Dissection of the molecular circuitry controlling virulence in Francisella tularensis.

Francisella tularensis, the etiological agent of tularemia, is one of the most infectious bacteria known. Because of its extreme pathogenicity, F. tularensis is classified as a category A bioweapon by the US government. F. tularensis virulence stems from genes encoded on the Francisella pathogenicity island (FPI). An unusual set of Francisella regulators-the heteromeric macrophage growth locus protein A (MglA)-stringent starvation protein A (SspA) complex and the DNA-binding protein pathogenicity island gene regulator (PigR)-activates FPI transcription and thus is essential for virulence. Intriguingly, the second messenger, guanosine-tetraphosphate (ppGpp), which is produced during infection, is also involved in coordinating Francisella virulence; however, its role has been unclear. Here we identify MglA-SspA as a novel ppGpp-binding complex and describe structures of apo- and ppGpp-bound MglA-SspA. We demonstrate that MglA-SspA, which binds RNA polymerase (RNAP), also interacts with the C-terminal domain of PigR, thus anchoring the (MglA-SspA)-RNAP complex to the FPI promoter. Furthermore, we show that MglA-SspA must be bound to ppGpp to mediate high-affinity interactions with PigR. Thus, these studies unveil a novel pathway different from those described previously for regulation of transcription by ppGpp. The data also indicate that F. tularensis pathogenesis is controlled by a highly interconnected molecular circuitry in which the virulence machinery directly senses infection via a small molecule stress signal.

Poxviruses as Agents of Biological Warfare: The Importance of Ensuring Ethical Standards for Research with Viruses.

Historically, biological agents have been used to target various populations. One of the earliest examples could be the catastrophic effect of smallpox in Australia in the eighteenth century (as alleged by some historians). Modern biological techniques can be used to both create or provide protection against various agents of biological warfare. Any microorganism (viruses, bacteria, and fungi) or its toxins can be used as biological agents. Minnesota Department of Health has listed Smallpox (variola major) as a category A bioterrorism agent, even though it has been eradicated in 1980 through an extensive vaccination campaign. Category A agents are considered the highest risk to public health. Laboratory-associated outbreaks of poxviruses could cause unprecedented occupational hazards. Only two WHO-approved BSL-4 facilities in the United States and Russia are allowed to perform research on the variola virus. So, poxviruses present themselves as a classical case of a dual-use dilemma, since research with them can be used for both beneficial and harmful purposes. Although the importance of ethics in scientific research requires no further elaboration, ethical norms assume greater significance during experimentation with poxviruses. In this chapter, we will update the readers on the sensitive nature of conducting research with poxviruses, and how these viruses can be a source of potential biological weapons. Finally, specified ethical guidelines are explored to ensure safe research practices in virology.

Epidemiology of Pathogens Listed as Potential Bioterrorism Agents, the Netherlands, 2009‒2019.

We provide incidences (cases/10 million persons) in the Netherlands during 2009-2019 for pathogens listed as potential bioterrorism agents. We included pathogens from the highest categories of the European Medicines Agency or the US Centers for Disease Control and Prevention. Notifiable diseases and recently published data were used to calculate the average annual incidence. Coxiella burnetii had the highest incidence because of a Q fever epidemic during 2007-2010. Incidence then decreased to 10.8 cases/. Pathogens with an incidence >1 were Brucella spp. (2.5 cases), Francisella tularensis (1.3 cases), and Burkholderia pseudomallei (1.1 cases). Pathogens with an incidence <1 were hemorrhagic fever viruses (0.3 cases), Clostridium botulinum (0.2 cases), and Bacillus anthracis (0.1 cases). Variola major and Yersinia pestis were absent. The generally low incidences make it unlikely that ill-meaning persons can isolate these pathogens from natural sources in the Netherlands. However, the pathogens are stored in laboratories, underscoring the need for biosecurity measures.

