Expression and identification of recombinant adenovirus type 5-Norovirus GⅡ.4-VP1 virus-like particles / 中国生物制品学杂志

@#Objective To express and identify recombinant adenovirus type 5-Norovirus(NoV)GⅡ.4-VP1 virus-like particles(VLPs)in 293T cells. Methods The recombinant adenovirus plasmids pAd5-eGFP and pAd5-NoV-GⅡ.4-VP1 were transfected into 293T cells respectively,and the recombinant adenovirus rAd5-eGFP and rAd5-NoV-GⅡ.4-VP1 were rescued. The rAd5-eGFP was subcultured in 293T cells to verify the function of the vector. The rAd5-NoV-GⅡ.4-VP1 was subcultured in293T cells,expressed and purified,and then NoV-GⅡ.4-VLP was formed by self-assembly,which was detected by Western blot,ELISA and observed by transmission electron microscope. Results The green fluorescence of the recombinant adenovirus rAd5-eGFP of various generations was observed under microscope,and the brightness increased with the increase of generations. NoV-GⅡ.4-VP1 protein was expressed in the harvested solution of recombinant adenovirus rAd5-NoV-GⅡ.4-VP1 of various generations,with a relative molecular mass of about 58 900. NoV-GⅡ.4-VLP showed specific binding to the rabbit anti-NoV-GⅡ.4-VP1 serum;it had similar conformation to natural NoV virus particles and can effectively identify NoV receptors in volunteer saliva samples;microscopic observation showed that the morphology was complete and spherical,with a diameter of 43. 5 — 58. 3 nm,while there were a few protrusions on the surface of the particles,which might be the P domain exposedon the particle surface during self-assembly of VP1. Conclusion The expressed recombinant adenovirus NoV-GⅡ. 4-VLP has complete VLP structure and good specificity,and is expected to be used in the related research of NoV adenovirus vector vaccine.

Epidemiological characteristics and influencing factors of adenovirus-associated acute conjunctivitis in Shenzhen，2014‒2022 / 上海预防医学

ObjectiveTo investigate the epidemic characteristics and influencing factors of acute conjunctivitis caused by human adenovirus（HAdv）in Yantian District of Shenzhen City， Guangdong Province from 2014 to 2022， and to provide evidence for formulating effective prevention and control measures. MethodsDescriptive epidemiology was used to analyze the epidemiological characteristics of acute hemorrhagic conjunctivitis （AHC） cases reported from the Chinese disease prevention and control information system. Etiological characteristics of laboratory-confirmed cases were analyzed， and a case-control study method of test-negative design （TND） was conducted as well. According to the result of HAdv detection， analysis was used to identify the influencing factors of morbidity. ResultsA total of 1 005 AHC cases were reported in Yantian District， Shenzhen City from 2014 to 2022， with an average annual incidence rate of 49.49/105. The incidence rate ranged from 4.67/105 to 117.28/105. The peak incidence occurred from July to October each year， with a male-to-female ratio of 1.42∶1. The median （P25，P75） age of onset was 29（12，40） years. A total of 716 eye swabs were collected for etiological detection from 2014 to 2022. HAdv positive detection rate was 36.45% （263/716）， Cox 24v positive detection rate was 0.28% （2/716）， while EV70 was not detected. The longest viral shedding time in eye swabs was 10 days， with a median of 2 days. The highest HAdv positive detection rate （47.47%） was observed when the sampling-to-onset interval was 4‒5 days， and the difference was statistically significant （P<0.05）， with a trend of first increasing and then decreasing. Multivariate logistic regressing analysis showed that aged 18‒ years， and average temperature higher than 24.90 ℃ 3 days before onset （P<0.05） were the risk factors for acute HAdv conjunctivitis. ConclusionAHC in Yantian District， Shenzhen City showed a low level of prevalence from 2014 to 2022， with HAdv being the predominant pathogen. The peak period of viral shedding occurred on the 4th to 5th day after symptom onset， suggesting the importance of adherence to strict home isolation for infected persons. Aged18‒ years and average temperature increased 3 days before onset are associated with an increased risk of adenoviral conjunctivitis. It is recommended to strengthen personal protection and keep doing a good job of AHC surveillance and early warning， so as to timely prevent the outbreaks of AHC.

