Control of highly pathogenic avian influenza through vaccination

The stamping-out strategy has been used to control highly pathogenic avian influenza viruses in many countries,driven by the belief that vaccination would not be successful against such viruses and fears that avian influenza virus in vaccinated birds would evolve more rapidly and pose a greater risk to humans.In this review,we summarize the successes in controlling highly pathogenic avian influenza in China and make suggestions regarding the requirements for vaccine selection and effectiveness.In addition,we present evidence that vaccination of poultry not only eliminates human infection with avian influenza virus,but also significantly reduces and abolishes some harmful characteristics of avian influenza virus.

Development and Assessment of Two Highly Pathogenic Avian Influenza(HPAI) H5N6 Candidate Vaccine Viruses for Pandemic Preparedness

Objective In China, 24 cases of human infection with highly pathogenic avian influenza(HPAI) H5 N6 virus have been confirmed since the first confirmed case in 2014. Therefore, we developed and assessed two H5 N6 candidate vaccine viruses(CVVs).Methods In accordance with the World Health Organization(WHO) recommendations, we constructed two reassortant viruses using reverse genetics(RG) technology to match the two different epidemic H5 N6 viruses. We performed complete genome sequencing to determine the genetic stability. We assessed the growth ability of the studied viruses in MDCK cells and conducted a hemagglutination inhibition assay to analyze their antigenicity. Pathogenicity attenuation was also evaluated in vitro and in vivo.Results The results showed that no mutations occurred in hemagglutinin or neuraminidase, and both CVVs retained their original antigenicity. The replication capacity of the two CVVs reached a level similar to that of A/Puerto Rico/8/34 in MDCK cells. The two CVVs showed low pathogenicity in vitro and in vivo, which are in line with the WHO requirements for CVVs.Conclusion We obtained two genetically stable CVVs of HPAI H5N6 with high growth characteristics,which may aid in our preparedness for a potential H5N6 pandemic.

Assessment of the Level of Knowledge of the Nature of Highly Pathogenic Avian Influenza Demonstrated by the Nigerian Veterinary Laboratory Staff Involved in HPAI Diagnosis in Nigeria

The study was designed to evaluate the level of knowledge of Nigerian Veterinary Laboratory Staff on the nature of Highly Pathogenic Avian Influenza (HPAI) disease using structured questionnaires. The study comprised the Staff of National Veterinary Research Institute (NVRI) and five reference Veterinary Teaching Hospitals (VTH) designated for HPAI diagnosis. A total of 69 questionnaires were distributed to the laboratory staff. Questions on the general nature of the disease such as the cause, signs, mode of transmission, methods of identification, lesions, control and prevention, etc. were asked. The results showed that 77.38% of the staff answered all the questions correctly indicating their considerable knowledge of the HPAI disease. Considerable percentage of the staff listed correctly the equipment used for serology (36.23%) and RT-PCR (31.88%). Interestingly only 13.04% of the staff listed correctly the equipment used in rapid tests despite the fact that they are simpler and recommended for all P2 laboratories. In conclusion, the veterinary laboratory staff assessed demonstrated a significant level of knowledge on HPAI diagnosis;however, most of their laboratories lack the structure, organization, funds and basic facilities required for effective HPAI diagnosis.

Establishment of a Risk Assessment Framework for Analysis of the Spread of Highly Pathogenic Avian Influenza

To evaluate the risk of highly pathogenic avian influenza （HPAI） in China's Mainland, a risk assessment framework was built. Risk factors were determined by analyzing the epidemic data using the brainstorming method; the analytic hierarchy process was designed to weigh risk factors, and the integrated multicriteria analysis was used to evaluate the final result. The completed framework included the risk factor system, data standards for risk factors, weights of risk factors, and integrated assessment methods. This risk assessment framework can be used to quantitatively analyze the outbreak and spread of HPAI in China's Mainland.

