医学病毒学生物安全三级动物实验室的建立

目前国内多个机构已建立了BSL-3实验室,这些实验室能够满足细胞水平和动物水平的病原微生物实验活动.医学病毒学生物安全三级动物实验室(ABSL-3)可分为小动物实验室和中(大)动物实验室.小动物实验室主要开展小鼠、大鼠、仓鼠等啮齿类动物的感染性实验活动;中(大)动物实验室主要开展小型猪、猴等实验.动物三级实验室特别是开展中(大)动物实验的高等级实验室,其实验室布局,设备设施,管理体系及流程会有很大的差别.本文从使用者的角度,通过所在机构医学病毒学生物安全三级动物实验室(ABSL-3)的建设与实践,对于筹备和建立以及使用过程中的所面对的问题以及应对,进行了提炼和归纳,分析和探讨,旨在为建设和使用医学病毒学高致病性生物安全三级动物实验室提供借鉴和参考.     

宏基因组学与动物病原微生物检测

宏基因组学（ metagenome）是直接从土壤、海水、人及动物胃肠道、口腔、呼吸道、皮肤等环境中获取样品DNA,利用载体将其克隆到替代宿主细胞中构建宏基因文库,以高通量检测为主要技术来研究特定环境中全部微生物的基因组及筛选活性物质和基因的新兴学科。利用宏基因组学技术不仅能够有效地检测特定环境的微生物群落结构,扩展了微生物资源的利用空间,发展了新兴的高通量检测技术,丰富了生物信息学内容。基于宏基因组学研究方法在环境微生物研究中的优势,对近年来相关领域、方法及其在人及动物病原微生物研究中的应用进行综述,以期将此方法用于实验动物病原微生物的调查分析及动物疫情、生物安全的监测。     

建立洁净区微生物数据库与无菌药品GMP生产过程控制的探讨

目的 建立洁净区微生物数据库,为追溯洁净区微生物污染来源提供依据,为无菌药品生产过程控制提供有力指导.方法 对乙型脑炎减毒活疫苗生产车间洁净区环境和操作人员进行微生物负载检测,并对该车间注射用水、纯化水系统样品进行微生物限度检测,鉴别研究相应分离菌,建立洁净区微生物数据库.同时对乙型脑炎减毒活疫苗中间产品进行无菌检查检测,并对阳性结果分离菌进行鉴别分析.然后根据建立的洁净区微生物数据库对阳性结果举例进行溯源探讨分析.结果 洁净区环境和人员主要存在里拉/藤黄微球菌、人葡萄球菌、表皮葡萄球菌以及科氏葡萄球菌科氏亚种等革兰阳性菌和蜡样芽胞杆菌等,水系统中则主要存在少动鞘氨醇单胞菌和铫子芽胞杆菌等.中间产品无菌阳性结果主要存在里拉/藤黄微球菌、蜡样芽胞杆菌以及奇异变形菌等,经调查分析,分别为操作过程中经环境偶然带入或试验动物操作不慎带入.结论 建立洁净区微生物数据库是追溯产品微生物污染来源的有效方法,能够为无菌药品GMP生产过程制定有效的控制措施提供依据,并使其更有针对性.     

关于高级别生物安全实验室若干管理要素的探讨

目前,我国已有两家生物安全四级实验室和五十多家生物安全三级实验室完成建造并通过实验室认可。生物安全管理体系的技术要素和管理要素是确保高级别生物安全实验室生物安全的关键。本文探讨了高级别生物安全实验室区别于其他体系实验室(如ISO 17025检测和校准实验室与ISO 15189医学实验室)的若干个特征性管理要素,这些要素包括潜在污染的周期性评估、失活验证实验、分子诊断实验的无生物风险的阳性对照、微生物的不同封闭措施及生物安全文化。潜在污染的周期性评估与失活验证实验是高级别生物安全实验室日常管理的强制性措施。分子诊断实验的无生物风险的阳性对照与微生物的不同封闭措施应用于微生物实验(尤其是病毒学实验)从而降低暴露风险。生物安全文化应全面施行,以预防生物安全事件的再次发生。     

实验动物与生物安全

实验动物是从事科学研究、教学、生产、检定等的重要工具和支撑条件.随着科学技术的飞速发展,科学研究中使用的实验动物和实验用动物的种类、品系越来越多,生物安全问题已经明显地威胁到生物多样性、生态环境和人类健康.本文分析了实验动物的潜在生物安全危害,探讨了实验动物生物安全管理方面存在的问题并就应对实验动物生物安全提出了一些建议.     

