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Sophisticated gene circuits built by synthetic biology can enable bacteria to sense their environment and respond predictably. Biosensing bacteria can potentially probe the human gut microbiome to prevent, diagnose, or treat disease. To provide robust biocontainment for engineered bacteria, we devised a Cas9-assisted auxotrophic biocontainment system combining thymidine auxotrophy, an Engineered Riboregulator (ER) for controlled gene expression, and a CRISPR Device (CD). The CD prevents the engineered bacteria from acquiring thyA via horizontal gene transfer, which would disrupt the biocontainment system, and inhibits the spread of genetic elements by killing bacteria harboring the gene cassette. This system tunably controlled gene expression in the human gut commensal bacterium Bacteroides thetaiotaomicron, prevented escape from thymidine auxotrophy, and blocked transgene dissemination for at least 10 days. These capabilities were validated in vitro and in vivo. This biocontainment system exemplifies a powerful strategy for bringing genetically engineered microorganisms safely into biomedicine.

Background Biocontainment systems in healthcare settings are intended to ensure isolation of contagious or potentially contagious patients so as to protect healthcare providers and control infection spread. The need for safe and effective biocontainment systems has emerged during the COVID-19 outbreak. In particular, transport of potentially contagious patients poses technical, logistical and medical challenges that are to be addressed to protect population health and enable safe healthcare services delivery. Methods As novel biocontainment systems become available it is important to assess their operational effectiveness and safety and to define their organizational impact. We applied the Health Technology Assessment (HTA) framework to evaluate a novel rigid bio-containment system (Billy-Cab) integrated with the stretcher support, for ambulance transport of contagious patients. Results We applied the EUnetHTA core model to conduct an HTA report of Billy-Cab. A multi-disciplinary working group was established. We assessed available biocontainment under the nine dimensions of the EUnetHTA core model. We derived and pooled: 1) original data derived from clinical practice, 2) secondary data derived from systematic reviews of the literature, conducted according to the PRISMA guidelines, 3) original data derived from validated questionnaires, 4) economic original data. Conclusions Preliminary data allow to conclude selected biocontainment systems are effective in ensuring infection control during ambulance transport, with manageable organizational impact. In addition, the use biocontainment systems positively influence healthcare professionals' attitudes and performances. The HTA methodology offers substantial support to evaluate the introduction of innovative technologies during health emergencies and beyond. Key messages Selected biocontainment systems are effective in ensuring infection control during ambulance transport. HTA is to be applied to evaluate the introduction of innovative technologies during health emergencies and beyond.

Patient care and public health require timely, reliable laboratory testing. However, clinical laboratory professionals rarely know whether patient specimens contain infectious agents, making ensuring biosafety while performing testing procedures challenging. The importance of biosafety in clinical laboratories was highlighted during the 2014 Ebola outbreak, where concerns about biosafety resulted in delayed diagnoses and contributed to patient deaths. This review is a collaboration between subject matter experts from large and small laboratories and the federal government to evaluate the capability of clinical laboratories to manage biosafety risks and safely test patient specimens. We discuss the complexity of clinical laboratories, including anatomic pathology, and describe how applying current biosafety guidance may be difficult as these guidelines, largely based on practices in research laboratories, do not always correspond to the unique clinical laboratory environments and their specialized equipment and processes. We retrospectively describe the biosafety gaps and opportunities for improvement in the areas of risk assessment and management; automated and manual laboratory disciplines; specimen collection, processing, and storage; test utilization; equipment and instrumentation safety; disinfection practices; personal protective equipment; waste management; laboratory personnel training and competency assessment; accreditation processes; and ethical guidance. Also addressed are the unique biosafety challenges successfully handled by a Texas community hospital clinical laboratory that performed testing for patients with Ebola without a formal biocontainment unit. The gaps in knowledge and practices identified in previous and ongoing outbreaks demonstrate the need for collaborative, comprehensive solutions to improve clinical laboratory biosafety and to better combat future emerging infectious disease outbreaks.

Introduction: Positive pressure breathing air-fed protective suits from three vendors are commonly used in biosafety level 4 (BSL-4) laboratories: they are Dover Chemturion suits (ILC Dover, DE), Delta suits (Honeywell Safety, NC), and HVO suits (HVO-ISSI-Deutschland GmbH, Germany). To address the potential risk of infectious agents being introduced through the supplied breathing air stream, some suit manufacturers incorporate protective filters on the suits themselves. However, these integrated filters are not amenable to in situ testing for efficacy verification. We have been using external filters from Matheson USA on the positive pressure suits since our BSL-4 laboratories were commissioned two decades ago. As part of our BSL-4 protective suit management program, we test these filters before them being put into use, and annually thereafter. In the past few years, we have observed these filters failing at a higher rate, as high as two out of three of the new filters tested at one point. Objective: The purpose of this study was to procure personal protective filters from other sources and validate their efficacy long-term. Methods: Filters from Respirex, HVO, and Honeywell were validated for filter integrity and filter loading. Results: Respirex filters performed well during the initial testing and periodic testing thereafter. Regular testing of the Respirex filters has now been ongoing for 30 months with continued successful performance. Conclusion: Filters from Respirex are a suitable option to protect personnel wearing positive pressure suits in BSL-4 laboratories.

