Using thermodynamic equilibrium models to predict the effect of antiviral agents on infectivity: Theoretical application to SARS-CoV-2 and other viruses.

Thermodynamic equilibrium models predict the infectivity of novel and emerging viruses using molecular data including the binding affinity of the virus to the host cell (as represented by the association constant Ka_virus_T) and the probability, pvirogenesis, of the virus replicating after entry to the cell. Here those models are adapted based on the principles of ligand binding to macromolecules to assess the effect on virus infectivity of inhibitor molecules which target specific proteins of the virus. Three types of inhibitor are considered using the thermodynamic equilibrium model for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection of the human lung with parameters for the strength and nature of the interaction between the target virus protein and the inhibitor molecule. The first is competitive inhibition of the SARS-CoV-2 spike glycoprotein (SGP) trimer binding to its human angiotensin converting enzyme 2 (ACE2) receptor by unfractionated heparin (UFH). Using a novel approach presented here, a value of Ka_virus_T = 3.53 × 1017 M-1 is calculated for SARS-CoV-2 from the IC50 for inhibition by UFH of SARS-CoV-2 plaque formation in cell culture together with the dissociation constant KVI of 0.73 × 10-10 M reported for heparin binding to SARS-CoV-2 SGP trimer. Such a high Ka_virus_T limits the effectiveness of competitive inhibitors such as UFH. The second is the attachment of a nanoparticle such as a zinc oxide tetrapod (ZnOT) to the virus shell as for herpes simplex virus (HSV). The increase in molecular weight through ZnOT attachment is predicted to decrease Ka_virus_T by orders of magnitude by making the entropy change (ΔSa_immob) on immobilisation of the ZnOT:virus complex on cell binding more negative than for the virus alone. According to the model, ZnOT acts synergistically with UFH at the IC50 of 33 µg/cm3 which together decrease viral infectivity by 61,000-fold compared to the two-fold and three-fold decreases predicted for UFH alone at the IC50 and for ZnOT alone respectively. According to the model here, UFH alone at its peak deliverable dose to the lung of 1,000 µg/cm3 only decreases infectivity by 31-fold. Practicable approaches to target and decrease ΔSa_immob for respiratory viruses should therefore be considered. The combination of decreasing ΔSa_immob together with blocking the interaction of virus surface protein with its host cell receptor may achieve synergistic effects for faecal-oral viruses and HSV. The third is reversible noncompetitive inhibition of the viral main protease (Mpro) for which the decrease in pvirogenesis is assumed to be proportional to the decrease in enzyme activity as predicted by enzyme kinetic equations for a given concentration of inhibitor which binds to Mpro with dissociation constant Ki. Virologists reporting viral inhibition studies are urged to report the concentration of cells in the cell culture experiment as this is a key parameter in estimating Ka_virus_T here.

Predicting spread and effective control measures for African swine fever-Should we blame the boars?

An ongoing, continually spreading, outbreak of African swine fever (ASF), following its identification in Georgia in 2007, has resulted in 17 European and 12 Asian countries reporting cases by April 2020, with cases occurring in both wild boar and domestic pigs. Curtailing further spread of ASF requires understanding of the transmission pathways of the disease. ASF is self-sustaining in the wild boar population, and they have been implicated as one of the main drivers of transmission within Europe. We developed a spatially explicit model to estimate the risk of infection with ASF in wild boar and pigs due to natural movement of wild boar that is applicable across the whole of Europe. We demonstrate the model by using it to predict the probability that early cases of ASF in Poland were caused by wild boar dispersion. The risk of infection in 2015 is computed due to wild boar cases in Poland in 2014, compared against reported cases in 2015, and then the procedure is repeated for 2015-2016. We find that long- and medium-distance spread of ASF (i.e. >30 km) is unlikely to have occurred due to wild boar dispersal, due in part to the generally short distances wild boar will travel (<20 km on average). We also predict the relative success of different control strategies in 2015, if they were implemented in 2014. Results suggest that hunting of wild boar reduces the number of new cases, but a larger region is at risk of ASF compared with no control measure. Alternatively, introducing wild boar-proof fencing reduces the size of the region at risk in 2015, but not the total number of cases. Overall, our model suggests wild boar movement is only responsible for local transmission of disease; thus, other pathways are more dominant in medium- and long-distance spread of the disease.