喷气燃料中污染微生物检测方法概况

微生物污染不仅降解喷气燃料,还严重威胁储存和飞行安全.喷气燃料中污染微生物的检测是有效治理微生物污染的前提.基于此,本文将喷气燃料中污染微生物检测方法分为传统法、分析生物学法以及分子生物学方法,并对三种方法优缺点进行了详细介绍.最后对喷气燃料中污染微生物检测方法进行了展望.     

人间传染的病原微生物菌(毒)种保藏机构运行与管理探讨

2017年我国人间传染的病原微生物菌(毒)种保藏机构完成指定工作,作为国家生物安全工作的重要组成部分,做好病原微生物菌(毒)种保藏机构运行与管理,确保国家生物安全,已成为坚持和落实总体国家安全观一项重要内容和具体体现。保藏机构运行管理应将病原微生物菌(毒)种的合理利用和安全管理两方面相结合,在确保国家生物安全的前提下,统一病原微生物保藏技术标准和数据规范体系,提升保藏机构能力和水平,不断适应国家生物安全战略,以及传染性疾病防控、生物技术和生物产业发展需求。     

生物芯片技术及其在水体环境生物监测中的应用

水源微生物污染严重威胁着人类的健康。为有效控制水体环境生物安全,水体环境中微生物快速而准确地监测是关键的技术基础之一。生物芯片(biochip)技术是20世纪90年代初期发展起来的一门新兴技术,能迅速检测出水中的微生物。本文阐述了生物芯片的基本概念,对基因芯片技术作了简介。重点叙述了生物芯片技术在水体环境生物监测方面的应用,并就其应用前景作了展望。     

啮齿类实验动物行为学视觉测量方法

啮齿类动物广泛运用于视觉疾病的造模;为了合理评估动物模型的视觉疾病损害程度和治疗手段干预效果,需要掌握合适的啮齿类实验动物视觉检测手段。视觉行为学检测具有简明、直观、非侵入、可重复的优良特性,能够弥补其他视觉测量手段的缺陷。从定性以及定量两个角度对常用的啮齿类实验动物行为学视觉测量方法进行综述,有助于实验者选择适当的检测方法,提升动物视觉检测效率。     

生物分析测试:探索生命科学奥秘的工具

生物分析测试主要包括开发生物分析方法和研制标准物质,是生物学研究、应用及生物安全监管与检测的重要组成部分。其主要是用分析化学、生物化学和分子生物学等手段,对核酸、蛋白质、代谢物及细胞、微生物等生物量的含量或数量进行快速或精准分析。     

Role of housing modalities on management and surveillance strategies for adventitious agents of rodents

Specific pathogen-free (SPF) rodents for modern biomedical research need to be free of pathogens and other infectious agents that may not produce disease but nevertheless cause research interference. To meet this need, rodents have been rederived to eliminate adventitious agents and then housed in room- to cage-level barrier systems to exclude microbial contaminants. Because barriers can and do fail, routine health monitoring (HM) is necessary to verify the SPF status of colonies. Testing without strict adherence to biosecurity practices, however, can lead to the inadvertent transfer of unrecognized, inapparent agents among institutions and colonies. Microisolation caging systems have become popular for housing SPF rodents because they are versatile and provide a highly effective cage-level barrier to the entry and spread of adventitious agents. But when a microisolation-caged colony is contaminated, the cage-level barrier impedes the spread of infection and so the prevalence of infection is often low, which increases the chance of missing a contamination and complicates the corroboration of unexpected positive findings. The expanding production of genetically engineered mutant (GEM) rodent strains at research institutions, where biosecurity practices vary and the risk of microbial contamination can be high, underscores the importance of accurate HM results in mitigating the risk of the introduction and spread of microbial contaminants with the exchange of mutant rodent strains among investigators and institutions.     

Epidemiology and disease control in everyday beef practice

It is important for food animal veterinarians to understand the interaction among animals, pathogens, and the environment, in order to implement herd-specific biosecurity plans. Animal factors such as the number of immunologically protected individuals influence the number of individuals that a potential pathogen is able to infect, as well as the speed of spread through a population. Pathogens differ in their virulence and contagiousness. In addition, pathogens have various methods of transmission that impact how they interact with a host population. A cattle population's environment includes its housing type, animal density, air quality, and exposure to mud or dust and other health antagonists such as parasites and stress; these environmental factors influence the innate immunity of a herd by their impact on immunosuppression. In addition, a herd's environment also dictates the "animal flow" or contact and mixing patterns of potentially infectious and susceptible animals. Biosecurity is the attempt to keep infectious agents away from a herd, state, or country, and to control the spread of infectious agents within a herd. Infectious agents (bacteria, viruses, or parasites) alone are seldom able to cause disease in cattle without contributing factors from other infectious agents and/or the cattle's environment. Therefore to develop biosecurity plans for infectious disease in cattle, veterinarians must consider the pathogen, as well as environmental and animal factors.     