The tools of synthetic biology have enormous potential to help us uncover the fundamental mechanisms controlling development and metabolism in plants. However, their effective utilization typically requires transgenesis, which is plagued by long timescales and high costs. In this review we explore how transgenesis can be minimized by delivering foreign genetic material to plants with systemically mobile and persistent vectors based on RNA viruses. We examine the progress that has been made thus far and highlight the hurdles that need to be overcome and some potential strategies to do so. We conclude with a discussion of biocontainment mechanisms to ensure these vectors can be used safely as well as how these vectors might expand the accessibility of plant synthetic biology techniques. RNA vectors stand poised to revolutionize plant synthetic biology by making genetic manipulation of plants cheaper and easier to deploy, as well as by accelerating experimental timescales from years to weeks.

The need for increased testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus that causes coronavirus disease 2019 (COVID-19), has resulted in an increase of testing facilities outside of traditional clinical settings and sample handling by individuals without appropriate biohazard and biocontainment training. During the repatriation and quarantine of passengers from the Grand Princess cruise ship at a US military base, biocontainment of a potentially infectious sample from a passenger was compromised. This study describes the steps taken to contain the spill, decontaminate the area, and discusses the needs for adequate training in a biohazard response.

Studying potentially dangerous microbes in animal models of the disease they cause takes some extra considerations, as researchers and staff must perform their work under the careful restrictions of different biosafety level laboratories.

The aim of Australia’s Security Sensitive Biological Agents (SSBAs) Regulatory Scheme is to limit opportunities for acts of bioterrorism or criminal acts using harmful biological agents. The scheme is based on a two-tiered list of agents, of which many can cause disease in animals and thereby have the potential to significantly impact Australian agricultural industries and the economy. Laboratories handling these agents have clear responsibilities for their biocontainment, biosafety and biosecurity. Importantly, through research and ongoing improvements to diagnostic assays and techniques, these facilities also play an integral role in threat preparedness, mitigation, diagnosis and control of natural, accidental or deliberate outbreaks of disease.

Recent synthetic biology efforts have raised biosafety concerns for possible release of engineered cyanobacteria into natural environments. To address the issues, we developed a controllable metal ion induced biocontainment system for two model cyanobacteria. First, six ion-inducible promoters were respectively evaluated in both Synechococcus elongatus PCC 7942 and the fast-growing cyanobacterium Synechococcus elongatus UTEX 2973, leading to the identification of an iron ion-repressed promoter PisiAB with low leakage and a reduction-fold of 5.4 and 7.9, respectively. Second, holin-endolysin and nuclease NucA systems were introduced, the inhibition rate of which against two Synechococcus strains varied from 61% to 86.4%. Third, two toxin/antitoxin modules were identified capable of inducing programmed suicide in both Synechococcus strains after induction. Furthermore, an escape experiment was conducted and the results showed that the system was able to achieve an escape frequency below the detection limit of 10-9 after 3 days' duration, demonstrating the strategy integrating iron ion-inducible promoter PisiAB and that toxin/antitoxin modules could be a useful tool for cyanobacterium biocontainment.

Background More than 28 000 people were infected with Ebola virus during the 2014–2015 West African outbreak, resulting in more than 11 000 deaths. Better methods are needed to reduce the risk of self-contamination while doffing personal protective equipment (PPE) to prevent pathogen transmission. Methods A set of interventions based on previously identified failure modes was designed to mitigate the risk of self- contamination during PPE doffing. These interventions were tested in a randomized controlled trial of 48 participants with no prior experience doffing enhanced PPE. Contamination was simulated using a fluorescent tracer slurry and fluorescent polystyrene latex spheres (PLSs). Self-contamination of scrubs and skin was measured using ultraviolet light visualization and swabbing followed by microscopy, respectively. Doffing sessions were videotaped and reviewed to score standardized teamwork behaviors. Results Participants in the intervention group contaminated significantly fewer body sites than those in the control group (median [interquartile range], 6 [3–8] vs 11 [6–13], P = .002). The median contamination score was lower for the intervention group than the control group when measured by ultraviolet light visualization (23.15 vs 64.45, P = .004) and PLS swabbing (72.4 vs 144.8, P = .001). The mean teamwork score was greater in the intervention group (42.2 vs 27.5, P < .001). Conclusions An intervention package addressing the PPE doffing task, tools, environment, and teamwork skills significantly reduced the amount of self-contamination by study participants. These elements can be incorporated into PPE guidance and training to reduce the risk of pathogen transmission., An intervention package that addressed components of the task, tools, environment, and teamwork skills during personal protective equipment (PPE) doffing significantly reduced the amount of self-contamination by study participants. These elements can be incorporated into PPE guidance and training to prevent pathogen transmission.

The 2014 to 2016 Ebola outbreak response resulted in many lessons learned about biocontainment patient care, leading to enhanced domestic capabilities for highly infectious and hazardous communicable diseases. However, additional opportunities for improvement remain. The article identifies and describes key considerations and challenges for laboratory analysis, clinical management, transportation, and personnel management during the care of patients infected with Ebola or other special pathogens. Dedication to maintaining preparedness enables biocontainment patient care teams to perform at the highest levels of safety and confidence.

Bovine tuberculosis is a major economic problem for the cattle farmers of the UK, particularly the endemically infected are as of the south and west. Attempts at eradication have failed to effectively control the disease: of farms that suffer a breakdown and are then deemed clear of disease, more than half will suffer recurrence of the disease within 3 years, while a significant number of farms remain chronically infected. These herds provide a major reservoir of infection and robust, farm specific, disease control strategies are required to manage the disease from them, economically and sustainably. Novel technologies in surveillance, resilience and risk management are required to effectively control bovine tuberculosis in these endemically infected herds.

