A Portrait of Sentinel Surveillance Networks for Vector-Borne Diseases: A Scoping Review Supporting Sentinel Network Design.

Vector-borne diseases (VBDs) are continuing to emerge globally, requiring new surveillance systems to follow increasing VBD risk for human populations. Sentinel surveillance is an approach that allows tracking of disease risk through time using limited resources. However, there is no consensus on how best to design a sentinel surveillance network in the context of VBDs. We conducted a scoping review to compare VBD sentinel surveillance systems worldwide with the aim of identifying key design features associated with effective networks. Overall, VBD surveillance networks were used most commonly for malaria, West Nile virus, and lymphatic filariasis. A total of 45 criteria for the selection of sentinel unit location were identified. Risk-based criteria were the most often used, and logistic regression showed that using risk-based criteria dependent on host animals is particularly correlated with surveillance system sensitivity (p < 0.018). We identify tools that could prove valuable for sentinel surveillance network design, including a standardized approach for evaluating surveillance systems and a tool to prioritize criteria for selecting optimal geographic locations for spatial sentinel units.

A qualitative program evaluation of the Publicly Available International Foodborne Outbreak Database.

BACKGROUND: The Publicly Available International Foodborne Outbreak Database (PAIFOD) is a regularly updated repository that contains international outbreak data collected from multiple surveillance systems and sources. As of February 2020, the database contained more than 13,000 entries spanning over 20 years. PAIFOD is the only known database that captures international foodborne outbreak data. OBJECTIVE: To explore user perceptions and identify potential directions for PAIFOD and make recommendations for databases with food safety information. METHODS: Between January and March 2020, 16 semistructured telephone interviews were conducted with 24 previous, current and potential PAIFOD users. Interviewees were asked about their knowledge of and experience of using PAIFOD as well as about its strengths and limitations and recommendations for the database. An inductive thematic analysis approach was used to analyze qualitative data and generate themes. RESULTS: Four main themes were generated based on the 24 interviewees' accounts of their experience with and recommendations for PAIFOD: participants viewed PAIFOD as a useful tool; they weren't familiar with its contents or purpose; they stated it should become an open-access platform or linked with another information-sharing initiative; and they considered that PAIFOD had the potential to enhance the Agency's reputation by becoming widely recognized and used. CONCLUSION: This work, along with the ever-changing landscape of foodborne surveillance, supports the need to ensure that PAIFOD is updated to meet the modern-day demands of food safety experts.

Eastern Equine Encephalitis Virus: A Scoping Review of the Global Evidence.

Background: Eastern equine encephalitis virus (EEEV) is a mosquito-borne virus that is primarily found in North America and the Caribbean. Over the past decade there has been an increase in virus activity, including large outbreaks in human and horse populations. Predicted climate change is expected to affect the range of mosquitoes including vectors of EEEV, which may alter disease risk posing a public health concern. Methods: A scoping review (ScR) was conducted to identify and characterize the global evidence on EEEV. A thorough search was conducted in relevant bibliographic databases and government websites. Two reviewers screened titles and abstracts for relevance and the characteristics of relevant articles were extracted using a uniformly implemented data collection form. The study protocol was developed a priori and described the methods and tools used and this article follows the PRISMA-ScR guidelines for reporting ScRs. Results: The ScR included 718 relevant research articles. The majority of the articles originated from North America (97%) between 1933 and 2019. EEEV has been identified in 35 species of mosquitoes, over 200 species of birds, various domestic animals, wild mammals, reptiles, and amphibians. Articles identified in this ScR primarily covered three topic areas: epidemiology of hosts and vectors (344 articles) including surveillance results (138), pathogenesis of EEEV in hosts (193), and in vitro studies characterizing EEEV (111). Fewer articles evaluated the accuracy of diagnostic tests (63), the efficacy of mitigation strategies (62), transmission dynamics (56), treatment of EEEV in hosts (10), societal knowledge, attitudes, and perceptions (4), and economic burden (2). Conclusion: With the projected impact of climate change on mosquito populations, it is expected that the risk of EEEV could change resulting in higher disease burden or spread into previously unaffected areas. Future research efforts should focus on closing some of the important knowledge gaps identified in this ScR.