Characterization of Highly Pathogenic Avian Influenza H5N1 Viruses Isolated from Domestic Poultry in China

The highly pathogenic avian influenza （HPAI） H5N1 virus has caused several outbreaks in domestic poultry. Despite great efforts to control the spread of this virus, it continues to evolve and poses a substantial threat to public health because of a high mortality rate. In this study, we sequenced whole genomes of eight H5N1 avian influenza viruses isolated from domestic poultry in eastern China and compared them with those of typical influenza virus strains. Phylogenetic analyses showed that all eight genomes belonged to clade 2.3.2.1 and clade 7.2, the two main circulating clades in China. Viruses that clustered in clade 2.3.2.1 shared a high degree of homology with H5N1 isolates located in eastern Asian. Isolates that clustered in clade 7.2 were found to circulate throughout China, with an east-to-west density gradient. Pathogenicity studies in mice showed that these isolates replicate in the lungs, and clade 2.3.2.1 viruses exhibit a notably higher degree of virulence compared to clade 7.2 viruses. Our results contribute to the elucidation of the biological characterization and pathogenicity of HPAI H5N1 viruses.

Highly pathogenic avian influenza H5N1 Clade 2.3.2.1c virus in migratory birds,2014–2015

A novel Clade 2.3.2.1c H5N1 reassortant virus caused several outbreaks in wild birds in some regions of China from late 2014 to 2015.Based on the genetic and phylogenetic analyses,the viruses possess a stable gene constellation with a Clade 2.3.2.1c HA,a H9N2-derived PB2 gene and the other six genes of Asian H5N1-origin.The Clade 2.3.2.1c H5N1 reassortants displayed a high genetic relationship to a human H5N1 strain(A/Alberta/01/2014).Further analysis showed that similar viruses have been circulating in wild birds in China,Russia,Dubai(Western Asia),Bulgaria and Romania(Europe),as well as domestic poultry in some regions of Africa.The affected areas include the Central Asian,East Asian-Australasian,West Asian-East African,and Black Sea/Mediterranean flyways.These results show that the novel Clade 2.3.2.1c reassortant viruses are circulating worldwide and may have gained a selective advantage in migratory birds,thus posing a serious threat to wild birds and potentially humans.

Knowledge,attitudes and practices towards avian influenza A(H5N1) among Cambodian women:A cross-sectional study

Objective:To measure highly pathogenic avian influenza(HPAI)-related knowledge,attitudes, and practices(KAPs) among Cambodian women.Methods:This cross-sectional study selected 246 married women aged between 18-55 years who had backyard poultry and lived at least one year in the areas of the survey through multi-stage cluster sampling.An average score of correct answers was generated to evaluate respondents’ knowledge(Good/Poor).attitudes(Positive/ Negative),and practices(Good/Bad).Results:We reported that about half of the respondents had good knowledge and good practices and four-fifth of them had positive attitudes towards HPAl.Odds ratios(ORs) and 95%confidence intervals(CIs) were estimated through a logistic regression model to explore contributing factors that raise their KAP levels.Most of the sources were significant in increasing knowledge of the respondent,like television(OR=l.6.95%CI=1.0- 2.7),radio(OR=2.5.95%CI=l.3-4.9).leaflets/booklets(OR=2.1,95%CI=1.2-3.9),school students (OR=18.4.95%CI=2.4-142.9).village health volunteers(OR=4.5.95%CI=2.2-10.9) etc.Factors such as television(OR=3.7.95%CI=2.1-6.4).leaflets/booklets(OR=2.6,95%CI7=1.4-5.1).and public health staff(OR=2.2,95%CI=1.2-4.1) had similar influence on practices.Although,we found similar effect on raising the attitudes of the responded,it was not significant.Conclusions: We report a satisfactory level of positive attitudes,and moderate level of knowledge and practices related to HPAI among Cambodian women.Raising KAPs through television,radio and other medias may be more efficient than using usual information,education and communication materials to prevent HPAI.

International chicken trade and increased risk for introducing or reintroducing highly pathogenic avian influenza A (H5N1) to uninfected countries

Every year billions of chickens are shipped thousands of miles around the globe in order to meet the ever increasing demands for this cheap and nutritious protein source.Unfortunately,transporting chickens internationally can also increase the chance for introducing zoonotic viruses,such as highly pathogenic avian influenza A(H5N1)to new countries.Our study used a retrospective analysis of poultry trading data from 2003 through 2011 to assess the risk of H5N1 poultry infection in an importing country.We found that the risk of infection in an importing country increased by a factor of 1.3(95%CI:1.1e1.5)for every 10-fold increase in live chickens imported from countries experiencing at least one H5N1 poultry case during that year.These results suggest that the risk in a particular country can be significantly reduced if imports from countries experiencing an outbreak are decreased during the year of infection or if biosecurity measures such as screening,vaccination,and infection control practices are increased.These findings show that limiting trade of live chickens or increasing infection control practices during contagious periods may be an important step in reducing the spread of H5N1 and other emerging avian influenza viruses.

