） was identified by Western blot analysis and flow cytometry. Sialidase activity of NDVcc HN expressed by Ad-HN was assayed by the periodate-resorcinol method. The in vivo anti-tumor effects of NDVcc HN were evaluated in the H22 solid tumor model. Regional lymph nodes of the mouse model treated with Ad-HN were removed to harvest T lymphocytes and evaluating the specific cytotoxicity of cytotoxic T lymphocyte（CTL） and natural killer（NK） cells by an L-lactate dehydrogenase（LDH） assay, in the mean time, the secretion of cytokines was analyzed by enzyme linked immunosorbent assays（ELISA）. The results show that NDVcc HN was effectively expressed by Ad-HN in HepG-2 and H22 cells. The sialidase activity assay showed that Ad-HN significantly reduced sialic acid level of the hepatoma cells compared with the cells infected the empty adenovirus vector（Ad-mock）. When treated with Ad-HN, the growth of subcutaneous H22 primary tumors in C57BL/6 mice was suppressed, and the mean mice survival increased. In addition, the treatment of Ad-HN elicited strong NK and CTL responses, and high levels of Th1 cytokines, such as IL-2 and IFN-γ. In conclusion, NDVcc HN effectively elicits T cell-mediate anti-tumor cytotoxicity via sialidase activity and may be a novel strategy for cancer immunotherapy.

Ebola virus disease: From epidemiology to prophylaxis

The outbreak of Ebola virus disease(EVD) continues to spread through West Africa. Since the first reported EVD in March 2014, the number of cases has increased rapidly, with the fatality rate of >50%. The most prevalent Ebola virus belongs to the species of Zaire ebolavirus, with a mortality rate as high as 90%. Although there were introduced cases in other continents, Africa is the endemic area where fruit bats and apes are suspected to be Ebola virus carriers. The virus might be transmitted from the host animals to humans if humans consume relative raw and contaminated meats; however, human-to-human transmission via close contact is the major route of current outbreaks. EVD happens at any seasons and affected people of any race in any age groups. Direct contact with body fluids of EVD patients and living in the contaminated environment greatly increase the risk of being infected. Transmission viaaerosol is less possible but the transmission via droplet is possible in humans. Thus, health care providers are facing danger of getting Ebola virus infection. So far, there are limited vaccines, drugs and/or therapies to prevent Ebola virus infection or treat EVD. Medical workers should follow the current standard prophylactic procedures. Military forces can orchestrate efficient care to mass EVD casualties. Although it is necessary to speed up the pace of developing effective vaccine and therapeutics for the prevention and treatment of EVD, public health prophylaxis is the most important issue at present to control the spread of this disease cost-effectively.

The Complete Genomic Sequence of a Foot-and-Mouth Disease Virus Isolated from the Swine

The complete genomic sequence of foot-and-mouth disease virus (FMDV) Chinese strain OH/CHA/99 was determined. The 8040 nt sequence and the deduced amino acid sequence werecompared with FMDV sequences published. The results showed that OH/CHA/99 shared highersequence homology with OTYTW/97, indicating their close genetic relationship. However,the strain had lower sequence identity with O1/Kaufbeuren/66 strain. Besides, largedeletions in 3A coding region were observed in OH/CHA/99. It was shown that the poly (A)tail of OH/CHA/99 had 56 As at least.

Complete Nucleotide Sequence of a Newly Avirulent Newcastle Disease Virus Hubei 92(HB92) Strain

A new avirulent, heat-resistance HB92 strain of newcastle Disease Virus (NDV) was acquired from Australia V4 strain. Its complete nucleotides sequence was first determined. The entire genome of NDV HB92 consists of 15 186 nucleotides (GenBank accession no. AY225110). It was demonstrated by sequence analysis that nucleotides homology of HB92 strain with virulent strain ZJ1 were 83.9%, and the homology compared with avirulent vaccine strain La Sota and B1 were 94.0% and 93.5%, respectively. These results might be contributive to the study of the molecular mechanism of evolution of the NDV strain HB92 and to develop the engineered recombinant vaccine. Key words newcastle disease virus - genomic sequence - sequence analysis CLC number S 852. 65 Foundation item: Supported by Hubei Natural Science Foundation (2002AB144)Biography: Pan Zhi-shu(1961-), male, Ph. D, Associate professor, research direction: molecular biology and pathogenesis of eucaryotic viruses.