Zoonotic Babesia: A scoping review of the global evidence.

BACKGROUND: Babesiosis is a parasitic vector-borne disease of increasing public health importance. Since the first human case was reported in 1957, zoonotic species have been reported on nearly every continent. Zoonotic Babesia is vectored by Ixodes ticks and is commonly transmitted in North America by Ixodes scapularis, the tick species responsible for transmitting the pathogens that also cause Lyme disease, Powassan virus, and anaplasmosis in humans. Predicted climate change is expected to impact the spread of vectors, which is likely to affect the distribution of vector-borne diseases including human babesiosis. METHODS: A scoping review has been executed to characterize the global evidence on zoonotic babesiosis. Articles were compiled through a comprehensive search of relevant bibliographic databases and targeted government websites. Two reviewers screened titles and abstracts for relevance and characterized full-text articles using a relevance screening and data characterization tool developed a priori. RESULTS: This review included 1394 articles relevant to human babesiosis and/or zoonotic Babesia species. The main zoonotic species were B. microti, B. divergens, B. duncani and B. venatorum. Articles described a variety of study designs used to study babesiosis in humans and/or zoonotic Babesia species in vectors, animal hosts, and in vitro cell cultures. Topics of study included: pathogenesis (680 articles), epidemiology (480), parasite characterization (243), diagnostic test accuracy (98), mitigation (94), treatment (65), transmission (54), surveillance (29), economic analysis (7), and societal knowledge (1). No articles reported predictive models investigating the impact of climate change on Babesia species. CONCLUSION: Knowledge gaps in the current evidence include research on the economic burden associated with babesiosis, societal knowledge studies, surveillance of Babesia species in vectors and animal hosts, and predictive models on the impact of climate change. The scoping review results describe the current knowledge and knowledge gaps on zoonotic Babesia which can be used to inform future policy and decision making.

Cache Valley virus: A scoping review of the global evidence.

Cache Valley virus (CVV) is a mosquito-borne RNA virus detected throughout North America, Central America and parts of South America. A limited number of human case reports have described severe illness. CVV infection has been associated with outbreaks of congenital defects in small ruminants in Canada and the United States. A scoping review was conducted to identify, characterize and summarize research on CVV, and to identify research gaps. A structured search was conducted in eight electronic databases, with additional search verification and grey literature investigation. All captured studies were independently appraised by two reviewers for relevance and data characterization. The review captured 143 relevant studies investigating CVV epidemiology (n = 104), pathogenesis (n = 37), viral characteristics (n = 24), transmission (n = 14), diagnostic test performance (n = 8) and mitigation strategies (n = 2). Evidence of CVV infection was found in mosquito studies (n = 47), and serological evidence of exposure was demonstrated in animals (n = 41), as well as human (n = 20) studies. In sheep, five outbreaks of birth defects following asymptomatic dam CVV infection during the first 50 days of pregnancy were reported. Only six human cases of CVV-associated illness were captured, with case symptoms described as initially non-specific, progressing to more severe clinical signs (e.g., meningitis). No research was identified investigating treatment, societal knowledge and risk perception, economic burden or predictive models related to the impact of climate change on CVV. CVV circulates in mosquito and animal species across a large area of the Americas. Small ruminants are the only animals in which CVV-associated clinical disease has been extensively studied. It is likely that human cases are under-reported or misdiagnosed. Future research should focus on the impact of CVV infection in human and animal populations.

A systematic review of individual and community mitigation measures for prevention and control of chikungunya virus.