Construction of recombinant adenovirus expression vector of avian reovirus σC protein and its effect on proliferation of hepatocellular carcinoma cells / 中国生物制品学杂志

@#Objective To construct the recombinant adenovirus vector pAd-σC for the expression of avian reovirus(ARV)aC protein and to detect its effects on the proliferation of hepatocellular carcinoma cells,in order to build up a basis for the development of novel anti-tumor vaccines.Methods The recombinant shuttle vector pShuttle-σC was constructed by PCR amplification of ARV σC gene,and then transformed into competent BJ5183 cells containing the adenovirus vector pAdessy-1.The recombi-nant adenovirus vector pAd-σC was obtained by homologous recombination,and the virus was packaged in HEK293 cells.The virus titer was measured by TCID_(50),the expression of σC protein was determined by Western blot and ELISA,and the effect of virus on the proliferation of human hepatocellular carcinoma cells SMMC7721 was detected by CCK-8 assay.Results The recombinant shuttle vector pShuttle-σC was confirmed to be constructed correctly by double enzyme digestion and sequen-cing,and the recombinant adenovirus vector pAd-σC was constructed correctly as identified by colony PCR.σC protein was successfully expressed in hepatocellular carcinoma cells SMMC7721.The recombinant adenovirus Ad-σC had a titer of 10~(7.5)/0.1 mL,which inhibited the proliferation of hepatocellular carcinoma cells SMMC7721.Conclusion The recombinant adenovirus vector pAd-trC containing ARV σC gene was successfully constructed,and its inhibitory effect on tumor cell proliferation was preliminarily analyzed,which lays a foundation for revealing the molecular mechanism of ARV oncolytic effect and further developing novel anti-tumor biological preparation.

Bronquiolitis obliterante postinfecciosa en niños: serie de casos en un hospital pediátrico de Perú / Postinfectious bronchiolitis obliterans in children: case series at a pediatric hospital in Peru

Resumen Introducción: La bronquiolitis obliterante postinfecciosa es una enfermedad pulmonar poco frecuente; existen limitados reportes en Sudamérica. Caso clínico: En esta serie se reportan 10 pacientes con esta enfermedad diagnosticados en el Instituto Nacional de Salud del Niño-Breña (Lima-Perú). La mediana de edad al diagnóstico fue de 19 meses. Todos los pacientes presentaron el antecedente de infección respiratoria aguda grave. Los síntomas más frecuentes fueron tos, dificultad respiratoria, sibilancias e hipoxemia; el patrón de atenuación en mosaico fue la característica más frecuente en la tomografía. Todos tenían serología positiva para adenovirus. Se administró tratamiento con pulsos de metilprednisolona, azitromicina, hidroxicloroquina y corticoides inhalados. Ningún paciente falleció durante el seguimiento. Conclusiones: En los niños previamente sanos con antecedente de infección respiratoria aguda grave y sintomatología obstructivo bronquial persistente se debe considerar el diagnóstico de bronquiolitis obliterante postinfecciosa. Este es el primer reporte en Perú con un régimen terapéutico adaptado a nuestra institución.

Abstract Background: Postinfectious bronchiolitis obliterans is a rare lung disease; there are limited reports in South America. Case report: We report 10 patients with this disease diagnosed at the Instituto Nacional de Salud del Niño-Breña (Lima-Peru). The median age at diagnosis was 19 months and all patients had a history of severe acute respiratory infection. The most frequent symptoms were cough, respiratory distress, wheezing, and hypoxemia. The mosaic attenuation pattern was the most frequent on the tomography. All the patients had positive serology for adenovirus. The treatment received was methylprednisolone pulses, azithromycin, hydroxychloroquine, and inhaled corticosteroids. No patient died during the follow-up. Conclusions: In previously healthy children with a history of severe acute respiratory infection and persistent bronchial obstructive symptoms, the diagnosis of postinfectious bronchiolitis obliterans should be considered. This is the first report in Peru with a therapeutic regimen adapted to our institution.