Thermodynamic equilibrium dose-response models for MERS-CoV infection reveal a potential protective role of human lung mucus but not for SARS-CoV-2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and Middle East respiratory syndrome coronavirus (MERS-CoV) infect the human respiratory tract. A prototype thermodynamic equilibrium model is presented here for the probability of the virions getting through the mucus barrier and infecting epithelial cells based on the binding affinity (Kmucin) of the virions to mucin molecules in the mucus and parameters for binding and infection of the epithelial cell. Both MERS-CoV and SARS-CoV-2 bind strongly to their cellular receptors, DDP4 and ACE2, respectively, and infect very efficiently both bronchus and lung ex vivo cell cultures which are not protected by a mucus barrier. According to the model, mucin binding could reduce the infectivity for MERS-CoV compared to SARS-CoV-2 by at least 100-fold depending on the magnitude of Kmucin. Specifically Kmucin values up to 106 M-1 have little protective effect and thus the mucus barrier would not remove SARS-CoV-2 which does not bind to sialic acids (SA) and hence would have a very low Kmucin. Depending on the viability of individual virions, the ID50 for SARS-CoV-2 is estimated to be ~500 virions (viral RNA genomic copies) representing 1 to 2 pfu. In contrast MERS-CoV binds both SA and human mucin and a Kmucin of 5 × 109 M-1 as reported for lectins would mop up 99.83% of the virus according to the model with the ID50 for MERS-CoV estimated to be ~295,000 virions (viral RNA genomic copies) representing 819 pfu. This could in part explain why MERS-CoV is poorly transmitted from human to human compared to SARS-CoV-2. Some coronaviruses use an esterase to escape the mucin, although MERS-CoV does not. Instead, it is shown here that "clustering" of virions into single aerosol particles as recently reported for rotavirus in extracellular vesicles could provide a co-operative mechanism whereby MERS-CoV could theoretically overcome the mucin barrier locally and a small proportion of 10 µm diameter aerosol particles could contain ~70 virions based on reported maximum levels in saliva. Although recent evidence suggests SARS-CoV-2 initiates infection in the nasal epithelium, the thermodynamic equilibrium models presented here could complement published approaches for modelling the physical entry of pathogens to the lung based on the fate and transport of the pathogen particles (as for anthrax spores) to develop a dose-response model for aerosol exposure to respiratory viruses. This would enable the infectivity through aerosols to be defined based on molecular parameters as well as physical parameters. The role of the spike proteins of MERS-CoV and SARS-CoV-2 binding to SA and heparan sulphate, respectively, may be to aid non-specific attachment to the host cell. It is proposed that a high Kmucin is the cost for subsequent binding of MERS-CoV to SAs on the cell surface to partially overcome the unfavourable entropy of immobilisation as the virus adopts the correct orientation for spike protein interactions with its protein cellular receptor DPP4.

Assessing the aggregated probability of entry of a novel prion disease agent into the United Kingdom.

In 2018 prion disease was detected in camels at an abattoir in Algeria for the first time. The emergence of prion disease in this species made it prudent to assess the probability of entry of the pathogen into the United Kingdom (UK) from this region. Potentially contaminated products were identified as evidenced by other prion diseases. The aggregated probability of entry of the pathogen was estimated as very high and high for legal milk and cheese imports respectively and very high, high and high for illegal meat, milk and cheese products respectively. This aggregated probability represents a qualitative assessment of the probability of one or more entry events per year into the UK; it gives no indication of the number of entry events per year. The uncertainty associated with these estimates was high due to the unknown variation in prevalence of infection in camels and an uncertain number and type of illegal products entering the UK. Potential public health implications of this pathogen are unknown although there is currently no evidence of zoonotic transmission of prion diseases other than bovine spongiform encephalopathy to humans.

How virus size and attachment parameters affect the temperature sensitivity of virus binding to host cells: Predictions of a thermodynamic model for arboviruses and HIV.