Gain of Function and Loss of Control: Genetic Modification of Microbial Agents - Viruses "On Steroids".

Gain of Function refers to genetic modification to enhance certain properties of a biological agent. "Dual use research" refers to experiments which have a primary goal of benefitting humanity, but which could produce harm if misapplied. So, for example, a virus which was being genetically modified (GM) for altruistic reasons might become more transmissible or resistant to vaccines or antimicrobial medications. Such a GM virus has bioterrorism potential. The UN Biological Weapons Convention has not been universally approved and 10 States are not signatories to the Convention. The control of such experiments is variously controlled in certain jurisdictions but in Australia these experiments are well regulated through the Gene Technology Act 2000 (Cth), the National Health Security Act 2007 (Cth) and the Crimes (Biological Weapons) Act 1976 (Cth). The controls on such experiments in Europe and the United States are less precise. There are examples in the United States and Europe where the security provisions to contain microorganisms undergoing research including genetic modification have been breached. This threatens the health and safety of laboratory workers and the wider community.

Assessing dual use research of concern (DURC)-lessons learned from the United States government institutional DURC policy.

Life science research was analyzed for potential misuse in the 2004 report "Biotechnology Research in an Age of Bioterrorism". However, it was not until 2015 that the United States Government (USG) Institutional Dual Use Research of Concern (DURC) policy went into effect. Institutions receiving USG funding for life science research are required to scan their research portfolios for research involving one of 15 agents and subsequent 7 experimental effects described in the policy. In practice, this policy was implemented in a variety of ways with varying outcomes and lessons learned. First and foremost, reviewing research for potential DURC is a highly subjective process that differs depending on the risk tolerance, experience, and training of the individuals charged with reviewing research for an institution as well as the review process itself. The information being reviewed also lends to the subjectivity of the process, that is, the experimental data provided. It is difficult to determine whether research is potential DURC without experimental data. Any review process is hypothetical until there is data. Lastly, reviewers of the research should look beyond the research proposals, like how compounding existing research information can create new risks, potential use in other organisms or systems, or the creation of a roadmap that, for example, shows how to create a concerning organism or could be used in a pathogen.

Addressing Biodisaster X Threats With Artificial Intelligence and 6G Technologies: Literature Review and Critical Insights.

BACKGROUND: With advances in science and technology, biotechnology is becoming more accessible to people of all demographics. These advances inevitably hold the promise to improve personal and population well-being and welfare substantially. It is paradoxical that while greater access to biotechnology on a population level has many advantages, it may also increase the likelihood and frequency of biodisasters due to accidental or malicious use. Similar to "Disease X" (describing unknown naturally emerging pathogenic diseases with a pandemic potential), we term this unknown risk from biotechnologies "Biodisaster X." To date, no studies have examined the potential role of information technologies in preventing and mitigating Biodisaster X. OBJECTIVE: This study aimed to explore (1) what Biodisaster X might entail and (2) solutions that use artificial intelligence (AI) and emerging 6G technologies to help monitor and manage Biodisaster X threats. METHODS: A review of the literature on applying AI and 6G technologies for monitoring and managing biodisasters was conducted on PubMed, using articles published from database inception through to November 16, 2020. RESULTS: Our findings show that Biodisaster X has the potential to upend lives and livelihoods and destroy economies, essentially posing a looming risk for civilizations worldwide. To shed light on Biodisaster X threats, we detailed effective AI and 6G-enabled strategies, ranging from natural language processing to deep learning-based image analysis to address issues ranging from early Biodisaster X detection (eg, identification of suspicious behaviors), remote design and development of pharmaceuticals (eg, treatment development), and public health interventions (eg, reactive shelter-at-home mandate enforcement), as well as disaster recovery (eg, sentiment analysis of social media posts to shed light on the public's feelings and readiness for recovery building). CONCLUSIONS: Biodisaster X is a looming but avoidable catastrophe. Considering the potential human and economic consequences Biodisaster X could cause, actions that can effectively monitor and manage Biodisaster X threats must be taken promptly and proactively. Rather than solely depending on overstretched professional attention of health experts and government officials, it is perhaps more cost-effective and practical to deploy technology-based solutions to prevent and control Biodisaster X threats. This study discusses what Biodisaster X could entail and emphasizes the importance of monitoring and managing Biodisaster X threats by AI techniques and 6G technologies. Future studies could explore how the convergence of AI and 6G systems may further advance the preparedness for high-impact, less likely events beyond Biodisaster X.