Hepatitis aguda de origen desconocido. Una serie de casos probables atendidos en un hospital pediátrico de referencia en Panamá

Introducción: Durante el año 2022 se objetivó un aumento de casos de hepatitis de origen desconocido en la población pediátrica. Su etiología a día de hoy todavía no ha sido esclarecida. Este estudio tiene como objetivo presentar la clínica y los resultados de los análisis de laboratorio de los pacientes con hepatitis atendidos en el Hospital del Niño Dr. José Renán Esquivel durante el año 2022.   Material y métodos: Estudio descriptivo de 10 pacientes pediátricos con criterios de hepatitis aguda de origen desconocido atendidos en nuestro centro durante el año 2022. Se ha llevado a cabo un análisis de las principales variables clínicas, analíticas, demográficas, vacunales y serológicas.   Resultados:  Se encontraron 10 pacientes, con una media de edad de 2,8 años. La clínica predominante ha sido la de ictericia, fiebre y clínica gastrointestinal. Analíticamente se ha detectado un aumento de los niveles de transaminasas así como de bilirrubina. Tan sólo un paciente contaba con historia de vacunación para el SARS-CoV-2. El principal agente etiológico identificado ha sido el adenovirus, encontrándose en el 40% de los casos.   Conclusiones: Los pediatras deben tener en cuenta el diagnóstico de hepatitis aguda de origen desconocido en pacientes con clínica de ictericia y fiebre, pudiendo asociar vómitos, diarrea, hepatomegalia, dolor abdominal y coluria, entre otros. La elevación de transaminasas (>500 U/L de AST o ALT) y bilirrubina debe aumentar el índice de sospecha. A pesar de que el principal agente etiológico parece guardar relación con el adenovirus, son necesarios más estudios para confirmar dicha hipótesis. (provisto por Infomedic International)

Introduction: During the year 2022 an increase in cases of hepatitis of unknown origin was observed in the pediatric population. Its etiology has not yet been clarified. This study aims to present the clinical and laboratory results of patients with hepatitis treated at the Hospital del Niño Dr. José Renán Esquivel during the year 2022.   Material and methods: Descriptive study of 10 pediatric patients with criteria of acute hepatitis of unknown origin seen in our center during the year 2022. An analysis of the main clinical, analytical, demographic, vaccination and serological variables was carried out.   Results: The final sample consisted of 10 patients, with a mean age of 2.8 years. The predominant clinical manifestations were jaundice, fever and gastrointestinal symptoms. Laboratory tests showed increased levels of transaminases and bilirubin. Only one patient had a history of vaccination for SARS-CoV-2. The main etiological agent identified was adenovirus, found in 40% of the patients studied.   Conclusions: Pediatricians should consider the diagnosis of acute hepatitis of unknown origin in patients with jaundice and fever, which may be associated with vomiting, diarrhea, hepatomegaly, abdominal pain and choluria, among others. Elevated transaminases (>500 U/L of AST or ALT) and bilirubin should increase the index of suspicion. Although the main etiological agent to date seems to be related to adenovirus, further studies are needed to confirm this hypothesis. (provided by Infomedic International)

Sobre la hepatitis aguda grave de etiología desconocida: la situación en México / About severe acute hepatitis of unknown etiology: the situation in Mexico

Introducción: en México, las hepatitis virales son de notificación epidemiológica obligatoria, pero no existe un sistema especial de vigilancia. La información disponible se limita a la distribución por edad y sexo. Ante la alerta de casos de hepatitis aguda grave de etiología desconocida, en la Unión Europea el Consejo Nacional de Vigilancia Epidemiológica (CONAVE) alertó al Sistema Nacional de Salud (SNS) para la atención y vigilancia de estos casos. Desarrollo: la hipótesis más convincente sobre la etiología está relacionada con una respuesta inmunitaria exacerbada que es mediada por superantígenos relacionados con la proteína espiga del SARS-CoV-2, activados por una infección por adenovirus que desencadena una respuesta de linfocitos T que provoca apoptosis de hepatocitos. Con base en la presentación clínica (niños menores de 16 años, con diarrea, dolor abdominal, ictericia, vómito e hipertransaminasemia) se han diseñado definiciones operacionales para su identificación y notificación al Sistema Nacional de Vigilancia Epidemiológica (SINAVE). Hasta junio del 2022, se han identificado 56 casos en México. Conclusiones: este brote de hepatitis representa un reto para el SINAVE. Es necesario incluir la identificación de adenovirus en el algoritmo diagnóstico de enfermedad respiratoria viral, implementar un sistema especial de vigilancia epidemiológica de hepatitis virales y sensibilizar a los profesionales sanitarios en el tema.