BACKGROUND: Chikungunya is a mosquito-borne virus transmitted by mosquitoes from the Aedes genus. The virus, endemic to parts of Asia and Africa, has recently undergone an emergence in other parts of the world where it was previously not found including Indian Ocean Islands, Europe, the Western Pacific and the Americas. There is no vaccine against chikungunya virus, which means that prevention and mitigation rely on personal protective measures and community level interventions including vector control. METHODOLOGY/PRINCIPAL FINDINGS: A systematic review (SR) was conducted to summarize the literature on individual and community mitigation and control measures and their effectiveness. From a scoping review of the global literature on chikungunya, there were 91 articles that investigated mitigation or control strategies identified at the individual or community level. Of these, 81 were confirmed as relevant and included in this SR. The majority of the research was published since 2010 (76.5%) and was conducted in Asia (39.5%). Cross sectional studies were the most common study design (36.6%). Mitigation measures were placed into six categories: behavioural protective measures, insecticide use, public education, control of blood and blood products, biological vector control and quarantine of infected individuals. The effectiveness of various mitigation measures was rarely evaluated and outcomes were rarely quantitative, making it difficult to summarize results across studies and between mitigation strategies. Meta-analysis of the proportion of individuals engaging in various mitigation measures indicates habitat removal is the most common measure used, which may demonstrate the effectiveness of public education campaigns aimed at reducing standing water. CONCLUSIONS/SIGNIFICANCE: Further research with appropriate and consistent outcome measurements are required in order to determine which mitigation measures, or combination of mitigation measures, are the most effective at protecting against exposure to chikungunya virus.

A scoping review of published literature on chikungunya virus.

Chikungunya virus (CHIKV) has caused several major epidemics globally over the last two decades and is quickly expanding into new areas. Although this mosquito-borne disease is self-limiting and is not associated with high mortality, it can lead to severe, chronic and disabling arthritis, thereby posing a heavy burden to healthcare systems. The two main vectors for CHIKV are Aedes aegypti and Aedes albopictus (Asian tiger mosquito); however, many other mosquito species have been described as competent CHIKV vectors in scientific literature. With climate change, globalization and unfettered urban planning affecting many areas, CHIKV poses a significant public health risk to many countries. A scoping review was conducted to collate and categorize all pertinent information gleaned from published scientific literature on a priori defined aspects of CHIKV and its competent vectors. After developing a sensitive and specific search algorithm for the research question, seven databases were searched and data was extracted from 1920 relevant articles. Results show that CHIKV research is reported predominantly in areas after major epidemics have occurred. There has been an upsurge in CHIKV publications since 2011, especially after first reports of CHIKV emergence in the Americas. A list of hosts and vectors that could potentially be involved in the sylvatic and urban transmission cycles of CHIKV has been compiled in this scoping review. In addition, a repository of CHIKV mutations associated with evolutionary fitness and adaptation has been created by compiling and characterizing these genetic variants as reported in scientific literature.

Powassan virus, a scoping review of the global evidence.

BACKGROUND: Powassan virus (POWV), a flavivirus discovered in 1958, causes sporadic but severe cases of encephalitis in humans. Since 2007, the number of human Powassan cases diagnosed each year in the USA has steadily increased. This is in agreement with predictions that Powassan cases may increase in North America as a result of increased exposure to infected ticks. However, the increase may also reflect improved diagnostics and reporting among other factors. METHODS: A scoping review was prioritized to identify and characterize the global literature on POWV. Following an a priori developed protocol, a comprehensive search strategy was implemented. Two reviewers screened titles and abstracts for relevant research and the identified full papers were used to characterize the POWV literature using a predetermined data characterization tool. RESULTS: One hundred and seventy-eight articles were included. The majority of the studies were conducted in North America (88.2%) between 1958 and 2017. Both genotypes of POWV (Powassan lineage 1 and Deer Tick virus) were isolated or studied in vitro, in vectors, nonhuman hosts and human populations. To date, POWV has been reported in 147 humans in North America. The virus has also been isolated from five tick species, and several animals have tested positive for exposure to the virus. The relevant articles identified in this review cover the following eight topics: epidemiology (123 studies), pathogenesis (66), surveillance (33), virus characterization (22), POWV transmission (8), diagnostic test accuracy (8), treatment (4) and mitigation strategies (3). CONCLUSION: The literature on POWV is relatively small compared with other vector-borne diseases, likely because POWV has not been prioritized due to the small number of severe sporadic human cases. With the projected impact of climate change on tick populations, increases in the number of human cases are expected. It is recommended that future research efforts focus on closing some of the important knowledge gaps identified in this scoping review.