Virus binding to host cells involves specific interactions between viral (glyco)proteins (GP) and host cell surface receptors (Cr) (protein or sialic acid (SA)). The magnitude of the enthalpy of association changes with temperature according to the change in heat capacity (ΔCp) on GP/Cr binding, being little affected for avian influenza virus (AIV) haemagglutinin (HA) binding to SA (ΔCp = 0 kJ/mol/K) but greatly affected for HIV gp120 binding to CD4 receptor (ΔCp = -5.0 kJ/mol/K). A thermodynamic model developed here predicts that values of ΔCp from 0 to ~-2.0 kJ/mol/K have relatively little impact on the temperature sensitivity of the number of mosquito midgut cells with bound arbovirus, while intermediate values of ΔCp of ~-3.0 kJ/mol/K give a peak binding at a temperature of ~20 °C as observed experimentally for Western equine encephalitis virus. More negative values of ΔCp greatly decrease arbovirus binding at temperatures below ~20 °C. Thus to promote transmission at low temperatures, arboviruses may benefit from ΔCp ~ 0 kJ/mol/K as for HA/SA and it is interesting that bluetongue virus binds to SA in midge midguts. Large negative values of ΔCp as for HIV gp120:CD4 diminish binding at 37 °C. Of greater importance, however, is the decrease in entropy of the whole virus (ΔSa_immob) on its immobilisation on the host cell surface. ΔSa_immob presents a repulsive force which the enthalpy-driven GP/Cr interactions weakened at higher temperatures struggle to overcome. ΔSa_immob is more negative (less favourable) for larger diameter viruses which therefore show diminished binding at higher temperatures than smaller viruses. It is proposed that small size phenotype through a less negative ΔSa_immob is selected for viruses infecting warmer hosts thus explaining the observation that virion volume decreases with increasing host temperature from 0 °C to 40 °C in the case of dsDNA viruses. Compared to arboviruses which also infect warm-blooded vertebrates, HIV is large at 134 nm diameter and thus would have a large negative ΔSa_immob which would diminish its binding at human body temperature. It is proposed that prior non-specific binding of HIV through attachment factors takes much of the entropy loss for ΔSa_immob so enhancing subsequent specific gp120:CD4 binding at 37 °C. This is consistent with the observation that HIV attachment factors are not essential but augment infection. Antiviral therapies should focus on increasing virion size, for example through binding of zinc oxide nanoparticles to herpes simplex virus, hence making ΔSa_immob more negative, and thus reducing binding affinity at 37 °C.

Risk assessments for quality-assured, source-segregated composts and anaerobic digestates for a circular bioeconomy in the UK.

A circular economy relies on demonstrating the quality and environmental safety of wastes that are recovered and reused as products. Policy-level risk assessments, using generalised exposure scenarios, and informed by stakeholder communities have been used to appraise the acceptability of necessary changes to legislation, allowing wastes to be valued, reused and marketed. Through an extensive risk assessment exercise, summarised in this paper, we explore the burden of proof required to offer safety assurance to consumer and brand-sensitive food sectors in light of attempts to declassify, as wastes, quality-assured, source-segregated compost and anaerobic digestate products in the United Kingdom. We report the residual microbiological and chemical risks estimated for both products in land application scenarios and discuss these in the context of an emerging UK bioeconomy worth £52bn per annum. Using plausible worst case assumptions, as demanded by the quality food sector, risk estimates and hazard quotients were estimated to be low or negligible. For example, the human health risk of E. coli 0157 illness from exposure to microbial residuals in quality-assured composts, through a ready-to-eat vegetable consumption exposure route, was estimated at ~10-8 per person per annum. For anaerobic digestion residues, 7 × 10-3cases of E. coli 0157 were estimated per annum, a potential contribution of 0.0007% of total UK cases. Hazard quotients for potential chemical contaminants in both products were insufficient in magnitude to merit detailed quantitative risk assessments. Stakeholder engagement and expert review was also a substantive feature of this study. We conclude that quality-assured, source-segregated products applied to land, under UK quality protocols and waste processing standards, pose negligible risks to human, animal, environmental and crop receptors, providing that risk management controls set within the standards and protocols are adhered to.