Antivirals in medical biodefense.

The viruses historically implicated or currently considered as candidates for misuse in bioterrorist events are poxviruses, filoviruses, bunyaviruses, orthomyxoviruses, paramyxoviruses and a number of arboviruses causing encephalitis, including alpha- and flaviviruses. All these viruses are of concern for public health services when they occur in natural outbreaks or emerge in unvaccinated populations. Recent events and intelligence reports point to a growing risk of dangerous biological agents being used for nefarious purposes. Public health responses effective in natural outbreaks of infectious disease may not be sufficient to deal with the severe consequences of a deliberate release of such agents. One important aspect of countermeasures against viral biothreat agents are the antiviral treatment options available for use in post-exposure prophylaxis. These issues were adressed by the organizers of the 16th Medical Biodefense Conference, held in Munich in 2018, in a special session on the development of drugs to treat infections with viruses currently perceived as a threat to societies or associated with a potential for misuse as biothreat agents. This review will outline the state-of-the-art methods in antivirals research discussed and provide an overview of antiviral compounds in the pipeline that are already approved for use or still under development.

Biological agents of bioterrorism - preparedness is vital.

Bioterrorist threats and attacks are still an issue of concern in the world. Biological agents are divided into three categories. The highest priority agents classified in category A pose a massive risk to public health and national security. The society should be prepared for this risk. Health professionals in the Czech Republic should be aware of the diseases caused by category A agents, which are not common in the country. In this context, the project of the Ministry of the Interior of the Czech Republic “Decontamination of the injured persons” has been implemented at the Faculty of Health Sciences, Palacký University, Olomouc. The article provides an overview of selected serious infectious risks and information on the project the aim of which is to create certified methodical procedures and guidelines on situations related to bioterrorism.

Next-Generation Sequencing for Biodefense: Biothreat Detection, Forensics, and the Clinic.

BACKGROUND: Next-generation sequencing (NGS) is revolutionizing a variety of molecular biology fields including bioforensics, biosurveillance, and infectious disease diagnostics. For pathogen detection, the ability to sequence all nucleic acids in a sample allows near limitless multiplexability, free from a priori knowledge regarding an etiologic agent as is typically required for targeted molecular assays such as real-time PCR. Furthermore, sequencing capabilities can generate in depth genomic information, allowing detailed molecular epidemiological studies and bioforensics analysis, which is critical for source agent identification in a biothreat outbreak. However, lack of analytical specificity, inherent to NGS, presents challenges for regulated applications such as clinical diagnostics and molecular attribution. CONTENT: Here, we discuss NGS applications in the context of preparedness and biothreat readiness. Specifically, we investigate current and future applications of NGS technologies to affect the fields of biosurveillance, bioforensics, and clinical diagnostics with specific focus on biodefense. SUMMARY: Overall, there are many advantages to the implementation of NGS for preparedness and readiness against biowarfare agents, from forensics to diagnostics. However, appropriate caveats must be associated with any technology. This includes NGS. While NGS is not the panacea replacing all molecular techniques, it will greatly enhance the ability to detect, characterize, and diagnose biowarfare agents, thus providing an excellent addition to the biodefense toolbox of biosurveillance, bioforensics, and biothreat diagnosis.

Information Hazards in Biotechnology.