The 2014 West African Ebola virus disease (EVD) outbreak is the largest and deadliest EVD epidemic to date, resulting in fivefold more cases than all other outbreaks combined. This outbreak was particularly devastating to healthcare workers in West Africa and resulted in several EVD patients being medically evacuated for treatment in the U.S. and Europe. Governmental agencies provide recommendations for triaging and testing patients with EVD, however best laboratory practices are still unknown and are very resource dependent.

Introduction: Keeping a contamination free environment in the laboratory has commonly been achieved by one of two ways: a flame or a biosafety cabinet (BSC). However, it has been frequently observed that these two practices have been combined, where a heat source has been used within the BSC. As flames require flammable gasses and cause hot air to rise, it was hypothesized that these could lead to a loss of BSC containment, as BSCs rely on unidirectional downflow air. Objectives: The objective of this study was to determine whether BSCs can maintain containment when a heat source is operated within the work area. Methods: Several heat sources (Bunsen burner, High Heat Bunsen Burner, Spirit Lamp and Bacti-cinerator) were placed within two sizes of BSCs (4-foot and 6-foot), and smoke was used to visualize airflow disturbances, air cleanliness was measured by particle counting , and aerosol microbiological testing was conducted to ascertain containment. The risk of introducing a flammable gas into a BSC was also calculated. Results: Large flamed Bunsen burners were found to have the most detrimental effects on the ability of the BSC to maintain containment, especially in the center of the work area, while the smaller heat sources were more variable. Containment was completely lost in the 4-foot BSC, whereas the 6-foot BSC was capable of maintaining containment in only a few conditions. The BSC was also calculated to be able to maintain the required volume of flammable gas needed to operate the burners, not taking into consideration unintended leaks. Conclusions: Overall, it was determined that BSCs cannot operate safely and reliably while housing a heat source, as it could cause unexpected contamination of the work or the worker, or BSC ignition or explosion.

The UK Government has committed to a strategy of Bovine Tuberculosis eradication, with the stated aim of England becoming TB free by 2038. A review of the progress of the programme was conducted in 2018, (published in October 2018) which made specific and relatively radical changes to the current plan, including changes to governance, technical innovation and better farmer engagement in more effective biosecurity for their cattle herds. This personal opinion on the possible changes to the current programme outlines some radical improvements that could be made to the statutory controls, providing a practical and effective way of implementing the recommendations of the Strategy Review and re-engaging the industry in a robust and effective TB control and eradication strategy.

During the 2014-2016 Ebola virus epidemic, more than 500 health care workers (HCWs) died in spite of the use of personal protective equipment (PPE). The Johns Hopkins University Center for Bioengineering Innovation and Design (CBID) and Jhpiego, an international nongovernmental organization affiliate of Johns Hopkins, collaborated to create new PPE to improve the ease of the doffing process.HCWs in Liberia and a US biocontainment unit compared standard Médecins Sans Frontière PPE (PPE A) with the new PPE (PPE B). Participants wore each PPE ensemble while performing simulated patient care activities. Range of motion, time to doff, comfort, and perceived risk were measured.Overall, 100% of participants preferred PPE B over PPE A (P.0001); 98.1% of respondents would recommend PPE B for their home clinical unit (P.0001). There was a trend towards greater comfort in PPE B. HCWs at both sites felt more at risk in PPE A than PPE B (71.9% vs 25% in Liberia, P.0001; 100% vs 40% in the US biocontainment unit, P.0001).HCWs preferred a new PPE ensemble to Médecins Sans Frontière PPE for high-consequence pathogens. Further studies on the safety of this new PPE need to be conducted.

Worldwide, public health systems are continually challenged by emerging and re-emerging viruses. It is therefore important that high-containment labs coordinate and communicate globally to share their experiences and lessons to improve their capacity to respond to threats. The National Biosafety (Level 4) Laboratory in Wuhan, CAS, which is the first Level 4 biosafety laboratory (BSL-4) in China, has been certified recently and it is expected to play an important role in the prevention and control of highly infectious agents in future. Trained and experienced staff in such organizations is the most important factor contributing to safety and security. Therefore, it is imperative to develop a standard training program. Accordingly, in the present study, we developed an improved training program and assessment system based on policies and practices developed by BSL-3/4s in other countries. It included the following three components: (1) A flexible modularized theoretical training: This training comprised 14 modularized theoretical topics such that staffs with different backgrounds could take this theoretical training with different topic combinations according to their knowledge and skill levels; (2) A standardized practical training assessment: This comprehensive assessment, which could be used with biosafety laboratory staff before, during, and after training, included standard operation procedures to meet the special requirements of trainees with different scores; and (3) An applicable documentation system: A certification system was established to evaluate the ability of all staff working inside or outside the laboratory, implemented by a special committee. This certification was approved and authorized by the director of the laboratory and was classified into three grades with corresponding minimal requirements. Further, the present study examined the importance of and need for networking for training BSL-3/4 staff. The establishment of rigorous standards for training BSL-3/4 staff will instill confidence in the public, policy makers, and security officials. Additionally, the expanded international network of BSL-3/4s will continue to be operated safely and will pose no risk to scientific staff, local communities, surrounding environment, and host nations. The clarification and coordination of training standards will help develop a highly-qualified biocontainment workforce and will result in a series of programs that will enable scientists to develop measures to deal with existing threat agents and new diseases that emerge. Keywords: Level 3/4 biosafety laboratory, Biosafety training program, Networking

