Applied Biosafety最新文献

Introduction: The Biosafety Research Road Map reviewed the scientific literature on a viral respiratory pathogen, avian influenza virus, and a bacterial respiratory pathogen, Mycobacterium tuberculosis. This project aims at identifying gaps in the data required to conduct evidence-based biorisk assessments, as described in Blacksell et al. One significant gap is the need for definitive data on M. tuberculosis sample aerosolization to guide the selection of engineering controls for diagnostic procedures.

Methods: The literature search focused on five areas: routes of inoculation/modes of transmission, infectious dose, laboratory-acquired infections, containment releases, and disinfection and decontamination methods.

Results: The available data regarding biosafety knowledge gaps and existing evidence have been collated and presented in Tables 1 and 2. The guidance sources on the appropriate use of biosafety cabinets for specific procedures with M. tuberculosis require clarification. Detecting vulnerabilities in the biorisk assessment for respiratory pathogens is essential to improve and develop laboratory biosafety in local and national systems.

Introduction: Decontamination of farms affected by bovine tuberculosis could be very challenging during outbreaks occurring in the winter with freezing temperatures. Steam treatment has been of practical interest, but information is needed on whether such treatment is able to inactivate the causative agent, Mycobacterium bovis. This study was to evaluate the use of pressurized steam for inactivation of Mycobacterium terrae, a surrogate for M. bovis on various surfaces.

Methods: Carrier disks made of steel, wood, or rubber were inoculated with 6.32 ± 0.38 log₁₀ M. terrae. While being held at background temperatures of -20°C, 4°C, or 21°C, these carrier disks were treated with pressurized steam (120°C ± 5°C) for 5, 10, 15, or 20 s. Reduction in colony forming units of M. terrae and temperatures on the top and bottom surfaces of the disks were determined.

Results: Complete inactivation of 6 log₁₀ M. terrae on steel and wood disks was achieved by 10 s of steam treatment at all three background temperatures. In comparison, 20 s of steam treatment was needed for the complete inactivation of mycobacteria on rubber disks. Corresponding to the longer treatment time required for mycobacterial inactivation, temperatures on the bottom surface of the rubber disks rose substantially slower than those of the steel and wood disks at all three background temperatures.

Conclusion: The results suggested that treatment with pressurized steam has potential for efficient and effective disinfection of surfaces contaminated by mycobacteria at or below freezing temperatures in winter.

Introduction: This article provides a strategy by which a manufacturing process with a Biosafety Level 2 (BL2) designation can be downgraded to Biosafety Level 1 (BL1). The principles of the downgrading process are based on the robust contamination controls in clinical and commercial manufacturing, which typically are not part of Research and Development processes. These strict requirements along with the application of current Good Manufacturing Practice (cGMP) principles provide a framework by which processes can be suitably managed and controlled to mitigate biohazard risk, specifically for cell lines that may be contaminated with human pathogenic viral agents.

Purpose: We demonstrate how a risk assessment guide was used to define the risk profile of a theoretical process with a human cell line intended for clinical/commercial application. Based on the risk assessment, key BL2 elements were identified as suitable for downgrading, including facility containment controls, emergency spill response plans, and storage and shipping requirements. For various reasons, some aspects of the systems were deemed unsuitable for downgrading due to the severity of the control risk and, therefore, remained at BL2.

Summary and conclusions: We have used an established risk assessment guide to show how cGMP compliments and augments biosafety containment. We provide justification for downgrading from BL2 to BL1 for clinical and commercial cell and gene therapy manufacturing with human cell lines.

Introduction: Snap-cap microcentrifuge tubes are ubiquitous in biological laboratories. However, limited data are available on how frequently splashes occur when opening them. These data would be valuable for biorisk management in the laboratory.

Methods: The frequency of splashes from opening snap-cap tubes using four different methods was tested. The splash frequency for each method was measured on the benchtop surface and on the experimenter's gloves and smock, using a Glo Germ solution as a tracer.