With the advance of biotechnology, biological information, rather than biological materials, is increasingly the object of principal security concern. We argue that both in theory and in practice, existing security approaches in biology are poorly suited to manage hazardous biological information, and use the cases of Mousepox, H5N1 gain of function, and Botulinum toxin H to highlight these ongoing challenges. We suggest that mitigation of these hazards can be improved if one can: (1) anticipate hazard potential before scientific work is performed; (2) consider how much the new information would likely help both good and bad actors; and (3) aim to disclose information in the manner that maximally disadvantages bad actors versus good ones.

Smallpox Readiness: Modern Strategies Against an Ancient Disease.

This Viewpoint highlights the potential for unintentional or deliberate release of variola virus (smallpox), discusses current medical countermeasures for smallpox, and calls for greater flexibility from the US and its partners in developing safe, reliable, affordable, and equitable countermeasures.

Engineering H5N1 avian influenza viruses to study human adaptation.

Two studies of H5N1 avian influenza viruses that had been genetically engineered to render them transmissible between ferrets have proved highly controversial. Divergent opinions exist about the importance of these studies of influenza transmission and about potential 'dual use' research implications. No consensus has developed yet about how to balance these concerns. After not recommending immediate full publication of earlier, less complete versions of the studies, the United States National Science Advisory Board for Biosecurity subsequently recommended full publication of more complete manuscripts; however, controversy about this and similar research remains.

Experimental adaptation of an influenza H5 HA confers respiratory droplet transmission to a reassortant H5 HA/H1N1 virus in ferrets.

Highly pathogenic avian H5N1 influenza A viruses occasionally infect humans, but currently do not transmit efficiently among humans. The viral haemagglutinin (HA) protein is a known host-range determinant as it mediates virus binding to host-specific cellular receptors. Here we assess the molecular changes in HA that would allow a virus possessing subtype H5 HA to be transmissible among mammals. We identified a reassortant H5 HA/H1N1 virus-comprising H5 HA (from an H5N1 virus) with four mutations and the remaining seven gene segments from a 2009 pandemic H1N1 virus-that was capable of droplet transmission in a ferret model. The transmissible H5 reassortant virus preferentially recognized human-type receptors, replicated efficiently in ferrets, caused lung lesions and weight loss, but was not highly pathogenic and did not cause mortality. These results indicate that H5 HA can convert to an HA that supports efficient viral transmission in mammals; however, we do not know whether the four mutations in the H5 HA identified here would render a wholly avian H5N1 virus transmissible. The genetic origin of the remaining seven viral gene segments may also critically contribute to transmissibility in mammals. Nevertheless, as H5N1 viruses continue to evolve and infect humans, receptor-binding variants of H5N1 viruses with pandemic potential, including avian-human reassortant viruses as tested here, may emerge. Our findings emphasize the need to prepare for potential pandemics caused by influenza viruses possessing H5 HA, and will help individuals conducting surveillance in regions with circulating H5N1 viruses to recognize key residues that predict the pandemic potential of isolates, which will inform the development, production and distribution of effective countermeasures.

Microorganisms: Good or Evil, MIRRI Provides Biosecurity Awareness.

The life-science community is a key stakeholder in the effort to ensure that the advances in biotechnology are not misused. Unfortunately, to date, the engagement of life scientists with issues of biosecurity has been limited. Microorganisms have been harnessed for the benefit of humankind but in the wrong hands could be used in direct or indirect acts against humans, livestock, crops, food, water infrastructure and other economically valuable entities. The Microbial Resources Research Infrastructure in its preparatory phase has addressed the topic implementing a code of conduct as part of its programme of prevention of malicious use and continues to work with the international community to raise awareness of best practice to avoid misuse of microorganisms. Biosecurity has become a major concern for several countries creating numerous activities to put in place counter measures, risk assessment, legislation and emergency response. The goal is to implement measures to protect us against malicious use of microorganisms, their products, information and technology transfer. Through this paper, we wish to discuss some of the activities that are underway, mention key educational tools and provide scientists with information on addressing biosecurity issues.