This article describes a large-scale scenario designed to test the capabilities of a US biocontainment unit to manage a pregnant woman infected with a high-consequence pathogen, and to care for a newborn following labor and spontaneous vaginal delivery. We created and executed a multidisciplinary functional exercise with simulation to test the ability of the Johns Hopkins Hospital biocontainment unit (BCU) to manage a pregnant patient in labor with an unknown respiratory illness and to deliver and stabilize her neonate. The BCU Exercise and Drill Committee established drill objectives and executed the exercise in partnership with the Johns Hopkins Simulation Center in accordance with Homeland Security and Exercise Program guidelines. Exercise objectives were assessed by after-action reporting and objective measurements to detect contamination, using a fluorescent marker to simulate biohazardous fluids that would be encountered in a typical labor scenario. The immediate objectives of the drill were accomplished, with stabilization of the mother and successful delivery and resuscitation of her newborn. There was no evidence of contamination when drill participants were inspected under ultraviolet light at the end of the exercise. Simulation optimizes teamwork, communication, and safety, which are integral to the multidisciplinary care of the maternal-fetal unit infected, or at risk of infection, with a high-consequence pathogen. Lessons learned from this drill regarding patient transportation, safety, and obstetric and neonatal considerations will inform future exercises and protocols and will assist other centers in preparing to care for pregnant patients under containment conditions.

The Ebola epidemic of 2014 demonstrated that outbreaks of high-consequence infectious diseases, even in remote parts of the world, can affect communities anywhere in the developed world and that every healthcare facility must be prepared to identify, isolate, and provide care for infected patients. The Nebraska Biocontainment Unit (NBU), located at Nebraska Medicine in Omaha, Nebraska, cared for 3 American citizens exposed in West Africa and confirmed with Ebola virus disease (EVD). Symptom monitoring of healthcare workers caring for these patients was implemented, which included twice daily contact to document the absence or presence of signs of fever or illness. This article describes the symptom monitoring experience of the NBU and local and state public health agencies. Based on lessons learned from that experience, we sought a more efficient solution to meet the needs of both the healthcare facility and public health authorities. REDCap, an open-source application used commonly by academic health centers, was used to develop an inexpensive symptom monitoring application that could reduce the burden of managing these activities, thus freeing up valuable time. Our pilot activities demonstrated that this novel use of REDCap holds promise for minimizing costs and resource demands associated with symptom monitoring while offering a more user-friendly experience for people being monitored and the officials managing the response.

Summary Synthetic biology offers new tools and capabilities of engineering cells with desired functions for example as new biosensing platforms leveraging engineered microbes. In the last two decades, bacterial cells have been programmed to sense and respond to various input cues for versatile purposes including environmental monitoring, disease diagnosis and adaptive biomanufacturing. Despite demonstrated proof‐of‐concept success in the laboratory, the real‐world applications of microbial sensors have been restricted due to certain technical and societal limitations. Yet, most limitations can be addressed by new technological developments in synthetic biology such as circuit design, biocontainment and machine learning. Here, we summarize the latest advances in synthetic biology and discuss how they could accelerate the development, enhance the performance and address the present limitations of microbial sensors to facilitate their use in the field. We view that programmable living sensors are promising sensing platforms to achieve sustainable, affordable and easy‐to‐use on‐site detection in diverse settings., Synthetic biology offers new tools and capabilities of engineering microbes with defined functions as new biosensing platforms for environmental monitoring, disease diagnosis and adaptive biomanufacturing. Here, we summarize the latest advances in synthetic biology and discuss how they could accelerate the development, enhance the performance and address the present limitations of microbial sensors to facilitate their use in the field. We view that programmable living sensors are promising sensing platforms to achieve sustainable, affordable, and easy‐to‐use on‐site detection in diverse settings.

Inadequate management of household and industrial wastes pose major challenges to human and environmental health. Advances in synthetic biology may help address these challenges by engineering biological systems to perform new functions such as biomanufacturing of high-value compounds from low-value waste streams and bioremediation of industrial pollutants. The current emphasis on microbial systems for biomanufacturing, which often require highly pre-processed inputs and sophisticated infrastructure, is not feasible for many waste streams. Concerns about transgene biocontainment have limited the release of engineered microbes or plants for bioremediation. Engineering animals may provide opportunities for utilizing various waste streams that are not suitable for microbial biomanufacturing while effective transgene biocontainment options should enable in situ bioremediation. Here, we engineer the model insect Drosophila melanogaster to express a functional laccase from the fungus Trametes trogii. Laccase expressing flies reduced concentrations of the endocrine disruptor bisphenol A by more than 50% when present in their growth media. A lyophilized powder made from engineered adult flies retained substantial enzymatic activity, degrading more than 90% of bisphenol A and the textile dye indigo carmine in aqueous solutions. Our results demonstrate that transgenic animals may be used to bioremediate environmental contaminants in vivo and serve as novel production platforms for industrial enzymes. These results support further development of insects, and possibly other animals, as bioproduction platforms and their potential use in bioremediation.