Biosecurity on cattle farms: a study in north-west England

Background

Few studies have considered in detail the range of biosecurity practices undertaken on cattle farms, particularly within the UK. In this study, 56 cattle farmers in a 100 km² area of north-west England were questioned regarding their on-farm biosecurity practices, including those relating to animal movements, equipment sharing and companies and contractors visiting the farms.

Methodology/Principal Findings

There was great variation between farms in terms of the type of, and extent to which, biosecurity was carried out. For example, the majority of farmers did not isolate stock bought onto the farm, but a small proportion always isolated stock. Many farmers administered treatments post-movement, primarily vaccinations and anthelmintics, but very few farms reported carrying out any health checks after moving animals on. In addition, there appeared to be much variation in the amount of biosecurity carried out by the different companies and contractors visiting the farms. Deadstock collectors and contracted animal waste spreaders, although likely to have a high potential for contact with infectious agents, were reported to infrequently disinfect themselves and their vehicles.

Conclusions/Significance

These findings suggest that although certain biosecurity practices are undertaken, many are carried out infrequently or not at all. This may be due to many factors, including cost (in time and money), lack of proven efficacies of practices and lack of relevant education of veterinary surgeons, producers and other herd health specialists. Further research exploring the reasons for the lack of uptake is imperative if preventive medicine is to be utilised fully by the farming industry.     

Relative risks and approaches to biosecurity in the use of embryo technologies in livestock

Embryo technologies have been integrated into production systems for a variety of livestock species. As relates to transmission of infectious diseases, our working hypothesis has been that use of embryo transfer for distribution of germ plasm within and between herds and flocks is likely safer than the movement of postnatal animals. Indeed, research and experience generally have been supportive of this hypothesis. However, the relative risks of transmitting infectious agents via embryo transfer vary among donor species. Further, different methods of producing embryos appear to present different risks. This paper provides a comparative overview of the risks of transmitting infectious diseases via transfer of both in vivo- and in vitro-derived embryos in common domesticated livestock species. Also discussed are universal approaches to biosecurity in embryo production and transfer.     

Expert Opinion on the Perceived Effectiveness and Importance of On-Farm Biosecurity Measures for Cattle and Swine Farms in Switzerland

Biosecurity is crucial for safeguarding livestock from infectious diseases. Despite the plethora of biosecurity recommendations, published scientific evidence on the effectiveness of individual biosecurity measures is limited. The objective of this study was to assess the perception of Swiss experts about the effectiveness and importance of individual on-farm biosecurity measures for cattle and swine farms (31 and 30 measures, respectively). Using a modified Delphi method, 16 Swiss livestock disease specialists (8 for each species) were interviewed. The experts were asked to rank biosecurity measures that were written on cards, by allocating a score from 0 (lowest) to 5 (highest). Experts ranked biosecurity measures based on their importance related to Swiss legislation, feasibility, as well as the effort required for implementation and the benefit of each biosecurity measure. The experts also ranked biosecurity measures based on their effectiveness in preventing an infectious agent from entering and spreading on a farm, solely based on transmission characteristics of specific pathogens. The pathogens considered by cattle experts were those causing Bluetongue (BT), Bovine Viral Diarrhea (BVD), Foot and Mouth Disease (FMD) and Infectious Bovine Rhinotracheitis (IBR). Swine experts expressed their opinion on the pathogens causing African Swine Fever (ASF), Enzootic Pneumonia (EP), Porcine Reproductive and Respiratory Syndrome (PRRS), as well as FMD. For cattle farms, biosecurity measures that improve disease awareness of farmers were ranked as both most important and most effective. For swine farms, the most important and effective measures identified were those related to animal movements. Among all single measures evaluated, education of farmers was perceived by the experts to be the most important and effective for protecting both Swiss cattle and swine farms from disease. The findings of this study provide an important basis for recommendation to farmers and policy makers.     