Antiviral Effects of Compound Traditional Chinese Medicine on Newcastle Disease Virus

[Objective] To investigate the mechanism of compound traditional Chinese medicine （TCM） on Newcastle disease virus （NDV） and to provide a scientific basis for the reasonable usage of antiviral drugs in clinic. [Method] The compound TCM was composed of Hedyotis diffusa, Lonicera japonica Thunb, Radix astragali and Glycyrrhiza uralensis. Different dilutions of fluid extract were prepared. Its antiviral effects on NDV were observed through three inoculation ways, first, inoculation with the medicine and NDV mixture which had been incubated at 37 ℃; second, incubating chicken embryo fibroblasts （CEF） with the medicine followed by inoculation with NDV; third, inoculation with N DV followed by incubating CEF with the medicine. The A,= was determined by M]-r [ 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide） ~ method. Therapeutic indexes were used to evaluate the antiviral effects. [ Result] The minimum effective concentration of the compound TCM which acted through the three ways was 1.0 × 2^-10 1.0 × 2^-8 and 1.0 × 2^-7 g/ml, respectively. The antiviral effects of the compound TCM were the best through inoculation with the incubated medicine and NDV mixture, followed by the second method and the third method. [ Conclusion] The compound TCM can not only kill NDV directly in vitro but also inhibit viral propagation.

Development of a Liquid Chip Technique to Simultaneously Detect Taura Syndrome Virus( TSV) and Yellow Head Disease Virus( YHDV)

The aim is to develop a liquid chip technique to detect Taura syndrome virus( TSV) and yellow head disease virus( YHDV) on Penaeus orientalis simultaneously. The CP2 gene of TSV and N gene of YHDV in Gen Bank was analysed by using the software DNAStar 7. 0 to design the TSV-and YHDV-specific primers. The primers were labeled with biotin and subjected to amination modification. They were then coupled with fluorescence-coded microspheres and then used for hybridization with RT- PCR products of TSV and YHDV. The liquid chip detection technique for detection of TSV and YHDV was established by using BD FACSArray to detect fluorescence signal in the reaction system. This assay system had a high sensitivity to TSV and YHDV,with the detection of limit of 100 pg. Moreover,the assay was specific for the detection of TSV,YHDV and was not susceptible to cross with other viruses,including white spot syndrome virus( WSSV),spring viremia of carp virus( SVCV),infectious haematopoietic necrosis virus( IHNV). In conclusion,the liquid chip assay technique established in this study is highly sensitive and specific to TSV and YHDV detection. Moreover,it provides a novel,convenient and rapid approach for the detection of TSV and YHDV.

Advances in Research of Garlic Virus Diseases

Garlic virus infection is an important disease which affects garlic production,with the increasing years of planting,harm of virus is serious year by year,which seriously affect yield and quality of garlic.In order to know the garlic virus effectively,the paper reviewed the research situation of several important garlic virus in virus species,origin,distribution,host range,symptom,route of transmission,classification,genome and detection technique and the prevention technology of garlic viruses.At the same ...

The circulation of unique reassortment strains of infectious bursal disease virus in Pakistan

Infectious bursal disease(IBD),caused by IBD virus(IBDV),is one of the most devastating and immunosuppressive diseases of the poultry and has been a constraint on the sustainable poultry production around the globe including Pakistan.While the disease is threatening the poulty industry,the nature of predominant strains of IBDV in Pakistan remained l-defined.In this study,an epidemiology survey was conducted in the main chicken-farming regions of Pakistan.The batch of Pakistan IBDVs genes simultaneously covering both VP1 and VP2 were amplified,sequenced,and analyzed.The unique segment-reassortant IBDVs(vv-A/Uniq-B),carrying segmentA from vvIBDV and segment B from one unique ancestor,were identifed as one important type of circulating strains in Pakistan.The data also discovered the characteristic molecular features of Pakistan IBDVs,which will contribute to scientific vaccine selection and effective prevention of the disease.