Environmental and social determinants of population vulnerability to Zika virus emergence at the local scale.

BACKGROUND: Zika virus (ZIKV) spread rapidly in the Americas in 2015. Targeting effective public health interventions for inhabitants of, and travellers to and from, affected countries depends on understanding the risk of ZIKV emergence (and re-emergence) at the local scale. We explore the extent to which environmental, social and neighbourhood disease intensity variables influenced emergence dynamics. Our objective was to characterise population vulnerability given the potential for sustained autochthonous ZIKV transmission and the timing of emergence. Logistic regression models estimated the probability of reporting at least one case of ZIKV in a given municipality over the course of the study period as an indicator for sustained transmission; while accelerated failure time (AFT) survival models estimated the time to a first reported case of ZIKV in week t for a given municipality as an indicator for timing of emergence. RESULTS: Sustained autochthonous ZIKV transmission was best described at the temporal scale of the study period (almost one year), such that high levels of study period precipitation and low mean study period temperature reduced the probability. Timing of ZIKV emergence was best described at the weekly scale for precipitation in that high precipitation in the current week delayed reporting. Both modelling approaches detected an effect of high poverty on reducing/slowing case detection, especially when inter-municipal road connectivity was low. We also found that proximity to municipalities reporting ZIKV had an effect to reduce timing of emergence when located, on average, less than 100 km away. CONCLUSIONS: The different modelling approaches help distinguish between large temporal scale factors driving vector habitat suitability and short temporal scale factors affecting the speed of spread. We find evidence for inter-municipal movements of infected people as a local-scale driver of spatial spread. The negative association with poverty suggests reduced case reporting in poorer areas. Overall, relatively simplistic models may be able to predict the vulnerability of populations to autochthonous ZIKV transmission at the local scale.

Risk perceptions, attitudes, and knowledge of chikungunya among the public and health professionals: a systematic review.

BACKGROUND: Recently, attention to chikungunya has increased due to its spread into previously non-endemic areas. Since there is no available treatment or vaccine, most intervention strategies focus on mosquito bite prevention and mosquito control, which require community involvement to be successful. Thus, our objective was to systematically review the global primary literature on the risk perceptions, attitudes, and knowledge of chikungunya among the public and health professionals to inform future research and improve our understanding on which intervention strategies are likely to be successful. METHODS: Potentially relevant articles were identified through a standardized systematic review (SR) process consisting of the following steps: comprehensive search strategy in seven databases (Scopus, PubMed, CINAHL, CAB, LILACS, Agricola, and Cochrane) and a grey literature search of public health organizations, relevance screening, risk of bias assessment, and data extraction. Two independent reviewers performed each step. Reporting of this SR follows PRISMA reporting guidelines. RESULTS: Thirty-seven relevant articles were identified. The majority of the articles were published since 2011 (83.8%) and reported on studies conducted in Asia (48.7%) and the Indian Ocean Islands (24.3%). The results were separated into four categories: general knowledge and perceptions on chikungunya; perceptions on the risk and severity of chikungunya; knowledge of chikungunya-harboring vectors and transmission; and knowledge, perceptions, and attitudes on mitigation practices. Overall, the systematic review found that risk perceptions, attitudes, and knowledge of chikungunya among the public and health professionals vary across populations and countries and knowledge is higher in areas that have experienced an outbreak. CONCLUSION: The results suggest that most of the affected populations in this study do not understand mosquito borne diseases or chikungunya and are therefore less likely to protect themselves from mosquito bites. While more research is required to improve the generalizability of this dataset, it appears that a lack of knowledge is an important barrier for motivating community level interventions and personal protection against mosquitoes.