Results: Splashes occurred very frequently when opening microcentrifuge snap-cap tubes, no matter which method was used to open the tube. The highest rate of splashes on all surfaces was observed with the one-handed (OH) opening method compared with two-handed methods. Across all methods, the highest rate of splashes was observed on the opener's gloves (70-97%) compared with the benchtop (2-40%) or the body of the researcher (0-7%).

Conclusions: All tube opening methods we studied frequently caused splashes, with the OH method being the most error-prone but no two-handed method being clearly superior to any other. In addition to posing an exposure risk to laboratory personnel, experimental repeatability may be affected due to loss of volume when using snap-cap tubes. The rate of splashes underscores the importance of secondary containment, personal protective equipment, and good protocols for decontamination. When working with especially hazardous materials, alternatives to snap-cap tubes (such as screw cap tubes) should be strongly considered. Future studies can examine other methods of opening snap-cap tubes to determine whether a truly safe method exists.

Introduction: Shigella bacteria cause shigellosis, a gastrointestinal infection most often acquired from contaminated food or water.

Methods: In this review, the general characteristics of Shigella bacteria are described, cases of laboratory-acquired infections (LAIs) are discussed, and evidence gaps in current biosafety practices are identified.

Results: LAIs are undoubtedly under-reported. Owing to the low infectious dose, rigorous biosafety level 2 practices are required to prevent LAIs resulting from sample manipulation or contact with infected surfaces.

Conclusions: It is recommended that, before laboratory work with Shigella, an evidence-based risk assessment be conducted. Particular emphasis should be placed on personal protective equipment, handwashing, and containment practices for procedures that generate aerosols or droplets.

Introduction: Viral RNA replicons are self-amplifying RNA molecules generated by deleting genetic information of one or multiple structural proteins of wild-type viruses. Remaining viral RNA is used as such (naked replicon) or packaged into a viral replicon particle (VRP), whereby missing genes or proteins are supplied via production cells. Since replicons mostly originate from pathogenic wild-type viruses, careful risk consideration is crucial.

Methods: A literature review was performed compiling information on potential biosafety risks of replicons originating from positive- and negative-sense single-stranded RNA viruses (except retroviruses).

Results: For naked replicons, risk considerations included genome integration, persistence in host cells, generation of virus-like vesicles, and off-target effects. For VRP, the main risk consideration was formation of primary replication competent virus (RCV) as a result of recombination or complementation. To limit the risks, mostly measures aiming at reducing the likelihood of RCV formation have been described. Also, modifying viral proteins in such a way that they do not exhibit hazardous characteristics in the unlikely event of RCV formation has been reported.

Discussion and conclusion: Despite multiple approaches developed to reduce the likelihood of RCV formation, scientific uncertainty remains on the actual contribution of the measures and on limitations to test their effectiveness. In contrast, even though effectiveness of each individual measure is unclear, using multiple measures on different aspects of the system may create a solid barrier. Risk considerations identified in the current study can also be used to support risk group assignment of replicon constructs based on a purely synthetic design.

Introduction: The SARS-CoV-2 virus emerged as a novel virus and is the causative agent of the COVID-19 pandemic. It spreads readily human-to-human through droplets and aerosols. The Biosafety Research Roadmap aims to support the application of laboratory biological risk management by providing an evidence base for biosafety measures. This involves assessing the current biorisk management evidence base, identifying research and capability gaps, and providing recommendations on how an evidence-based approach can support biosafety and biosecurity, including in low-resource settings.

Methods: A literature search was conducted to identify potential gaps in biosafety and focused on five main sections, including the route of inoculation/modes of transmission, infectious dose, laboratory-acquired infections, containment releases, and disinfection and decontamination strategies.

Results: There are many knowledge gaps related to biosafety and biosecurity due to the SARS-CoV-2 virus's novelty, including infectious dose between variants, personal protective equipment for personnel handling samples while performing rapid diagnostic tests, and laboratory-acquired infections. Detecting vulnerabilities in the biorisk assessment for each agent is essential to contribute to the improvement and development of laboratory biosafety in local and national systems.