Case report for human infection with a highly pathogenic avian influenza A(H5N6) virus in Beijing, China 2019

Bird infections with highly pathogenic avian influenza A(H5N6) viruses have been identified since 2014. With very limited occasion, the virus could sporadically spilled over to infect humans. It has been recognized that all human infections were within southern region of China's Mainland until the case reported here in Beijing in Aug. 2019. This was the first human case infected with highly pathogenic avian influenza A(H5N6) virus in northern China. The infection was confirmed by real-time RT-PCR assay. The whole genome sequences were obtained from clinical sample. Genetic characteristics of the virus were identified similar to those of previous avian influenza A(H5N6) viruses, retaining the main features of the avian influenza virus.

The Evidence of Clade 7.1 Avian Influenza Virus (H5N1) in Qinghai Lake

The highly pathogenic influenza A virus subtype H5N1 spread throughout Asia since 2003, reached to Europe in 2005, and the Middle East, as well as Africa and caused a global concern for a potential pandemic threat last decade. A Clade 2.3.2 H5N1 virus became dominate in the Qinghai Lake region in 2009 with sporadic mammal cases of infection and transferred to Russia and Europe through wild migratory birds. Currently, HPAI H5N1 of clades 2.3.4, 2.3.2, and 7 are the dominant co-circulating H5N1 viruses in poultry in Asia. 2.3.2 Clade is dominant in wild birds through the world whereas there is no evident data about Clade 7 circulation in wild birds. We detected HPAI H5N1 virus of Clade 7.1 in Qinghai Lake, that closely related to Shanxi-like and Vietnam viruses co-circulating in poultry. This is the first report of Clade 7.1 H5N1 in wild birds. Based on phylogenetic analyses, the virus can be originated from Clade 7.1 virus gene pool that spread in Vietnam and Chinese poultry and could spread with migratory birds to Qinghai Lake. The Qinghai Lake continues to be significant hotspot for H5N1 surveillance since the regular outbreaks occurred there in wild birds and mammals. Based on these facts and findings, the related researchers should pay more attention to the Qinghai Lake basin as significant hotspot for H5N1 avian influenza surveillance since the regular H5N1 outbreaks occurred there in wild birds with sporadic mammal cases of infection.

Surveillance for highly pathogenic avian influenza in wild birds in the USA

As part of the USA’s National Strategy for Pandemic Influenza,an Interagency Strategic Plan for the Early Detection of Highly Pathogenic H5N1 Avian Influenza in Wild Migratory Birds was developed and implemented.From1April2006 through 31 March 2009,261946 samples fromwild birds and 101457 wild bird fecalsamples were collected in the USA;no highly pathogenic avian influenza was detected.The United States Department of Agriculture,and state and tribal cooperators accounted for 213115(81%)of the wild bird samples collected;31,27,21 and 21%of the samples were collected from theAtlantic,Pacific,Central and Mississippi flyways,respectively.More than 250 species of wild birds in all 50 states were sampled.The majority of wild birds(86%)were dabbling ducks,geese,swans and shorebirds.The apparent prevalence of low pathogenic avian influenza viruses during biological years 2007 and 2008 was 9.7 and 11.0%,respectively.The apparent prevalence of H5 and H7 subtypes across all species sampled were 0.5 and 0.06%,respectively.The pooled fecal samples(n=101539)positive for low pathogenic avian influenza were 4.0,6.7 and 4.7%for biological years 2006,2007 and 2008,respectively.The highly pathogenic early detection system for wild birds developed and implemented in the USA represents the largest coordinated wildlife disease surveillance system ever conducted.This effort provided evidence that wild birds in the USA were free of highly pathogenic avian influenza virus(given the expected minimum prevalence of 0.001%)at the 99.9%confidence level during the surveillance period.

Assessment of vaccination strategies against highly pathogenic avian influenza in China

Vaccination for highly pathogenic avian influenza(HPAI)has been implemented in China for a decade,however,the virus is still present in poultry.A series of recombinant vaccines,Re-1 to Re-7,have been developed and used,and Re-8 will also be used in clinical settings to prevent the prevailing flu strains.The question remains,when can China eradicate the disease?Here,we review the epidemiology of H5 HPAI along with the development,usage and problems of vaccines.Further suggestions for controlling the disease in China are provided.