Introduction: In Mexico viral hepatitis requires mandatory epidemiological notification, but there is no special surveillance system. Available information is limited to distribution of cases by age and sex. Given the alert of cases of severe acute hepatitis of unknown etiology in the European Union, the National Council for Epidemiological Surveillance (Consejo Nacional de Vigilancia Epidemiológica) alerted the entire National Health System to care for and monitor these cases in Mexico. Development: The most convincing hypothesis is an exacerbated immune response mediated by superantigens related to the spike protein of SARS-CoV-2, activated by adenovirus infection that ends in a response of T lymphocytes that causes apoptosis of hepatocytes. Based on clinical presentation (children under 16 years of age, with diarrhoea, abdominal pain, jaundice, vomiting and increase in transaminases) the operational case definitions have been designed for their timely identification and notification to the National System of Epidemiological Surveillance (Sistema Nacional de Vigilancia Epidemiológica). Until June 2022, 56 cases have been identified in Mexico. Conclusions: This hepatitis outbreak represents a challenge for the National System of Epidemiological Surveillance. It is necessary to include the identification of adenovirus in the diagnostic algorithm for viral respiratory disease, to implement a special epidemiological surveillance system for viral hepatitis, and to sensitize health professionals on this subject.

Severe Acute Hepatitis: An Emerging Grave Illness in Children

Acute hepatitis of unknown origin in children has been recently described in the literature, and a case definition has also been proposed for this condition. The exact etiology is unknown and exclusion of infectious, metabolic, autoimmune and toxin mediated injuries is essential. Management for this condition is supportive, but some may require liver transplantation. Infection prevention and control practices are important as the etiology remains unidentified.

Surveillance results of viral diarrhea in children under five years of age in Shaoxing City / 预防医学

Objective@#To investigate the epidemiological and etiological characteristics of viral diarrhea among children under 5 years of age in Shaoxing City, Zhejiang Province, so as to provide insights into management of viral diarrhea.@*Methods@#The surveillance data on viral diarrhea among children under 5 years of age in Shaoxing City from 2019 to 2022 were collected, including demographics and stool testing results. The epidemiological and etiological characteristics of viral diarrhea were analyzed using a descriptive epidemiological method. @*Results@#A total of 763 diarrheal children under 5 years of age were reported in Shaoxing City from 2019 to 2022, and 236 children were tested positive for virus (30.93%). The detection of virus was 49.01%, 31.61%, 20.43% and 21.89% from 2019 to 2022, which appeared an overall tendency towards a decline (P<0.05). The incidence of viral diarrhea peaked from November to March of the next year and from May to July, and the detection of virus was lower among children with diarrhea living in urban areas (Yuecheng District, Keqiao District and Shangyu District) than among those in suburb areas (Zhuji City, Shengzhou City and Xinchang County) (30.22% vs. 52.00%, P<0.05). There were 206 children tested positive for a single virus, and the detection rates of rotavirus (RV), enteric adenovirus (EAdV), norovirus (NoV), and sapovirus (SaV) were 9.57%, 8.91%, 8.39%, and 0.13%, respectively. There were 25 children with virus co-infections, and the positive rates of EAdV and NoV, RV and EAdV and RV and NoV co-infections were 1.31%, 1.18% and 0.79%, respectively. There were 5 children with triple infections of RV, EAdV and NoV (0.66%). The highest detection of EAdV was seen in April, the highest detection of RV and NoV was seen in January, while SaV was only detected in April. @*Conclusion@#The incidence of viral diarrhea among children under 5 years of age peaked in winter and spring in Shaoxing City from 2019 to 2022, and the cases predominantly occurred in urban areas. The detection of virus appeared an overall tendency towards a decline, with high detection of RV, EAdV and NoV.