United Nations Secretary-General's Mechanism.

The credibility and legitimacy of the United Nations Secretary-General's Mechanism (UNSGM) for investigating the alleged use of biological weapons relies in part on qualified experts trained to undertake fact-finding missions using an empirical scientific approach. As observed in the 2013 investigation into the alleged use of chemical weapons in the Syrian Arab Republic, this mechanism can be an effective tool in the verification of and as a deterrent against the use of chemical and biological weapons. The World Organisation for Animal Health (OIE) has a memorandum of understanding with the United Nations Office for Disarmament Affairs, supporting the UNSGM and providing experts for its roster for missions. As the majority of biological agents are of animal origin, the expertise of the OIE in this area, and its involvement in the UNSGM, are of paramount importance. Since 2014, experts from the OIE have participated in relevant training and are available for deployment to investigate any alleged use of a biological weapon anywhere in the world if the UNSGM were triggered.

La crédibilité et la légitimité du Mécanisme du Secrétaire général des Nations Unies visant à enquêter rapidement sur les allégations relatives à l'emploi possible d'armes biologiques reposent en partie sur le travail d'experts qualifiés et formés aux enquêtes d'établissement des faits, qui font appel à une méthode scientifique empirique. Comme cela a été constaté lors de l'enquête de 2013 concernant l'emploi allégué d'armes chimiques en République arabe syrienne, ce mécanisme peut constituer un outil efficace de vérification et de dissuasion en matière d'utilisation d'armes chimiques et biologiques. L'Organisation mondiale de la santé animale (OIE) a conclu un Mémorandum d'accord avec le Bureau des affaires de désarmement des Nations Unies, par lequel l'OIE s'engage à soutenir le Mécanisme du Secrétaire général et à mettre à disposition des experts pour participer aux missions. Les agents biologiques étant majoritairement d'origine animale, les compétences de l'OIE dans ce domaine et sa contribution au Mécanisme du Secrétaire général des Nations Unies sont d'une importance capitale. Depuis 2014, les experts de l'OIE ont participé aux formations requises ; en cas d'activation du mécanisme, ils sont donc opérationnels pour prendre part aux missions d'enquête sur l'emploi allégué d'armes biologiques partout dans le monde.

La credibilidad y legitimidad del Mecanismo del Secretario General de las Naciones Unidas para la Investigación del Presunto Empleo de Armas Químicas, Biológicas o Toxínicas reposa en parte en la participación de expertos cualificados y formados para llevar a cabo misiones de esclarecimiento de los hechos empleando métodos científicos experimentales. Como dejó patente la investigación sobre el presunto uso de armas químicas en la República Árabe Siria realizada en 2013, este mecanismo puede constituir una eficaz herramienta de verificación y un factor disuasorio del uso de armas químicas o biológicas. La Organización Mundial de Sanidad Animal (OIE) tiene suscrito con la Oficina de Asuntos de Desarme de las Naciones Unidas un memorando de entendimiento en virtud del cual presta apoyo al Mecanismo del Secretario General y proporciona expertos que figuran en la lista de especialistas llamados a participar en sus misiones. Toda vez que la mayoría de los agentes biológicos son de origen animal, la competencia técnica de la OIE en este ámbito, así como su participación en el Mecanismo, revisten una importancia capital. Desde 2014, expertos de la OIE han participado en actividades de formación sobre el tema y están disponibles para desplegarse en cualquier lugar del mundo e investigar todo caso de presunta utilización de un arma biológica en cuanto se active el Mecanismo del Secretario General.

Links between law enforcement and veterinary animal health: a case study of the United Kingdom and the United States of America.