The Risk of Infection by African Swine Fever Virus in European Swine Through Boar Movement and Legal Trade of Pigs and Pig Meat.

African swine fever (ASF) is currently spreading westwards throughout Europe and eastwards into China, with cases occurring in both wild boar and domestic pigs. A generic risk assessment framework is used to determine the probability of first infection with ASF virus (ASFV) at a fine spatial scale across European Union Member States. The framework aims to assist risk managers across Europe with their ASF surveillance and intervention activities. Performing the risk assessment at a fine spatial scale allows for hot-spot surveillance, which can aid risk managers by directing surveillance or intervention resources at those areas or pathways deemed most at risk, and hence enables prioritization of limited resources. We use 2018 cases of ASF to estimate prevalence of the disease in both wild boar and pig populations and compute the risk of initial infection for 2019 at a 100 km2 cell resolution via three potential pathways: legal trade in live pigs, natural movement of wild boar, and legal trade in pig meat products. We consider the number of pigs, boar and amount of pig meat entering our area of interest, the prevalence of the disease in the origin country, the probability of exposure of susceptible pigs or boar in the area of interest to introduced infected pigs, boar, or meat from an infected pig, and the probability of transmission to susceptible animals. We provide maps across Europe indicating regions at highest risk of initial infection. Results indicate that the risk of ASF in 2019 was predominantly focused on those regions which already had numerous cases in 2018 (Poland, Lithuania, Hungary, Romania, and Latvia). The riskiest pathway for ASFV transmission to pigs was the movement of wild boar for Eastern European countries and legal trade of pigs for Western European countries. New infections are more likely to occur in wild boar rather than pigs, for both the pig meat and wild boar movement pathways. Our results provide an opportunity to focus surveillance activities and thus increase our ability to detect ASF introductions earlier, a necessary requirement if we are to successfully control the spread of this devastating disease for the pig industry.

Towards a thermodynamic mechanistic model for the effect of temperature on arthropod vector competence for transmission of arboviruses.

Arboviruses such as West Nile virus (WNV), bluetongue virus (BTV), dengue virus (DENV) and chikungunya virus (CHIKV) infect their arthropod vectors over a range of average temperatures depending on the ambient temperature. How the transmission efficiency of an arbovirus (i.e. vector competence) varies with temperature influences not only the short term risk of arbovirus outbreaks in humans and livestock but also the long term impact of climate change on the geographical range of the virus. The strength of the interaction between viral surface (glyco)protein (GP) and the host cell receptor (Cr) on binding of virus to host cell is defined by the thermodynamic dissociation constant Kd_receptor which is assumed to equal 10-3 M (at 37 °C) for binding of a sialic acid (SA) on the arthropod midgut epithelial cell surface to a SA-binding site on the surface of BTV, for example. Here virus binding affinity is modelled with increasing number of GP/Cr contacts at temperatures from 10 °C to 35 °C taking into account the change in entropy on immobilization of the whole virus on binding (ΔSa_immob). Based on published data, three thermodynamic GP/Cr binding scenarios, namely enthalpy-driven, entropy-assisted and entropy-driven, are shown to affect the temperature sensitivity of virus binding in different ways. Thus for enthalpy-driven GP/Cr binding, viruses bind host cells much more strongly at 10 °C than 35 °C. A mechanistic model is developed for the number of arthropod midgut cells with bound virus and by building in a kinetic component for the rate of arbovirus replication and subsequent spread to the arthropod salivary glands, a model for the effect of temperature on vector competence is developed. The model separates the opposing effects of temperature on midgut cell binding affinity from the kinetic component of virogenesis. It successfully accommodates both increases in vector competence with temperature as for DENV and WNV in mosquitoes and decreases as for the CHIKV 2010-1909 strain in various populations of Aedes albopictus mosquitoes. Enhanced cell binding at lower temperatures through enthalpy-driven GP/Cr binding compensates for the lower replication rate to some degree such that some transmission can still occur at lower temperatures. In contrast, the strength of entropy-driven GP/Cr binding diminishes at low temperatures although there is no minimum temperature threshold for transmission efficiency. The magnitude of ΔSa_immob is an important data gap. It is concluded that thermodynamic and kinetic data obtained at the molecular level will prove important in modelling vector competence with temperature.