Molecular diagnosis and surveillance of pathogens such as SARS‐CoV‐2 depend on nucleic acid isolation. Pandemics at the scale of COVID‐19 can cause a global shortage of proprietary commercial reagents and BSL‐2 laboratories to safely perform testing. Therefore, alternative solutions are urgently needed to address these challenges. An open‐source method, magnetic‐nanoparticle‐aided viral RNA isolation from contagious samples (MAVRICS), built upon readily available reagents, and easily assembled in any basically equipped laboratory, is thus developed. The performance of MAVRICS is evaluated using validated pathogen detection assays and real‐world and contrived samples. Unlike conventional methods, MAVRICS works directly in samples inactivated in phenol‐chloroform (e.g., TRIzol), thus allowing infectious samples to be handled safely without biocontainment facilities. MAVRICS allows wastewater biomass immobilized on membranes to be directly inactivated and lysed in TRIzol followed by RNA extraction by magnetic nanoparticles, thereby greatly reducing biohazard risk and simplifying processing procedures. Using 39 COVID‐19 patient samples and two wastewater samples, it is shown that MAVRICS rivals commercial kits in detection of SARS‐CoV‐2, influenza viruses, and respiratory syncytial virus. Therefore, MAVRICS is safe, fast, and scalable. It is field‐deployable with minimal equipment requirements and could become an enabling technology for widespread testing and wastewater monitoring of diverse pathogens., One important bottleneck in the diagnosis and surveillance of COVID‐19 is the shortage of kits for RNA extraction. Magnetic‐nanoparticle‐aided viral RNA isolation from contagious samples (MAVRICS) is an open‐source, safe, fast, and scalable method for RNA extraction. MAVRICS rivals commercial kits but requires minimal materials, and thus could become an enabling technology for widespread community testing of diverse pathogens.

Background: The most critically ill patients with coronavirus disease 2019 (COVID-19) may require advanced support modalities, such as veno-venous extracorporeal membrane oxygenation (VV-ECMO). A systematic, methodical approach to a respiratory pandemic on a state and institutional level is critical. Methods: We conducted retrospective review of our institutional response to the COVID-19 pandemic, focusing on the creation of a dedicated airlock biocontainment unit (BCU) to treat patients with refractory COVID-19 acute respiratory distress syndrome (CARDS). Data were collected through conversations with staff on varying levels in the BCU, those leading the effort to make the BCU and hospital incident command system, email communications regarding logistic changes being implemented, and a review of COVID-19 patient census at our institution from March through June 2020. Results: Over 2100 patients were successfully admitted to system hospitals, 29% of these patients required critical care. The response to this respiratory pandemic augmented intensive care physician staffing, created a 70-member nursing team, and increased the extracorporeal membrane oxygenation (ECMO) capability by nearly 200%. During this time period, 40 COVID-19 patients on VV-ECMO were managed in the BCU. Challenges in an airlock unit included communication, scarcity of resources, double-bunking, and maintaining routine care. Conclusions: Preparing for a surge of critically ill patients during a pandemic can be a daunting task. The implementation of a coordinated, system-level approach can help with the allocation of resources as needed. Focusing on established strengths of hospitals within the system can guide triage based on individual patient needs. The management of ECMO patients is still a specialty care, and a systematic and hospital based approach requiring an ECMO team composed of multiple experienced individuals is paramount during a respiratory viral pandemic.

A survey was conducted to investigate potential differences in biosecurity and health management practices on Irish dairy farms that sent their heifers for contract-rearing (source dairy farms, SDF; n = 62) and those rearing their own heifers (control farms, CF; n = 50). Participating farmers were surveyed by postal questionnaire between September and November 2018. The overall response rate was 93%. Results show that structurally, SDF were larger, less fragmented, and more specialized than CF. Outsourcing of labor-intensive activities to external contractors was more common among SDF than CF, exposing them to potentially increased biosecurity risks associated with animal movements, use of shared equipment, and increased frequency of farm visitors. The majority of SDF sent heifers to a single-origin rearing facility (70%), with heifers most commonly arriving at the rearing unit between 2 and 4 mo (53%) and returning to the dairy farm between 18 and 21 mo of age (56%). Despite the increased biosecurity risk associated with contract-rearing, implementation of disease prevention measures was not superior on SDF compared with CF. For both farm types, there was scope for improvement to visitor biosecurity protocols, quarantine procedures, colostrum feeding practices, and hygiene of calving areas. This research provides an overview of the demographics and farm management practices implemented by dairy farmers engaged in contract-rearing of replacement heifers, and will serve to inform farmers, veterinary advisors, and policy makers.

With the recent upswing of infectious disease outbreaks (coronavirus, influenza, Ebola, etc), there is an ever-increasing need for biocontainment animal use protocols to better address the research of emerging diseases and to increase the health of both animals and humans. It is imperative that we as a research community ensure these protocols are conducted with the utmost scrutiny and regulatory compliance for the welfare of the animals as well as the health and safety concerns of the individual conducting these studies. Both the welfare of the animals and the health and safety of the research staff must be balanced with the integrity of the science being studied. Even prior to reviewing biocontainment protocols, the research stakeholders should have professional and collegial interactions across all levels of the proposed project. These stakeholders should include the attending veterinarian, the principal investigator, the sponsor, and any organic institutional health and safety assets (environmental health and safety, occupational health, biosafety personnel, medical personnel, facilities operations and maintenance, etc). At most institutions, these stakeholders are members of the Institutional Animal Care and Use Committee and may not possess the necessary tools to properly assess an Animal Biosafety Level 3 and 4 animal use protocol. It is the goal of this article to review some basic concepts of biocontainment, discuss critical communications and preapprovals, clinical observations, medical interventions and supportive care, scientific and study endpoints, euthanasia criteria, animal manipulations, documentation, training, emergency response and contingency plans, security, and decontamination and provide a scenario-based and informative thought-provoking process Institutional Animal Care and Use Committee members and veterinary staff may consider during Animal Biosafety Level 3 and 4 protocol review. These topics will enhance the ability of all stakeholders to balance the protection of the people with the integrity of the science and ultimately the welfare of the animal.