Assessment of the Probability of Autochthonous Transmission of Chikungunya Virus in Canada under Recent and Projected Climate Change.

BACKGROUND: Chikungunya virus (CHIKV) is a reemerging pathogen transmitted by Aedes aegypti and Aedes albopictus mosquitoes. The ongoing Caribbean outbreak is of concern due to the potential for infected travelers to spread the virus to countries where vectors are present and the population is susceptible. Although there has been no autochthonous transmission of CHIKV in Canada, there is concern that both Ae. albopictus and CHIKV will become established, particularly under projected climate change. We developed risk maps for autochthonous CHIKV transmission in Canada under recent (1981­2010) and projected climate (2011­2040 and 2041­2070). METHODS: The risk for CHIKV transmission was the combination of the climatic suitability for CHIKV transmission potential and the climatic suitability for the presence of Ae. albopictus; the former was assessed using a stochastic model to calculate R0 and the latter was assessed by deriving a suitability indicator (SIG) that captures a set of climatic conditions known to influence the ecology of Ae. albopictus. R0 and SIG were calculated for each grid cell in Canada south of 60°N, for each time period and for two emission scenarios, and combined to produce overall risk categories that were mapped to identify areas suitable for transmission and the duration of transmissibility. FINDINGS: The risk for autochthonous CHIKV transmission under recent climate is very low with all of Canada classified as unsuitable or rather unsuitable for transmission. Small parts of southern coastal British Columbia become progressively suitable with short-term and long-term projected climate; the duration of potential transmission is limited to 1­2 months of the year. INTERPRETATION: Although the current risk for autochthonous CHIKV transmission in Canada is very low, our study could be further supported by the routine surveillance of Ae. albopictus in areas identified as potentially suitable for transmission given our uncertainty on the current distribution of this species in Canada. https://doi.org/10.1289/EHP669

The Accuracy of Diagnostic Tests for Lyme Disease in Humans, A Systematic Review and Meta-Analysis of North American Research.

There has been an increasing incidence of Lyme disease (LD) in Canada and the United States corresponding to the expanding range of the Ixodes tick vector and Lyme disease agent (Borrelia burgdorferi sensu stricto). There are many diagnostic tests for LD available in North America, all of which have some performance issues, and physicians are concerned about the appropriate use and interpretation of these tests. The objective of this systematic review is to summarize the North American evidence on the accuracy of diagnostic tests and test regimes at various stages of LD. Included in the review are 48 studies on diagnostic tests used in North America published since 1995. Thirteen studies examined a two-tier serological test protocol vs. clinical diagnosis, 24 studies examined single assays vs. clinical diagnosis, 9 studies examined single immunoblot vs. clinical diagnosis, 7 studies compared culture or PCR direct detection methods vs. clinical diagnosis, 22 studies compared two or more tests with each other and 8 studies compared a two-tiered serological test protocol to another test. Recent studies examining the sensitivity and specificity of various test protocols noted that the Immunetics® C6 B. burgdorferi ELISA™ and the two tier approach have superior specificity compared to proposed replacements, and the CDC recommended western blot algorithm has equivalent or superior specificity over other proposed test algorithms. There is a dramatic increase in test sensitivity with progression of B. burgdorferi infection from early to late LD. Direct detection methods, culture and PCR of tissue or blood samples were not as sensitive or timely compared to serological testing. It was also noted that there are a large number of both commercial (n = 42) and in-house developed tests used by private laboratories which have not been evaluated in the primary literature.

A systematic review and meta-analysis of the effectiveness of food safety education interventions for consumers in developed countries.