甘肃省肉仔鸡HPAIH5亚型母源抗体消长规律研究

用HI法对甘肃省肉仔鸡H5N1型禽流感(HPAI)母源抗体消长规律进行动态监测,结果显示,肉仔鸡母源抗体保护期长达67 d。其中,种蛋的母源抗体效价最高为9.48 log2,1日龄肉仔鸡母源抗体效价均值为7.95 log2,4 d降为7.81 log2,7 d母源抗体升至高峰,达8.57log2,接近父母代抗体效价8.80 log2;随后,抗体效价逐渐下降并略有波动;67-70 d,抗体效价迅速降至1.67 log2,79 d趋于消失,为1.83 log2。该测定结果提示,HI试验在HPAI诊断中意义不大,仅能作为参考,确诊需进行病毒的分离培养。

Adoption of HPAI biosecurity measures： The Chinese broiler industry

Highly pathogenic avian influenza （HPAI） contamination via wild birds and rodents poses a threat to food security and safety. As chicken meat comprises an increasing proportion of diet in China, it is useful to determine whether broiler farmers are adopting wild bird and rodent controls to minimize the risk of HPAI impacts on food supply. Our study surveyed a cross sectional sample of 331 Chinese broiler farmers in six provinces. We find that only 47% of farmers （mainly farmers with large herds） adopted control measures against wild birds and rodents, while 14% adopted no measures. Farm size was the biggest driver of adoption followed by proportion of farm revenue derived from broiler production. However, southern farmers were at a far greater probability of non-adoption. We suggest that assistance in the form of education/training programs and subsidized traps or baiting controls across smaller producers could help raise of the adoption level toward more effective HPAI control.

中国禽群高致病性禽流感发生状况及其风险预测

本研究旨在探讨中国高致病性禽流感的发生状况,并对其风险态势进行尝试性的预测。作者在流行病学数据挖掘和模型分析基础上,基于OIE风险评估理论,结合昆虫生态学格局分析和Bernoulli统计模型,对中国2004年以来高致病性禽流感的发生状况及其风险态势进行了分析和预测。结果显示:(1)中国家禽养殖数量大,养殖模式复杂,种类繁多,养殖的区域特征显著,给中国HPAI的防控带来了挑战;(2)中国自2004年发生HPAI以来,疫情发生的时-空连续性强,突出表现在疫情发生的季节、区域和宿主特征明显,提示科学的区域化管理和协作的重要性;(3)在发展态势上,由于区域周边疫情特点和中国禽群养殖及风险管理状况,使得中国局部地区禽群在相当长的一段时间内存在较大的暴露和感染的风险;(4)风险预测上,利用Bernoulli统计模型和信息技术,预测2008年中国禽群在90%的置信区间内最有可能会有2～10起HPAI疫情发生,发生6起左右的高致病性禽流感疫情可能性较大,其中在中国东南部分地区局部区域禽群具有较高的疫情暴露和发生风险。本研究表明中国禽群持续发生HPAI疫情的可能性大,开展基于风险的区划和管理对疫情发生和控制具有积极意义。

安徽省规模禽场高致病性禽流感血清学横断面研究与风险因素分析

为掌握安徽省禽群的禽流感免疫抗体水平以及引起免疫不合格的主要风险因素,在2018年春季集中免疫前,对安徽省7个市开展了H5和H7亚型禽流感血清学检测和问卷调查。按照分层随机抽样方法,在7个市选择种禽场、蛋禽场和肉禽场,共140个养禽场,在每个养殖场内随机抽取35份血清样品,进行HI抗体检测,并根据检测结果,计算群体水平的真实免疫合格率/阳性率;将H5或H7亚型禽流感抗体群体合格率低于70%的养禽场定义为免疫不合格场,进行单因素风险分析。结果显示:安徽省7个市H5(Re-8株)和H7N9(Re-1株)的场群真实抗体合格率/阳性率分别为67.7%(95%CI:60.0%~75.4%)和39.8%(95%CI:31.7%~47.9%),62个养禽场的H5或H7亚型抗体未达标(<70%);存栏量越小(P=0.046)、免疫次数越少(P=0.016),H5或H7亚型免疫抗体不合格的可能性越高(P<0.05);存栏量小于4000羽、免疫后时间≤21 d或≥91 d、家禽日龄≤26或≥2200、免疫次数≤1、规模化饲养鹅,以及不使用“H5+H7”二价灭活疫苗(OR=2.66,95%CI:1.34~5.35)、周边3 km内有野禽栖息地(OR=2.78,95%CI:1.10~7.65)的养禽场的H5或H7亚型免疫抗体不合格风险相对较高。结果表明:2018年春季安徽省7个市H5、H7亚型禽流感免疫抗体保护率下降幅度明显,需要及时开展春季集中免疫;养禽场规模、类型以及家禽种类、是否免疫疫苗、免疫次数、养禽场周边是否有野禽栖息地等,均与群体的禽流感免疫抗体合格率显著相关,因此在制定强制免疫计划时,应视情况进行分类指导,实时补免或增加免疫次数。此外,养禽场选址时应尽量避开野禽栖息地。本研究为决策部门制定禽流感免疫方案提供了技术支撑。