Background On 31 December 2019, the World Health Organization recognised clusters of pneumonia‐like cases due to a novel coronavirus disease (COVID‐19). COVID‐19 became a pandemic 71 days later. Aim To report the clinical and epidemiological features, laboratory data and outcomes of the first group of 11 returned travellers with COVID‐19 in Australia. Methods This is a retrospective, multi‐centre case series. All patients with confirmed COVID‐19 infection were admitted to tertiary referral hospitals in New South Wales, Queensland, Victoria and South Australia. Results The median age of our patient cohort was 42 years (IQR, 24–53 years) with six men and five women. Eight patients (72.7%) had returned from Wuhan, one from Shenzhen, one from Japan, and one from Europe. Possible human‐to‐human transmission from close family contacts in gatherings overseas occurred in two cases. Symptoms on admission were fever, cough and sore throat (n = 9, 81.8%). Co‐morbidities included hypertension (n = 3, 27.3%) and hypercholesterolaemia (n = 2, 18.2%). No patients developed severe acute respiratory distress nor required intensive care unit admission or mechanical ventilation. After a median hospital stay of 14.5 days (IQR, 6.75–21), all patients were discharged. Conclusions This is a historical record of the first COVID‐19 cases in Australia during the early biocontainment phase of the national response. These findings were invaluable for establishing early inpatient and outpatient COVID‐19 models of care and informing the management of COVID‐19 overtime as the outbreak evolved. Future research should extend this Australian case series to examine global epidemiological variation of this novel infection. This article is protected by copyright. All rights reserved.

Invasive bivalves can cause widespread ecological damage, but eradication has proved difficult. Identifying the pathways of dispersal is crucial to implementing more effective biocontainment measures. We examined the distribution of the highly invasive zebra mussel (Dreissena polymorpha) in Great Britain through Species Distribution Modelling to determine the drivers of distribution and generated suitability maps to predict future dispersal. Distance to boat ramps was the most important predictor of zebra mussel establishment, accounting for 27% of variation in occurrence. Probability of occurrence was highest within 3 km upstream of boat ramps, probably due to boating activity and the impounded waters typically associated with boat ramps. Our results highlight the need for implementing stringent control measures around boat ramps, and demonstrate the value of spatially modelling species distribution to create risk maps for targeting monitoring efforts at those locations most vulnerable to invasion.

The novel coronavirus SARS-CoV-2 is the causative agent of the acute respiratory disease COVID-19, which has become a global concern due to its rapid spread. Laboratory work with SARS-CoV-2 in a laboratory setting was rated to biosafety level 3 (BSL-3) biocontainment level. However, certain research applications in particular in molecular biology require incomplete denaturation of the proteins, which might cause safety issues handling contaminated samples. In this study, we evaluated lysis buffers that are commonly used in molecular biological laboratories for their ability to inactivate SARS-CoV-2. In addition, viral stability in cell culture media at 4 °C and on display glass and plastic surfaces used in laboratory environment was analyzed. Furthermore, we evaluated chemical and non-chemical inactivation methods including heat inactivation, UV-C light, addition of ethanol, acetone-methanol, and PFA, which might be used as a subsequent inactivation step in the case of insufficient inactivation. We infected susceptible Caco-2 and Vero cells with pre-treated SARS-CoV-2 and determined the tissue culture infection dose 50 (TCID50) using crystal violet staining and microscopy. In addition, lysates of infected cells and virus containing supernatant were subjected to RT-qPCR analysis. We have found that guanidine thiocyanate and most of the tested detergent containing lysis buffers were effective in inactivation of SARS-CoV-2, however, the M-PER lysis buffer containing a proprietary detergent failed to inactivate the virus. In conclusion, careful evaluation of the used inactivation methods is required especially for non-denaturing buffers. Additional inactivation steps might be necessary before removal of lysed viral samples from BSL-3. Supplementary Information The online version contains supplementary material available at 10.1007/s00430-021-00716-3.

Exploitation of plant growth promoting (PGP) rhizobacteria (PGPR) as crop inoculants could propel sustainable intensification of agriculture to feed our rapidly growing population. However, field performance of PGPR is typically inconsistent due to suboptimal rhizosphere colonisation and persistence in foreign soils, promiscuous host-specificity, and in some cases, the existence of undesirable genetic regulation that has evolved to repress PGP traits. While the genetics underlying these problems remain largely unresolved, molecular mechanisms of PGP have been elucidated in rigorous detail. Engineering and subsequent transfer of PGP traits into selected efficacious rhizobacterial isolates or entire bacterial rhizosphere communities now offers a powerful strategy to generate improved PGPR that are tailored for agricultural use. Through harnessing of synthetic plant-to-bacteria signalling, attempts are currently underway to establish exclusive coupling of plant-bacteria interactions in the field, which will be crucial to optimise efficacy and establish biocontainment of engineered PGPR. This review explores the many ecological and biotechnical facets of this research.