BACKGROUND: Foodborne illness has a large public health and economic burden worldwide, and many cases are associated with food handled and prepared at home. Educational interventions are necessary to improve consumer food safety practices and reduce the associated burden of foodborne illness. METHODS: We conducted a systematic review and targeted meta-analyses to investigate the effectiveness of food safety education interventions for consumers. Relevant articles were identified through a preliminary scoping review that included: a comprehensive search in 10 bibliographic databases with verification; relevance screening of abstracts; and extraction of article characteristics. Experimental studies conducted in developed countries were prioritized for risk-of-bias assessment and data extraction. Meta-analysis was conducted on data subgroups stratified by key study design-intervention-population-outcome categories and subgroups were assessed for their quality of evidence. Meta-regression was conducted where appropriate to identify possible sources of between-trial heterogeneity. RESULTS: We identified 79 relevant studies: 17 randomized controlled trials (RCTs); 12 non-randomized controlled trials (NRTs); and 50 uncontrolled before-and-after studies. Several studies did not provide sufficient details on key design features (e.g. blinding), with some high risk-of-bias ratings due to incomplete outcome data and selective reporting. We identified a moderate to high confidence in results from two large RCTs investigating community- and school-based educational training interventions on behaviour outcomes in children and youth (median standardized mean difference [SMD] = 0.20, range: 0.05, 0.35); in two small RCTs evaluating video and written instructional messaging on behavioural intentions in adults (SMD = 0.36, 95% confidence interval [CI]: 0.02, 0.69); and in two NRT studies for university-based education on attitudes of students and staff (SMD = 0.26, 95% CI: 0.10, 0.43). Uncontrolled before-and-after study outcomes were very heterogeneous and we have little confidence that the meta-analysis results reflect the true effect. Some variation in outcomes was explained in meta-regression models, including a dose effect for behaviour outcomes in RCTs. CONCLUSIONS: In controlled trials, food safety education interventions showed significant effects in some contexts; however, many outcomes were very heterogeneous and do not provide a strong quality of evidence to support decision-making. Future research in this area is needed using more robust experimental designs to build on interventions shown to be effective in uncontrolled before-and-after studies.

Scoping Review of Research on the Effectiveness of Food-Safety Education Interventions Directed at Consumers.

Improper food handling by consumers at home is a major cause of foodborne illness. Therefore, effective education strategies are essential to change consumers' food safety attitudes and behaviors. The purpose of this scoping review was to identify and characterize primary literature examining the effectiveness of consumer food-safety education interventions. Ten bibliographic databases were searched using a comprehensive search strategy. Citations were identified; two reviewers screened them for relevance and characterized relevant articles. To ensure results would be applicable to end users, stakeholders were engaged to provide input on the review scope, methods, and results. We identified 246 relevant articles, of which 150 were quantitative, 66 qualitative, and 30 mixed-method research studies. Most studies (64.2%) were published in the United States, using an uncontrolled before-and-after study design (31.3%), and investigated the effectiveness of community-based training sessions and workshops (52.0%). Research gaps were found in the number of randomized controlled studies conducted, academic- and school-based courses and curricula investigated, and interventions targeting high-risk populations (e.g., pregnant women, those who are immunocompromised) and using new media channels (e.g., social media). Key opportunities to enhance the utility of future primary research investigating consumer food-safety interventions include the following: using studies based on behavior-change theories and formative research; engaging the target population in the research; using validated instruments to measures outcomes; and reporting intervention characteristics and outcomes completely. Results of this review can be used to prioritize future primary research and decision-making in this area.

Novel repressor of Escherichia coli O157:H7 motility encoded in the putative fimbrial cluster OI-1.