Construction and identification of recombinant adenovirus expressing S protein receptor binding domain and N protein of SARS-CoV-2 Delta variant / 中国生物制品学杂志

@#ObjectiveTo construct and identify a recombinant adenovirus expressing S protein receptor binding domain(RBD)and N protein of severe acute respiratory symptom coronavirus 2(SARS-CoV-2)Delta variant.MethodsThe RBD and N gene fragments of SARS-CoV-2 were cloned into pcDNA3.0BA vector respectively to construct recombinant plasmid pcDNA3.0BA-RBD-N. The RBD-CMV-N fragment was amplified by PCR and inserted into shuttle vector pShuttle-CMV. The shuttle plasmid pShuttle-RBD-N was then homologously recombined with pAdeasy-1 to obtain recombinant plasmid pAdeasy-1-RBD-N,which was transfected into HEK293 cells for recombinant adenovirus Ad-RBD-N packaging. The transcription of RBD and N genes of recombinant adenovirus in HEK293 cells was detected by RT-PCR,while the expre-ssion of RBD and N proteins by Western blot and immunofluorescence assay. 12 female BALB/c mice were immunized with Ad-RBD-N by intramuscular injection at a dose of 5 × 109copies per mouse. Blood samples were collected 14 d after immunization,and the serum antibody titers were measured by ELISA.ResultsThe RBD and N genes of recombinant adenovirus were transcribed normally in HEK293 cells,and the RBD and N proteins were expressed normally in MA104 cells. Mice immunized with the recombinant adenovirus produced specific IgG antibodies against RBD and N proteins.ConclusionThe recombinant adenovirus expressing S protein RBD and N protein of SARS-CoV-2 Delta variant was succe-ssfully constructed,which laid a foundation of the follow-up research on Delta variant vaccines.

Research progress regarding status of human adenovirus infections and serotyping technology / 中华检验医学杂志

There are many serotypes of human adenovirus (HAdV), and different serotypes can cause infections in different systems of the human body, one example being community acquired pneumonia (CAP). However, the clinical symptoms of HAdV infections are similar to those caused by other pathogens. To detect and serotype adenovirus rapidly and accurately is crucial towards the clinical diagnosis, treatment and prevention of the virus. This can facilitates the control of nosocomial infection and epidemiological monitoring. This article briefly reviews the molecular characteristics, epidemiology, and typing of human adenovirus, aiming to help formulate effective treatments towards HAdV infection in a clinical setting.

Research progress of m 6A methylation modification in respiratory viruses replication and host immune response / 中华临床感染病杂志

N 6-methyladenosine (m 6A) modification is the most prevalent internal modification of eukaryotic mRNA and is dynamically regulated by a variety of m 6A modifying enzymes, including methylation transferases, demethylases and specific binding proteins. Respiratory viral infections have received much attention in recent years, and the process of virus replication and metabolism in host cells is regulated by m 6A. This article reviews the mechanism of m 6A-regulated enzymes, the roles of m 6A modifications in respiratory viruses replication and the host immune response to viruses, including adenovirus, influenza A virus, severe acute respiratory syndrome coronavirus 2, respiratory syncytial virus, and human metapneumovirus. It would provide a reference for exploring the regulatory role of viral episodic transcriptome modifications and antiviral targets or vaccine development.