Biocontainment is an integral part of biomedical research that aims to protect the environment and human health by containing hazardous or invasive organisms in the laboratory. Containment systems often rely on elaborate genetic circuits; however, cells may escape containment by developing mutations that render the genetic circuits inviable or resistant to killing mechanisms. The aim of this thesis was to create a site-specific nuclease for biocontainment of plasmids in the mammalian gastrointestinal tract. LAGLIDADG homing endonucleases would be good candidate nucleases for a biocontainment system as they are resistant to mutations in their coding sequence and their target sequence in comparison to other nucleases, such as Cas9. Screening mutagenic libraries of the I-OnuI nuclease yielded a single variant that displays thermosensitive properties in vivo that may prove useful for biocontainment purposes. Using thermosensitive LAGLIDADG homing endonucleases for this purpose may prove to be a new, more robust approach to biocontainment.

Supplemental Digital Content is available in the text., We describe the process converting half of our 40-bed PICU into a negative-pressure biocontainment ICU dedicated to adult coronavirus disease 2019 patients within a 1,003-bed academic quaternary hospital. We outline the construction, logistics, supplies, provider education, staffing, and operations. We share lessons learned of working with a predominantly pediatric staff blended with adult expertise staff while maintaining elements of family-centered care typical of pediatric critical care medicine. Critically ill coronavirus disease 2019 adult patients may be cared for in a PICU and care may be augmented by implementing elements of holistic, family-centered PICU practice.

INTRODUCTION: An infectious disease outbreak like the current COVID-19 pandemic can lead to particularly high infection rates in areas where diagnostic laboratory support is limited. The deployment of mobile laboratories can help to detect pathogens, monitor the presence in a population, and inform public health authorities to take measures aimed at reducing pathogen spread. MATERIALS AND METHODS: Available layouts and operational descriptions of mobile laboratories were analyzed for their suitability for the envisioned purpose and to assure high standards of biosafety and biosecurity. Recent media coverage on creative solutions for the diagnostics of SARS-CoV-2 (drive-through test centers, self-swab, inverse gloveboxes to protect health care workers) from various countries were considered. RESULTS: A minimalistic and optimized design to construct a multifunctional laboratory on the chassis of a regular-sized box truck is proposed and can serve as a blueprint to rapidly develop additional diagnostic capacities. DISCUSSION: For acute health threats including the current COVID-19 outbreak, rapid diagnosis of infection is key to recommend measures aimed at preventing the spread of the pathogen. Laboratory layouts that are similar to the one proposed here are used in stationary setups, and mobile laboratories have been built on varying platforms (trailers, shipping containers, etc).

Following the publication of the italian Ministerial recommendations relating to dentistry in Phase 2 of the coViD-19 pandemic (focusing on operational protocols for all dental staff), we believe that the patient/dentist relationship should increasingly take into account the heightened fears and anxious thoughts of patients. this particularly regards patients who are about to undergo dental work. Moreover, dentists should also pay close attention to recent events, which have determined the new recommendations regarding SarS-coV-2 biocontainment. Furthermore, the authors of this paper consider it appropriate to make suggestions and develop interventionist techniques regarding the interface with the pa- tient starting from the initial consultation. the latter is invariably determinant in establishing clear communication of the Ministerial recommendations in encouraging a relaxed atmosphere with the patient. this interface is also a decisive factor in promoting patient empowerment, including specifying the time period envisaged for treatment in the new coViD-19 era in as calm a manner as possible. Such an approach will have a positive impact on the dentistry team.

Engineering at microscopic scales has an immense effect on the modern bioeconomy. Microbes contribute to such disparate markets as chemical manufacturing, fuel production, crop optimization, and pharmaceutical synthesis, to name a few. Due to new and emerging synthetic biology technologies, and the sophistication and control afforded by them, we are on the brink of deploying engineered microbes to not only enhance traditional applications but also to introduce these microbes to sectors, contexts, and formats not previously attempted. In microbially managed medicine, microbial engineering holds promise for increasing efficacy, improving tissue penetration, and sustaining treatment. In the environment, the most effective areas for deployment are in the management of crops and protection of ecosystems. However, caution is warranted before introducing engineered organisms to new environments where they may proliferate without control and could cause unforeseen effects. We summarize ideas and data that can inform identification and assessment of the risks that these tools present to ensure that realistic hazards are described and unrealistic ones do not hinder advancement. Further, because modes of containment are crucial complements to deployment, we describe the state of the art in microbial biocontainment strategies, current gaps, and how these gaps might be addressed through technological advances in synthetic engineering. Collectively, this work highlights engineered microbes as a foundational and expanding facet of the bioeconomy, projects their utility in upcoming deployments outside the laboratory, and identifies knowns and unknowns that will be necessary considerations and points of focus in this endeavor.