Escherichia coli O157:H7 is a gastrointestinal pathogen that has become a serious public health concern, as it is associated with outbreaks and severe diseases such as hemolytic-uremic syndrome. The molecular basis of its greater virulence than that of other serotypes is not completely known. OI-1 is a putative fimbria-encoding genomic island that is found almost exclusively in O157:H7 Shiga toxin-producing E. coli strains and may be associated with the enhanced pathogenesis of these strains. In this study, we identified and characterized a novel repressor of flagellar synthesis encoded by OI-1. We showed that deletion of Z0021 increased the motility of E. coli O157:H7, which correlated with an increase in flagellin production and enhanced assembly of flagella on the cell surface. In contrast, overexpression of Z0021 inhibited motility. We demonstrated that Z0021 exerted its regulatory effects downstream of the transcription and translation of flhDC but prior to the activation of class II/III promoters. Furthermore, the master regulator of flagellar synthesis, FlhD(4)C(2), was shown to be a high-copy suppressor of the nonmotile phenotype associated with elevated levels of Z0021--a finding consistent with Z0021-FlhD(4)C(2) being a potential regulatory complex. This work provides insight into the mechanism by which Z0021, which we have named fmrA, represses flagellar synthesis and is the first report of a fimbrial-operon-encoded inhibitor of motility in E. coli O157:H7.

Genome sequence of adherent-invasive Escherichia coli and comparative genomic analysis with other E. coli pathotypes.

BACKGROUND: Adherent and invasive Escherichia coli (AIEC) are commonly found in ileal lesions of Crohn's Disease (CD) patients, where they adhere to intestinal epithelial cells and invade into and survive in epithelial cells and macrophages, thereby gaining access to a typically restricted host niche. Colonization leads to strong inflammatory responses in the gut suggesting that AIEC could play a role in CD immunopathology. Despite extensive investigation, the genetic determinants accounting for the AIEC phenotype remain poorly defined. To address this, we present the complete genome sequence of an AIEC, revealing the genetic blueprint for this disease-associated E. coli pathotype. RESULTS: We sequenced the complete genome of E. coli NRG857c (O83:H1), a clinical isolate of AIEC from the ileum of a Crohn's Disease patient. Our sequence data confirmed a phylogenetic linkage between AIEC and extraintestinal pathogenic E. coli causing urinary tract infections and neonatal meningitis. The comparison of the NRG857c AIEC genome with other pathogenic and commensal E. coli allowed for the identification of unique genetic features of the AIEC pathotype, including 41 genomic islands, and unique genes that are found only in strains exhibiting the adherent and invasive phenotype. CONCLUSIONS: Up to now, the virulence-like features associated with AIEC are detectable only phenotypically. AIEC genome sequence data will facilitate the identification of genetic determinants implicated in invasion and intracellular growth, as well as enable functional genomic studies of AIEC gene expression during health and disease.

Enterohemorrhagic Escherichia coli O157:H7 gene expression profiling in response to growth in the presence of host epithelia.

BACKGROUND: The pathogenesis of enterohemorrhagic Escherichia coli (EHEC) O157:H7 infection is attributed to virulence factors encoded on multiple pathogenicity islands. Previous studies have shown that EHEC O157:H7 modulates host cell signal transduction cascades, independent of toxins and rearrangement of the cytoskeleton. However, the virulence factors and mechanisms responsible for EHEC-mediated subversion of signal transduction remain to be determined. Therefore, the purpose of this study was to first identify differentially regulated genes in response to EHEC O157:H7 grown in the presence of epithelial cells, compared to growth in the absence of epithelial cells (that is, growth in minimal essential tissue culture medium alone, minimal essential tissue culture medium in the presence of 5% CO(2), and Penassay broth alone) and, second, to identify EHEC virulence factors responsible for pathogen modulation of host cell signal transduction. METHODOLOGY/PRINCIPAL FINDINGS: Overnight cultures of EHEC O157:H7 were incubated for 6 hr at 37 degrees C in the presence or absence of confluent epithelial (HEp-2) cells. Total RNA was then extracted and used for microarray analyses (Affymetrix E. coli Genome 2.0 gene chips). Relative to bacteria grown in each of the other conditions, EHEC O157:H7 cultured in the presence of cultured epithelial cells displayed a distinct gene-expression profile. A 2.0-fold increase in the expression of 71 genes and a 2.0-fold decrease in expression of 60 other genes were identified in EHEC O157:H7 grown in the presence of epithelial cells, compared to bacteria grown in media alone. CONCLUSION/SIGNIFICANCE: Microarray analyses and gene deletion identified a protease on O-island 50, gene Z1787, as a potential virulence factor responsible for mediating EHEC inhibition of the interferon (IFN)-gamma-Jak1,2-STAT-1 signal transduction cascade. Up-regulated genes provide novel targets for use in developing strategies to interrupt the infectious process.