Dynamic Pathology and Antigen Location Study on Broiler Breeders with Coinfection of Marek's Disease Virus and Reticuloendotheliosis Virus

To further understand the generation and development of coinfection of Marek＇s disease virus （MDV） and reticuloendotheliosis virus （REV） in broiler breeders, and then find the method and optimal time of differential diagnosis for complex clinic multiple infection, the authors studied the pathohistological changes, apoptosis, immunohistochemistry （immunofluorescence）, and ultrastructure of tumor tissues of broiler breeders inoculated with MDV and REV. The study showed that proliferation of small lymphocytes was seen in the main organs at the age of 1 week, then immature lymphocytes, all kinds of lymphocytes, primitive reticulum cells, and Marek＇s disease cells （MDCs） were observed at 2-9 weeks. Apoptosis of lymphocytes could not be seen until the age of 10 weeks in the immune system. Immunohistochemistry detection showed that the positive signs of MDV and REV antigen were observed in the main organs at 2 weeks of age. Multi-morphology lymphocytes, MDV, and REV, mitotic figures and apoptosis of lymphocytes were observed with the help of transmission electron microscopy. MDV cooperating with REV promotes the course of disease of coinfection. Differential diagnosis can be done by immunohistochemistry in the early stage （before 2 weeks）, and histopathology in the late stage （post 4 weeks）. MDCs, primitive reticulum cells, immature lymphocytes, and two kinds of virions can serve as a basis for bistopathology differential diagnosis.

Development of a Synthetic Peptide ELISA Assay for the Detection of Foot-and-Mouth Disease Virus Nonstructural Protein Antibodies

In order to develop an ELISA assay with synthetic peptides for the detection of antibody to the nonstructural proteins of foot-and-mouth disease virus, specific peptides were synthesized by a solid-phase method according to FMDV NSPs B-cell epitopes, and were conjugated to carrier protein BSA. An ELISA system was developed to detect FMDV NSPs antibody with the conjugated proteins as the coating antigen. The optimal coating concentration of the antigen was determined as 2.5 μg mL-1. The comparative study of this assay with UBI NSP ELISA kit and national commercial 3ABC ELISA kit in the detection of 199 serum samples showed that they were very coincident, and the identity rates were 96.48 and 97.48%, respectively. The development of ELISA using the synthetic peptides as coating antigen is specific, reproducible, stable, and easy, and can be used to differentiate FMDV infected pigs from immunized pigs.

Physicochemical Properties of Musk Deer Pneumonia and Purulent Disease Viruses

[ Objective] To understand the physicochemical properties of musk deer pneumonia and purulent disease viruses. [ Method] The pneu- monia and purulent disease viruses were isolated from the abnormal and purulent lung tissues of musk deer. Then the isolated viruses were inocula- ted into the Vero cells. After culturing, the virus solution was collected and used to determine TCID50 and genoma types. The sensitivity to fat sol- vent, resistance to hydrochloric acid and trypsin as well as tolerance to heat of the musk deer pneumonia and purulent disease viruses were detec- ted, respectively. [ Result] The obvious cytopathic effects （CPE） were found in Veto cells infected by the isolated viruses. The virus was 2-1.43 TCID50/ml and its genome was RNA. The virus was not sensitive to chloroform, 1% trypsin and heats, and it had a certain tolerance to 0.1 mol/L hy- drochloric acid. [ Conclusion] The study on the physicochemical properties of musk deer pneumonia and purulent disease viruses lays a foundation for prevention and control of the musk deer pneumonia and purulent diseases.