A generic framework for spatial quantitative risk assessments of infectious diseases: Lumpy skin disease case study.

The increase in availability of spatial data and the technological advances to handle such data allow for subsequent improvements in our ability to assess risk in a spatial setting. We provide a generic framework for quantitative risk assessments of disease introduction that capitalizes on these new data. It can be adopted across multiple spatial scales, for any pathogen, method of transmission or location. The framework incorporates the risk of initial infection in a previously uninfected location due to registered movement (e.g., trade) and unregistered movement (e.g., daily movements of wild animals). We discuss the steps of the framework and the data required to compute it. We then outline how this framework is applied for a single pathway using lumpy skin disease as a case study, a disease which had an outbreak in the Balkans in 2016. We calculate the risk of initial infection for the rest of Europe in 2016 due to trade. We perform the risk assessment on 3 spatial scales-countries, regions within countries and individual farms. We find that Croatia (assuming no vaccination occurred) has the highest mean probability of infection, with Italy, Hungary and Spain following. Including import detection of infected trade does reduce risk but this reduction is proportionally lower for countries with highest risk. The risk assessment results are consistent across the spatial scales, while in addition, at the finer spatial scales, it highlights specific areas or individual locations of countries on which to focus surveillance.

Using thermodynamic parameters to calibrate a mechanistic dose-response for infection of a host by a virus.

Assessing the risk of infection from emerging viruses or of existing viruses jumping the species barrier into novel hosts is limited by the lack of dose response data. The initial stages of the infection of a host by a virus involve a series of specific contact interactions between molecules in the host and on the virus surface. The strength of the interaction is quantified in the literature by the dissociation constant (Kd) which is determined experimentally and is specific for a given virus molecule/host molecule combination. Here, two stages of the initial infection process of host intestinal cells are modelled, namely escape of the virus in the oral challenge dose from the innate host defenses (e.g. mucin proteins in mucus) and the subsequent binding of any surviving virus to receptor molecules on the surface of the host epithelial cells. The strength of virus binding to host cells and to mucins may be quantified by the association constants, Ka and Kmucin, respectively. Here, a mechanistic dose-response model for the probability of infection of a host by a given virus dose is constructed using Ka and Kmucin which may be derived from published Kd values taking into account the number of specific molecular interactions. It is shown that the effectiveness of the mucus barrier is determined not only by the amount of mucin but also by the magnitude of Kmucin. At very high Kmucin values, slight excesses of mucin over virus are sufficient to remove all the virus according to the model. At lower Kmucin values, high numbers of virus may escape even with large excesses of mucin. The output from the mechanistic model is the probability (p1) of infection by a single virion which is the parameter used in conventional dose-response models to predict the risk of infection of the host from the ingested dose. It is shown here how differences in Ka (due to molecular differences in an emerging virus strain or new host) affect p1, and how these differences in Ka may be quantified in terms of two thermodynamic parameters, namely enthalpy and entropy. This provides the theoretical link between sequencing data and risk of infection. Lack of data on entropy is a limitation at present and may also affect our interpretation of Kd in terms of infectivity. It is concluded that thermodynamic approaches have a major contribution to make in developing dose-response models for emerging viruses.

Application of a quantitative entry assessment model to compare the relative risk of incursion of zoonotic bat-borne viruses into European Union Member States.