中国高致病性禽流感疫情流行特征及其防控措施

高致病性禽流感在中国持续流行,H7N9病毒感染人类出现新的疫情特征,需要进一步强化防控措施。针对疫情点多面广、H9亚型泛滥重组和交叉感染带毒等特点,应做到依法处置疫情,加强家禽企业生物安全隔离区建设,谨慎使用疫苗,完善监测系统和兽医管理体制。结合禽流感最新疫情流行特征,对中国禽流感防控体系建设进行了探讨。

H5N1亚型高致病性禽流感病毒在鸽子体内分布的研究

应用实验室分离的鸭源高致病性禽流感病毒(AIV)A/Duck/Harbin/01/2004(H5N1)毒株,经SPF鸡胚增殖病毒,通过滴鼻途径人工感染鸽子,通过临床观察、剖检、HA、HI、PCR等方法进行体内分布和毒性研究。结果病死鸽子的肺脏损害最为严重,其次为脑、法氏囊;在病死鸽子的肺、肝、肾、脑中检测到病毒,其中肺中的病毒含量最高,脑、肝、肾中的病毒含量次之,再次为心和脾;各组织HA和HI检测阳性;各组织PCR检测到目的基因;对照组所有检测均为阴性。结果表明AIV在鸽子体内可以复制,并在肺、脑内病毒含量较高,为预防AIV奠定了基础。

虎源高致病性禽流感病毒H5N1在实验小鼠体内分布与含量测定

将本实验室分离、鉴定的虎源高致病性禽流感病毒A/Tiger/Harbin/01/2003(H5N1)株第3代鸡胚尿囊液进行10-3倍稀释,接种小鼠,待小鼠死亡后无菌采取心、肝、脾、肺、肾、脑六种脏器分别研磨制成乳剂,接种MDCK细胞,以细胞病变为判定指标,计算病死小鼠不同脏器内病毒含量。同时用RT-PCR、血凝-血凝抑制试验以及电镜负染的方法进行鉴定和观察。结果显示,可在小鼠的肺脏、肝脏、肾脏、脑组织中检出所接种的病毒,肺脏中的病毒含量最高,脑中的病毒含量次之,肝脏和肾脏中的病毒含量最少。可在肺、肝、肾、脑组织与感染细胞培养物中扩增出与理论值一致的核酸条带,肺乳剂上清液感染细胞培养物中可检出1∶23的血凝效价,电镜观察在肺乳剂上清液及其感染细胞培养物中可见到典型的流感病毒颗粒。

NASBA快速检测禽流感H5亚型病毒

采用建立的依赖核酸序列的扩增(Nucleicacidsequencebasedamplification,NASBA)对禽流感病毒3株H5亚型、1株H1、H3、H6亚型、3株禽流感H9亚型、5株不同宿主来源的新城疫病毒、鸭肝炎病毒、鸭瘟病毒、SPF鸡胚尿囊液及禽流感(H9)疫苗、新城疫疫苗、传染性法氏囊病疫苗、传染性支气管炎疫苗进行检测,结果NASBA(H5试剂)仅检测到禽流感病毒H5亚型,表明方法的特异性强。采用已知禽流感病毒A/Chicken/HK/1000/97(H5N1)的鸡胚尿囊液(ELD5010-7.5/mL),经10倍连续稀释,将经典的鸡胚病原分离法和NASBA进行比较,二种方法的灵敏度相当。用A/Chicken/HK/1000/97(H5N1)病毒人工感染SPF鸡、商品鸡,采用NASBA和病原分离法同时对人工感染鸡的粪拭子、血液进行了动态检测;采集感染死亡鸡的组织脏器,共检测了101个组织脏器,两种方法的符合率为90%(87/97)。