Background: Good laboratory practices in terms of biosecurity and biosafety are essential for the protection of laboratory personnel, the populations and the environment. If not implemented, the risk of pathogen spreading from the laboratory is conceivable. In addition, in current African geopolitical context, the reliable practices such as the protection, control and tracking of valuable biological material will prevent their loss, theft, uncontrolled access and potential misuse. WAHO, with the support of KfW is implementing trainings of trainers on biosecurity in the ECOWAS region in a project called PROALAB. PROALAB team realized that many partners were aiming at implementing in parallel quite similar non internationally certified programs in the ECOWAS region. Therefore, PROALAB decided to organize a meeting of key stakeholders in order to pool efforts, avoid duplication, and harmonize strategies and procedures. Objective: Develop a harmonized strategy and harmonized procedures for the training of health professionals in biosecurity/biosafety and biohazard risk management in ECOWAS member countries to pass internationally certified biosafety and biosecurity examinations. Methods: Programs of each of the institutions were presented and discussed. A regional program on biosecurity/biosafety training was elaborated during group working and a regional program of certification of trainers was discussed and validated. A work plan and timeline were developed through group working. Results: In the ECOWAS region, some countries are more advanced in biosecurity training. Training is generally organized by national biosecurity associations with the support of the IFBA or other partners (WHO, FMx, PHE, etc.). Côte d’Ivoire, Nigeria and Mali seem more advanced and could be solicited to organize training for less-advanced countries. The IFBA certification covers five domains: (1) Biorisk management, (2) biosecurity, (3) biosafety cabinet, (4) biological waste management and (5) biocontainment facility. The technical working group decided consensually that basic training for biosecurity managers in the laboratories should cover the first two domains. More advanced experts advising governments should be trained in the other domains progressively. Progressive training could be built on the 3 levels Nigerian/Ivoirian system of “basic-advanced-consolidated (or expert)” system. Existing module from WHO, FMx, IPD, IPCI and other ECOWAS countries should be adapted to match the international certification needs. Practical training should also be organized in the laboratory by the same institutions that will be allowed to deliver a practical certificate on behalf of WAHO. Conclusion: With the extension of laboratory activities in the context of strengthening the surveillance programs in the ECOWAS region, more expertise on biosecurity is needed. With the support of national biosecurity associations, WAHO could play a role in harmonizing reinforcing training on biosecurity and legislation.

Diagnosis and surveillance of emerging pathogens such as SARS-CoV-2 depend on nucleic acid isolation from clinical and environmental samples. Under normal circumstances, samples would be processed using commercial proprietary reagents in Biosafety 2 (BSL-2) or higher facilities. A pandemic at the scale of COVID-19 has caused a global shortage of proprietary reagents and BSL-2 laboratories to safely perform testing. Therefore, alternative solutions are urgently needed to address these challenges. We developed an open-source method calledMagnetic-nanoparticle-AidedViralRNAIsolation ofContagiousSamples (MAVRICS) that is built upon reagents that are either readily available or can be synthesized in any molecular biology laboratory with basic equipment. Unlike conventional methods, MAVRICS works directly in samples inactivated in acid guanidinium thiocyanate-phenol-chloroform (e.g., TRIzol), thus allowing infectious samples to be handled safely without biocontainment facilities. Using 36 COVID-19 patient samples, 2 wastewater samples and 1 human pathogens control sample, we showed that MAVRICS rivals commercial kits in validated diagnostic tests of SARS-CoV-2, influenza viruses, and respiratory syncytial virus. MAVRICS is scalable and thus could become an enabling technology for widespread community testing and wastewater monitoring in the current and future pandemics.

Transfusion of SARS-CoV-2 convalescent plasma is a promising treatment for severe COVID-19 cases, with success of the intervention based on neutralizing antibody content. Measurement by serological correlates without biocontainment needs, and an understanding of donor characteristics that may allow for targeting of more potent donors would greatly facilitate effective collection.

Research with captive wildlife in Animal Biosafety Level 2 (ABSL2) and 3 (ABSL3) facilities is becoming increasingly necessary as emerging and re-emerging diseases involving wildlife have increasing impacts on human, animal, and environmental health. Utilizing wildlife species in a research facility often requires outside the box thinking with specialized knowledge, practices, facilities, and equipment. The USGS National Wildlife Health Center (NWHC) houses an ABSL3 facility dedicated to understanding wildlife diseases and developing tools to mitigate their impacts on animal and human health. This review presents considerations for utilizing captive wildlife for infectious disease studies, including, husbandry, animal welfare, veterinary care, and biosafety. Examples are drawn from primary literature review and collective 40-year experience of the NWHC. Working with wildlife in ABSL2 and ABSL3 facilities differs from laboratory animals in that typical laboratory housing systems, husbandry practices, and biosafety practices are not designed for work with wildlife. This requires thoughtful adaptation of standard equipment and practices, invention of customized solutions and development of appropriate enrichment plans using the natural history of the species and the microbiological characteristics of introduced and native pathogens. Ultimately, this task requires critical risk assessment, understanding of the physical and psychological needs of diverse species, creativity, innovation, and flexibility. Finally, continual reassessment and improvement are imperative in this constantly changing specialty area of infectious disease and environmental hazard research.

Synthetic biology is a powerful tool to create therapeutics which can be rationally designed to enable unique and combinatorial functionalities. Here we utilize non-pathogenic E coli Nissle as a versatile platform for the development of a living biotherapeutic for the treatment of cancer. The engineered bacterial strain, referred to as SYNB1891, targets STING-activation to phagocytic antigen-presenting cells (APCs) in the tumor and activates complementary innate immune pathways. SYNB1891 treatment results in efficacious antitumor immunity with the formation of immunological memory in murine tumor models and robust activation of human APCs. SYNB1891 is designed to meet manufacturability and regulatory requirements with built in biocontainment features which do not compromise its efficacy. This work provides a roadmap for the development of future therapeutics and demonstrates the transformative potential of synthetic biology for the treatment of human disease when drug development criteria are incorporated into the design process for a living medicine., Synthetic biology can be used to create rationally designed living therapeutics. Here the authors engineer E. coli Nissle to target STING activation in antigen presenting cells for the treatment of solid tumors and demonstrate preclinical activity in murine models.