This paper presents a quantitative assessment model for the risk of entry of zoonotic bat-borne viruses into the European Union (EU). The model considers four routes of introduction: human travel, legal trade of products, live animal imports and illegal import of bushmeat and was applied to five virus outbreak scenarios. Two scenarios were considered for Zaire ebolavirus (wEBOV, cEBOV) and other scenarios for Hendra virus, Marburg virus (MARV) and Middle East Respiratory Syndrome Coronavirus (MERS-CoV). The use of the same framework and generic data sources for all EU Member States (MS) allows for a relative comparison of the probability of virus introduction and of the importance of the routes of introduction among MSs. According to the model wEBOV posed the highest risk of an introduction event within the EU, followed by MARV and MERS-CoV. However, the main route of introduction differed, with wEBOV and MERS-CoV most likely through human travel and MARV through legal trade of foodstuffs. The relative risks to EU MSs as entry points also varied between outbreak scenarios, highlighting the heterogeneity in global trade and travel to the EU MSs. The model has the capability to allow for a continual updating of the risk estimate using new data as, and when, it becomes available. The model provides an horizon scanning tool for use when available data are limited and, therefore, the absolute risk estimates often have high uncertainty. Sensitivity analysis suggested virus prevalence in bats has a large influence on the results; a 90% reduction in prevalence reduced the risk of introduction considerably and resulted in the relative ranking of MARV falling below that for MERS-CoV, due to this parameter disproportionately affecting the risk of introduction from the trade route over human travel.

A Generic Quantitative Risk Assessment Framework for the Entry of Bat-Borne Zoonotic Viruses into the European Union.

Bat-borne viruses have been linked to a number of zoonotic diseases; in 2014 there have been human cases of Nipah virus (NiV) in Bangladesh and Ebola virus in West and Central Africa. Here we describe a model designed to provide initial quantitative predictions of the risk of entry of such viruses to European Union (EU) Member States (MSs) through four routes: human travel, legal trade (e.g. fruit and animal products), live animal movements and illegal importation of bushmeat. The model utilises available datasets to assess the movement via these routes between individual countries of the world and EU MSs. These data are combined with virus specific data to assess the relative risk of entry between EU MSs. As a case study, the model was parameterised for NiV. Scenario analyses showed that the selection of exporting countries with NiV and potentially contaminated trade products were essential to the accuracy of all model outputs. Uncertainty analyses of other model parameters identified that the model expected number of years to an introduction event within the EU was highly susceptible to the prevalence of NiV in bats. The relative rankings of the MSs and routes, however, were more robust. The UK, the Netherlands and Germany were consistently the most likely points of entry and the ranking of most MSs varied by no more than three places (maximum variation five places). Legal trade was consistently the most likely route of entry, only falling below human travel when the estimate of the prevalence of NiV in bats was particularly low. Any model-based calculation is dependent on the data available to feed into the model and there are distinct gaps in our knowledge, particularly in regard to various pathogen/virus as well as host/bat characteristics. However, the strengths of this model lie in the provision of relative comparisons of risk among routes and MSs. The potential for expansion of the model to include other routes and viruses and the possibility of rapid parameterisation demonstrates its potential for use in an outbreak situation.

Hyalomma ticks on northward migrating birds in southern Spain: Implications for the risk of entry of Crimean-Congo haemorrhagic fever virus to Great Britain.

Crimean-Congo haemorrhagic fever virus (CCHFV) is a zoonotic virus transmitted by Hyalomma ticks, the immature stages of which may be carried by migratory birds. In this study, a total of 12 Hyalomma ticks were recovered from five of 228 migratory birds trapped in Spring, 2012 in southern Spain along the East Atlantic flyway. All collected ticks tested negative for CCHFV. While most birds had zero Hyalomma ticks, two individuals had four and five ticks each and the statistical distribution of Hyalomma tick counts per bird is over-dispersed compared to the Poisson distribution, demonstrating the need for intensive sampling studies to avoid underestimating the total number of ticks. Rates of tick exchange on migratory birds during their northwards migration will affect the probability that a Hyalomma tick entering Great Britain is positive for CCHFV. Drawing on published data, evidence is presented that the latitude of a European country affects the probability of entry of Hyalomma ticks on wild birds. Further data on Hyalomma infestation rates and tick exchange rates are required along the East Atlantic flyway to further our understanding of the origin of Hyalomma ticks (i.e., Africa or southern Europe) and hence the probability of entry of CCHFV into GB.

Drivers for emerging issues in animal and plant health.