The anthrax attacks carried out in the United States of America in the latter part of 2001 served as a clarion call to most law enforcement agencies among developed countries as, until this time, they had not recognised the threat posed by the criminal use of pathogens and/or toxins. Law enforcement agencies include the local and federal police, Customs, Immigration and any other agencies that perform a law-enforcing role. That a criminal or terrorist group could commit such an act was considered nearly inconceivable, even though biological weapons had been used against humans and animals in warfare many times over the last several hundred years. Similarly, it is without doubt that the terrorist threat to all societies changed after the events of 11 September 2001 during which the clear intention had been to cause as many fatalities and casualties as possible. As a result, the biological threat to human health became a focus for many governments.

Les attaques à la fièvre charbonneuse perpétrés aux États-Unis d'Amérique à la fin de l'année 2001 ont lancé un signal d'alerte à la plupart des agences chargées de l'application des lois des pays développés, qui n'avaient jusqu'alors pas pris conscience des menaces posées par l'utilisation criminelle des agents pathogènes et/ou des toxines. Les agences chargées de l'application des lois sont notamment les forces de police locales et fédérales, les services des douanes, les services d'immigration et toute autre agence gouvernementale chargée du maintien de l'ordre et de la justice. Il était pratiquement inconcevable qu'un groupe criminel ou terroriste puisse commettre un tel acte, même si des armes biologiques avaient déjà été utilisées contre les populations humaines et animales au cours de certains conflits armés des siècles précédents. De même, il va sans dire que la menace terroriste pesant sur les sociétés dans leur ensemble a pris un nouveau visage après les événements du 11 septembre 2001, dont le but était clairement de faire le plus de victimes possible et de tuer le plus grand nombre de personnes possible. En conséquence, la menace biologique pesant sur la santé publique est devenue un domaine central d'attention pour nombre de gouvernements.

Los ataques con la bacteria del carbunco perpetrados en los Estados Unidos de América a finales de 2001 marcaron un antes y un después para la mayoría de los cuerpos de seguridad de los países desarrollados, que hasta la fecha no habían advertido la amenaza que planteaba el uso de patógenos y/o toxinas con fines criminales. Forman parte de los cuerpos de seguridad las fuerzas de policía local y federal, los cuerpos de aduanas, inmigración y demás instituciones cuya función consiste en velar por el cumplimiento de la ley. Que un grupo delictivo o terrorista pudiera cometer semejante acto era algo prácticamente inconcebible, aun cuando en los últimos siglos hayan menudeado los ejemplos de utilización de armas biológicas contra la población humana o animal como instrumento de guerra. Análogamente, no cabe duda alguna de que la amenaza terrorista que planea sobre todas las sociedades cambió a partir de los acontecimientos del 11 de septiembre de 2001, en los que la intención era claramente de causar tantos muertos y heridos como fuera posible. A resultas de aquello, las amenazas biológicas para la salud humana pasaron a ser una de las preocupaciones cardinales de muchos gobiernos.

Synthetic Biology, Genome Editing, and the Risk of Bioterrorism.

The SynBioSecurity argument says that synthetic biology introduces new risks of intentional misuse of synthetic pathogens and that, therefore, there is a need for extra regulations and oversight. This paper provides an analysis of the argument, sets forth a new version of it, and identifies three developments that raise biosecurity risks compared to the situation earlier. The developments include (1) a spread of the required know-how, (2) improved availability of the techniques, instruments and biological parts, and (3) new technical possibilities such as "resurrecting" disappeared pathogens. It is first shown that the general argument from SynBioSecurity needs to be qualified and that many improvements to biosecurity have already been implemented, most notably in the United States. Second, I suggest a new strain of the argument: the situation that most branches of synthetic biology fall under the gene technology regulation in the European Union and that this regulation in its current form does not adequately address SynBioSecurity risks together provide a weighty reason to review and possibly refine the legislation as well as the supervisory practices. Ethically speaking, the rise in the relative risk of bioterrorism brings to the fore new extrinsic issues.