Analysis of clinical characteristics and risk factors of severe adenovirus pneumonia in children / 中国小儿急救医学

Objective:To understand the epidemiology, clinical characteristics and associated risk factors of severe adenovirus(ADV)pneumonia in children, providing the basis for targeted prevention and treatment.Methods:Clinical features of children with ADV pneumonia at Children′s Hospital of Soochow University from January 2011 to December 2020 were retrospectively analyzed.According to the severity of the disease, cases were divided into severe ADV pneumonia group and common ADV pneumonia group.The epidemiological and clinical characteristics of two groups were compared, and risk factors for the occurrence of severe ADV pneumonia were analyzed.Results:A total of 1 158 patients with ADV pneumonia were enrolled, including severe ADV pneumonia 104 cases(8.98%) and ordinary ADV pneumonia 1 054 cases(91.02%).The median age of severe ADV pneumonia group was 1.17 (0.83, 2.73) years, which was significantly younger than that of common ADV pneumonia group 3.16 (1.50, 4.50) years( P<0.05), and 77.89% (81/104) of them were younger than 3 years old.The occurrence of severe ADV pneumonia was predominant in winter and spring, accounting for 71.15% (74/104).Cough was present in 89.42% (93/104) and fever in 99.01% (103/104) of the severe ADV pneumonia group.Compared with the common ADV pneumonia group, the severe ADV pneumonia group had a significantly longer febrile time[10(6, 14)d vs. 5(4, 7)d, P<0.05], significantly higher incidence of shortness of breath, wheezing, convulsions/coma[100% vs. 2.09%, 45.19% vs. 13.57%, 10.57% vs. 1.99%, P<0.05], and significantly higher incidences of emphysema, pleural effusion, bronchial signs, pulmonary solids, and atelectasis [21.15% vs. 2.09%, 5.77% vs. 0.19%, 4.81% vs. 0, 3.85% vs. 0.09%, P<0.05].Multivariable Logistic regression showed that age younger than 1.71 years old, wheezing, and the presence of underlying diseases (moderate to severe anaemia, congenital heart disease, neurological disease) were risk factors for the development of severe ADV pneumonia ( P<0.05).Receiver operating characteristic curve analysis showed that the sensitivity and specificity of age<1.71 years old(20 months old) for predicting the occurrence of severe ADV pneumonia were 65.4% and 71.5%, respectively. Conclusion:The age of most severe ADV pneumonia is less 3 years in Suzhou.It usually occurres in winter and spring, with fever, cough, shortness of breath, and wheezing as the main symptoms.Pulmonary manifestations such as pleural effusion, emphysema, pulmonary consolidation, and atelectasis may occur.The underlying disease, wheezing, and age of onset less than 1.71 years (20 months) old are independent risk factors for severe ADV pneumonia.

The latest progress on novel coronavirus vaccination in kidney transplant recipients / 器官移植

Due to long-term use of immunosuppressant, poor immune function and a higher risk of critical diseases after novel coronavirus pneumonia in kidney transplant recipients, it is of significance to deliver prophylactic vaccination for this high-risk population. Studies have shown that the immune reaction of kidney transplant recipients to novel coronavirus vaccine is significantly lower than that of healthy counterparts. Standard vaccination program in the United States, such as 2 doses of messenger RNA (mRNA) vaccine, fails to provide sufficient protection for kidney transplant recipients. Many studies have proven that increasing the frequency of vaccination for kidney transplant recipients may enhance the vaccine efficacy. Nevertheless, the role of adjusting immunosuppressive therapy in increasing vaccine efficacy remains to be elucidated. In this article, the importance, effectiveness and particularity of novel coronavirus vaccine for kidney transplant recipients and the effect of immunosuppressive therapy on the efficacy of novel coronavirus vaccine were reviewed, aiming to provide reference on the vaccination for kidney transplant recipients.

Research progress on the epidemiology of human adenovirus infections / 预防医学

Abstract@#Human adenovirus (HAdV), which is characterized by infectivity, complex pathogenesis and multiple target organs, causes multiple organ infections in the respiratory system, gastrointestinal system and eyes, which seriously endangers human health. Various subspecies of HAdV has different tissue tropism, which presents diverse clinical symptoms and epidemiological characteristics. Based on the molecule biological characteristics of HAdV, this review summarizes the clinical symptoms and epidemiological characteristics of HAdV infections depending on tissue tropism, and describes the trends in HAdV epidemiology, so as to provide insights into prevention and control of HAdV infections.