Pathobiology and management of laboratory rodents administered CDC category A agents

The Centers for Disease Control and Prevention Category A infectious agents include Bacillus anthracis (anthrax), Clostridium botulinum toxin (botulism), Yersinia pestis (plague), variola major virus (smallpox), Francisella tularensis (tularemia), and the filoviruses and arenaviruses that induce viral hemorrhagic fevers. These agents are regarded as having the greatest potential for adverse impact on public health and therefore are a focus of renewed attention in infectious disease research. Frequently rodent models are used to study the pathobiology of these agents. Although much is known regarding naturally occurring infections in humans, less is documented on the sources of exposures and potential risks of infection to researchers and animal care personnel after the administration of these hazardous substances to laboratory animals. Failure to appropriately manage the animals can result both in the creation of workplace hazards if human exposures occur and in disruption of the research if unintended animal exposures occur. Here we review representative Category A agents, with a focus on comparing the biologic effects in naturally infected humans and rodent models and on considerations specific to the management of infected rodent subjects. The information reviewed for each agent has been curated manually and stored in a unique Internet-based database system called HazARD (Hazards in Animal Research Database, http://helab.bioinformatics.med.umich.edu/hazard/) that is designed to assist researchers, administrators, safety officials, Institutional Biosafety Committees, and veterinary personnel seeking information on the management of risks associated with animal studies involving hazardous substances.     

Lurking in the shadows: emerging rodent infectious diseases

Rodent parvoviruses, Helicobacter spp., murine norovirus, and several other previously unknown infectious agents have emerged in laboratory rodents relatively recently. These agents have been discovered serendipitously or through active investigation of atypical serology results, cell culture contamination, unexpected histopathology, or previously unrecognized clinical disease syndromes. The potential research impact of these agents is not fully known. Infected rodents have demonstrated immunomodulation, tumor suppression, clinical disease (particularly in immunodeficient rodents), and histopathology. Perturbations of organismal and cellular physiology also likely occur. These agents posed unique challenges to laboratory animal resource programs once discovered; it was necessary to develop specific diagnostic assays and an understanding of their epidemiology and transmission routes before attempting eradication, and then evaluate eradication methods for efficacy. Even then management approaches varied significantly, from apathy to total exclusion, and such inconsistency has hindered the sharing and transfer of rodents among institutions, particularly for genetically modified rodent models that may not be readily available. As additional infectious agents are discovered in laboratory rodents in coming years, much of what researchers have learned from experiences with the recently identified pathogens will be applicable. This article provides an overview of the discovery, detection, and research impact of infectious agents recently identified in laboratory rodents. We also discuss emerging syndromes for which there is a suspected infectious etiology, and the unique challenges of managing newly emerging infectious agents.     

Clinical considerations in rodent bioimaging

Imaging modalities such as micro-computed tomography (micro-CT), micro-positron emission tomography (micro-PET), high-resolution magnetic resonance imaging (MRI), optical imaging, and high-resolution ultrasound are rapidly becoming invaluable research tools. These advanced imaging technologies are now commonly used to investigate rodent biology, metabolism, pharmacokinetics, and disease in vivo. Choosing an appropriate anesthetic regimen as well as monitoring and supporting the animal's physiologic balance is key to obtaining images that truly represent the biologic process or disease state of interest. However, there are many challenges in rodent bioimaging such as limited animal access, small sample volumes, anesthetic complications, strain and gender variability, and the introduction of image artifacts. Because each imaging study presents unique challenges, a thorough understanding of the imaging modality used, the animal's health status, and the research data desired is required. This article addresses these issues along with other common laboratory animal clinical considerations such as biosecurity and radiation safety in in vivo rodent bioimaging.     

Mass Mortality of Adult Male Subantarctic Fur Seals: Are Alien Mice the Culprits?

Background

Mass mortalities of marine mammals due to infectious agents are increasingly reported. However, in contrast to previous die-offs, which were indiscriminate with respect to sex and age, here we report a land-based mass mortality of Subantarctic fur seals with apparent exclusivity to adult males. An infectious agent with a male-predilection is the most plausible explanation for this die-off. Although pathogens with gender-biased transmission and pathologies are unusual, rodents are known sources of male-biased infectious agents and the invasive Mus musculus house mouse, occurs in seal rookeries.

Methodology/ Principal Findings

Molecular screening for male-biased pathogens in this potential rodent reservoir host revealed the absence of Cardiovirus and Leptospirosis genomes in heart and kidney samples, respectively, but identified a novel Streptococcus species with 30% prevalence in mouse kidneys.

Conclusions/ Significance

Inter-species transmission through environmental contamination with this novel bacterium, whose congenerics display male-bias and have links to infirmity in seals and terrestrial mammals (including humans), highlights the need to further evaluate disease risks posed by alien invasive mice to native species, on this and other islands.