Genomic O island 122, locus for enterocyte effacement, and the evolution of virulent verocytotoxin-producing Escherichia coli.

The locus of enterocyte effacement (LEE) and genomic O island 122 (OI-122) are pathogenicity islands in verocytotoxin-producing Escherichia coli (VTEC) serotypes that are associated with outbreaks and serious disease. Composed of three modules, OI-122 may occur as "complete" (with all three modules) or "incomplete" (with one or two modules) in different strains. OI-122 encodes two non-LEE effector (Nle) molecules that are secreted by the LEE type III secretion system, and LEE and OI-122 are cointegrated in some VTEC strains. Thus, they are functionally linked, but little is known about the patterns of acquisition of these codependent islands. To examine this, we conducted a population genetics analysis, using multilocus sequence typing (MLST), with 72 VTEC strains (classified into seropathotypes A to E) and superimposed on the results the LEE and OI-122 contents of these organisms. The wide distribution of LEE and OI-122 modules among MLST clonal groups corroborates the hypothesis that there has been lateral transfer of both pathogenicity islands. Sequence analysis of a pagC-like gene in OI-122 module 1 also revealed two nonsynonymous single-nucleotide polymorphisms that could help discriminate a subset of seropathotype C strains and determine the presence of the LEE. A nonsense mutation was found in this gene in five less virulent strains, consistent with a decaying or inactive gene. The modular nature of OI-122 could be explained by the acquisition of modules by lateral transfer, either singly or as a group, and by degeneration of genes within modules. Correlations between clonal group, seropathotype, and LEE and OI-122 content provide insight into the role of genomic islands in VTEC evolution.

Molecular analysis as an aid to assess the public health risk of non-O157 Shiga toxin-producing Escherichia coli strains.

Shiga toxin-producing Escherichia coli (STEC) strains are commensal bacteria in cattle with high potential for environmental and zoonotic transmission to humans. Although O157:H7 is the most common STEC serotype, there is growing concern over the emergence of more than 200 highly virulent non-O157 STEC serotypes that are globally distributed, several of which are associated with outbreaks and/or severe human illness such as hemolytic-uremic syndrome (HUS) and hemorrhagic colitis. At present, the underlying genetic basis of virulence potential in non-O157 STEC is unknown, although horizontal gene transfer and the acquisition of new pathogenicity islands are an expected origin. We used seropathotype classification as a framework to identify genetic elements that distinguish non-O157 STEC strains posing a serious risk to humans from STEC strains that are not associated with severe and epidemic disease. We report the identification of three genomic islands encoding non-LEE effector (nle) genes and 14 individual nle genes in non-O157 STEC strains that correlate independently with outbreak and HUS potential in humans. The implications for transmissible zoonotic spread and public health are discussed. These results and methods offer a molecular risk assessment strategy to rapidly recognize and respond to non-O157 STEC strains from environmental and animal sources that might pose serious public health risks to humans.

Applicability of phylogenetic methods for characterizing the public health significance of verocytotoxin-producing Escherichia coli strains.

Two phylogenetic methods (multilocus sequence typing [MLST] and a multiplex PCR) were investigated to determine whether phylogenetic classification of verocytotoxin-producing Escherichia coli serotypes correlates with their classification into groups (seropathotypes A to E) based on their relative incidence in human disease and on their association with outbreaks and serious complications. MLST was able to separate 96% of seropathotype D and E serotypes from those that cause serious disease (seropathotypes A to C), whereas the multiplex PCR lacked this level of seropathotype discrimination.