Effects of manipulating lactate dehydrogenase gene on metabolism of HEK-293 and production of human adenovirus / 生物工程学报

Reducing lactate accumulation has always been a goal of the mammalian cell biotechnology industry. When animal cells are cultured in vitro, the accumulation of lactate is mainly the combined result of two metabolic pathways. On one hand, glucose generates lactate under the function of lactate dehydrogenase A (LDHA); on the other hand, lactate can be oxidized to pyruvate by LDHB or LDHC and re-enter the TCA cycle. This study comprehensively evaluated the effects of LDH manipulation on the growth, metabolism and human adenovirus (HAdV) production of human embryonic kidney 293 (HEK-293) cells, providing a theoretical basis for engineering the lactate metabolism in mammalian cells. By knocking out ldha gene and overexpression of ldhb and ldhc genes, the metabolic efficiency of HEK-293 cells was effectively improved, and HAdV production was significantly increased. Compared with the control cell, LDH manipulation promoted cell growth, reduced the accumulation of lactate and ammonia, significantly enhanced the efficiency of substrate and energy metabolism of cells, and significantly increased the HAdV production capacity of HEK-293 cells. Among these LDH manipulation measures, ldhc gene overexpression performed the best, with the maximum cell density increased by about 38.7%. The yield of lactate to glucose and ammonia to glutamine decreased by 33.8% and 63.3%, respectively; and HAdV titer increased by at least 16 times. In addition, the ATP production rate, ATP/O2 ratio, ATP/ADP ratio and NADH content of the modified cell lines were increased to varying degrees, and the energy metabolic efficiency was significantly improved.

Hyperosmotic stress and perfusion culture strategies increase the yield of recombinant adenoviral vector produced by HEK 293 cells / 生物工程学报

With various diseases ravaging internationally, the demands for recombinant adenoviral vector (Adv) vaccines have increased dramatically. To meet the demand for Adv vaccine, development of a new cell culture process is an effective strategy. Applying hyperosmotic stress in cells before virus infection could increase the yield of Adv in batch culture mode. Emerging perfusion culture can significantly increase the yield of Adv as well. Therefore, combining the hyperosmotic stress process with perfusion culture is expected to improve the yield of Adv at high cell density. In this study, a shake flask combined with a semi-perfusion culture was used as a scaled-down model for bioreactor perfusion culture. Media with osmotic pressure ranging from 300 to 405 mOsm were used to study the effect of hyperosmotic stress on cell growth and Adv production. The results showed that using a perfusion culture process with a hyperosmotic pressure medium (370 mOsm) during the cell growth phase and an isosmotic pressure medium (300 mOsm) during the virus production phase effectively increased the yield of Adv. This might be due to the increased expression of HSP70 protein during the late phases of virus replication. The Adv titer in a bioreactor with such a process reached 3.2×1010 IFU/mL, three times higher than that of the traditional perfusion culture process. More importantly, this is the first time that a strategy of combining the hyperosmotic stress process with perfusion culture is applied to the production of Adv in HEK 293 cells. It also reveals the reason why the hyperosmotic stress process increased the yield of Adv, which may facilitate the process optimization of for producing other Adv in HEK 293 cells.

Research progress of flavivirus vaccines based on adenovirus vector / 预防医学

@#There are more than 70 species of flaviviruses, including Zika virus, Dengue virus and Japanese encephalitis virus, and more than 33 species are known to be capable of infecting humans. Only three flavivirus vaccines have been approved, and there is a lack of safe and effective vaccines for most flaviviruses. Adenovirus-vectored vaccines have high safety, low cost, and convenience to store and transport. Currently, two adenovirus-vectored Zika vaccines are under early clinical trials, and adenovirus-vectored vaccines for Dengue virus, Japanese encephalitis virus, West Nile virus and yellow fever virus are still in the phase of animal experiment. In the development of adenovirus-vectored flavivirus vaccines, there are still problems of pre-existing immunity to adenovirus, the insufficient immunogenicity of adenovirus vectors and the antibody-dependent enhancement effects among flavivirus. Based on relevant publications from January 2006 to June 2023, this article reviews the current status, challenges and solutions of the research into adenovirus-vectored flavivirus vaccines, so as to provide the reference for the development of relevant vaccines.