The history of agriculture includes many animal and plant disease events that have had major consequences for the sector, as well as for humans. At the same time, human activities beyond agriculture have often driven the emergence of diseases. The more that humans expand the footprint of the global population, encroach into natural habitats, alter these habitats to extract resources and intensify food production, as well as move animals, people and commodities along with the pathogens they carry, the greater the potential for pathogens and pests to spread and for infection to emerge or re-emerge. While essential to human well-being, producing food also plays a major role in disease dynamics. The risk of emergence of pests and pathogens has increased as a consequence of global changes in the way food is produced, moved and consumed. Climate change is likely to increase pressure on the availability of food and provide newly suitable conditions for invasive pests and pathogens. Human population displacements due to economic, political and humanitarian crises represent another set of potential drivers for emerging issues. The overlapping drivers of plant, animal and human disease emergence and environmental changes point towards the concept of 'One Health'. This paradigm underlines the urgent need to understand the influence of human behaviour and incorporate this understanding into our approach to emerging risks. For this, we face two major challenges. One is cultural; the second is methodological. We have to look at systems not under the narrow view of specific hazards but with a wider approach to system dynamics, and consider a broad spectrum of potential outcomes in terms of risk. In addition, we have to make sense of the vast amounts of data that are available in the modern age. This paper aims to help in preparing for the cultural and methodological shifts needed in our approach to emerging risks.

Potential for introduction of bat-borne zoonotic viruses into the EU: a review.

Bat-borne viruses can pose a serious threat to human health, with examples including Nipah virus (NiV) in Bangladesh and Malaysia, and Marburg virus (MARV) in Africa. To date, significant human outbreaks of such viruses have not been reported in the European Union (EU). However, EU countries have strong historical links with many of the countries where NiV and MARV are present and a corresponding high volume of commercial trade and human travel, which poses a potential risk of introduction of these viruses into the EU. In assessing the risks of introduction of these bat-borne zoonotic viruses to the EU, it is important to consider the location and range of bat species known to be susceptible to infection, together with the virus prevalence, seasonality of viral pulses, duration of infection and titre of virus in different bat tissues. In this paper, we review the current scientific knowledge of all these factors, in relation to the introduction of NiV and MARV into the EU.

A commentary on recent water safety initiatives in the context of water utility risk management.

Over the last decade, suppliers of drinking water have recognised the limitations of relying solely on end-product monitoring to ensure safe water quality and have sought to reinforce their approach by adopting preventative strategies where risks are proactively identified, assessed and managed. This is leading to the development of water safety plans; structured 'route maps' for managing risks to water supply, from catchment to consumer taps. This paper reviews the Hazard Analysis and Critical Control Point (HACCP) procedure on which many water safety plans are based and considers its appropriateness in the context of drinking water risk management. We examine water safety plans in a broad context, looking at a variety of monitoring, optimisation and risk management initiatives that can be taken to improve drinking water safety. These are cross-compared using a simple framework that facilitates an integrated approach to water safety. Finally, we look at how risk management practices are being integrated across water companies and how this is likely to affect the future development of water safety plans.

The effect of drinking water treatment on the spatial heterogeneity of micro-organisms: implications for assessment of treatment efficiency and health risk.

The effect of drinking water treatment (ferric coagulation, floc blanket clarification, rapid sand filtration) on the spatial heterogeneity of five species of micro-organism was studied at pilot scale. It was found that the spatial heterogeneity of vegetative bacteria (namely total coliform and heterotrophic (22 degrees C; 3 d) bacteria) was little affected by treatment. Indeed, counts of total coliform bacteria within 500 l volumes of treated water were Poisson distributed (i.e. showed minimum variation). In contrast, treatment appeared to increase the spatial heterogeneity (or clustering) of both aerobic spores indigenous to the raw water and Bacillus subtilis var niger spores added to the raw water. Furthermore, B. subtilis var niger spores added to the raw water were detected in the treated water 25 h after termination of spiking to the raw water. The effect on C. parvum oocysts added to the raw water could not be determined because few oocysts broke through treatment into the treated water. Indeed oocyst removals of 5-6 logs were apparent. "Species-specific" differences in the removal ratios were also demonstrated. It is concluded that audits for treatment processes based on single 100 ml "spot" samples for spores will tend to over-estimate the net spore removal and hence underestimate the public health risk. Spatial heterogeneity of counts in treated water contributes to explaining why no "ideal" surrogate has been identified for treatment plant performance.