Identification and complete genome sequencing of human adenovirus type 55 isolated from a patient with acute hemorrhagic conjunctivitis / 上海预防医学

ObjectiveTo determine the genomic characteristics of a subgenus B human adenovirus strain isolated in Shanghai in 2021. MethodsAn adenovirus type 55 strain was isolated and identified from a patient with acute hemorrhagic conjunctivitis （AHC）. Complete genome of the strain was obtained using the next-generation sequencing （NGS）. Phylogenetic trees were reconstructed based on the sequences of Hexon， Fiber， Penton and complete genome to genomically characterize this strain. ResultsPhylogenetic analysis based on the complete genome classified this strain （MH2021001） into subgenus B， subspecies B2 of HAdV-55. Hexon gene of MH2021001 had close phylogenetic relationship with HAdV-11， while Fiber and Penton genes had close relationship with HAdV-14. The MH2021001 showed high nucleotide identity with currently prevalent HAdV⁃55 strains （>99.90%）. The complete genome had 99.96% nucleotide identity to the 73-GD_CHN_2016 strain isolated in Guangdong. In addition， the amino acid sequence of MH2021001 had several substitutions in regions coding for E1B， L4， E3 and L5. ConclusionThis strain has been classified to HAdV-B55. No recombination event is identified in the complete genome. Due to multiple amino acid substitutions， the biological characteristics of the strain need to be further identified.

Role of Mitophagy in Myocardial Ischemia/Reperfusion Injury and Chinese Medicine Treatment / 中国结合医学杂志

Mitophagy is one of the important targets for the prevention and treatment of myocardial ischemia/reperfusion injury (MIRI). Moderate mitophagy can remove damaged mitochondria, inhibit excessive reactive oxygen species accumulation, and protect mitochondria from damage. However, excessive enhancement of mitophagy greatly reduces adenosine triphosphate production and energy supply for cell survival, and aggravates cell death. How dysfunctional mitochondria are selectively recognized and engulfed is related to the interaction of adaptors on the mitochondrial membrane, which mainly include phosphatase and tensin homolog deleted on chromosome ten (PTEN)-induced kinase 1/Parkin, hypoxia-inducible factor-1 α/Bcl-2 and adenovirus e1b19k Da interacting protein 3, FUN-14 domain containing protein 1 receptor-mediated mitophagy pathway and so on. In this review, the authors briefly summarize the main pathways currently studied on mitophagy and the relationship between mitophagy and MIRI, and incorporate and analyze research data on prevention and treatment of MIRI with Chinese medicine, thereby provide relevant theoretical basis and treatment ideas for clinical prevention of MIRI.

Establishment and preliminary application of dual fluorescent quantitative PCR for detection of RV and CAV2 / 公共卫生与预防医学

Objective To investigate the shedding of CAV2-ΔE3-CGS after immunization and the background of canine adenovirus (CAV) infection, and to establish a dual fluorescent quantitative PCR detection method for rabies virus (RV) and canine adenovirus type 2 (CAV2). Methods A dual fluorescent quantitative PCR detection method was established by designing specific primers and probes for E1 gene of CAV and G gene of RV for the detection of CAV2-ΔE3-CGS. Oral swabs, anal swabs and environmental samples of stray dogs from experimental animal farm and dog detention center were tested. Results The standard curves generated by this method were Y=-3.351 × logX + 44.895, R2 = .999 and Y=-3.413 × logX + 45.192, R2=0.996, respectively. The linear relationships were good, and the minimum detection limits were both 102 copies/μL. CAV2-ΔE3-CGS was not detected in experimental animal farm. CAV was detected in dog detention center, and the positive rates were 5.88% (5/85) in oral swabs, 8.24% (7/85) in anal swabs, and 4% (1/25) in environmental samples. Conclusion The dual fluorescent quantitative PCR method can be used for the detection of CAV2-ΔE3-CGS after immunization and the investigation of CAV infection. The present study has shown that no CAV2-ΔE3-CGS has been detected after immunization and CAV infection rate of stay dogs is low in Shanghai. CAV2-ΔE3-CGS oral immunization meets requirement and is applicable.