James P Nataro

University of Virginia, Charlottesville, Virginia, United States

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Publications (276)1257.28 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Molecular identification of the invasion plasmid antigen-H (ipaH) gene has been established as a useful detection mechanism for Shigella spp. The Global Enteric Multicenter Study (GEMS) identified the etiology and burden of moderate-to-severe diarrhea (MSD) in sub-Saharan Africa and south Asia using a case-control study and traditional culture techniques. Here, we used quantitative polymerase chain reaction (qPCR) to identify Shigella spp. in 2,611 stool specimens from GEMS and compared these results to those using culture. Demographic and nutritional characteristics were assessed as possible risk factors. The qPCR identified more cases of shigellosis than culture; however, the distribution of demographic characteristics was similar by both methods. In regression models adjusting for Shigella quantity, age, and site, children who were exclusively breast-fed had significantly lower odds of MSD compared with children who were not breast-fed (odds ratio [OR] = 0.47, 95% confidence interval (CI) = 0.28-0.81). The association between Shigella quantity and MSD increased with age, with a peak in children of 24-35 months of age (OR = 8.2, 95% CI = 4.3-15.7) and the relationship between Shigella quantity and disease was greatest in Bangladesh (OR = 13.2, 95% CI = 7.3-23.8). This study found that qPCR identified more cases of Shigella and age, site, and breast-feeding status were significant risk factors for MSD. © The American Society of Tropical Medicine and Hygiene.
    The American journal of tropical medicine and hygiene 08/2015; DOI:10.4269/ajtmh.14-0319 · 2.70 Impact Factor
  • Tracy H Hazen · James B Kaper · James P Nataro · David A Rasko ·
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    ABSTRACT: The attaching and effacing Escherichia coli (AEEC) are a genomically diverse group of diarrheagenic E. coli that are characterized by the presence of the locus of enterocyte effacement (LEE) genomic island, which encodes a type III secretion system that is essential to virulence. The AEEC can be further classified as either enterohemorrhagic E. coli (EHEC), typical enteropathogenic E. coli (EPEC), or atypical EPEC depending on the presence or absence of the Shiga-toxin genes or bundle-forming pilus (BFP) genes. Recent AEEC genomic studies have focused on the diversity of the core genome and less is known regarding the genetic diversity and relatedness of AEEC plasmids. Comparative genomic analyses in this study demonstrate there is genetic similarity among AEEC plasmid genes involved in plasmid replication conjugative transfer, and maintenance while the remainder of the plasmids had sequence variability. Investigation of the EPEC adherence factor (EAF) plasmids, which encodes the BFP genes demonstrated significant plasmid diversity even among isolates within the same phylogenomic lineage, suggesting that these EAF-like plasmids have undergone genetic modifications, or have been lost and acquired multiple times. Global transcriptional analyses of the EPEC prototype isolate E2348/69 and two EAF plasmid mutants of this isolate demonstrated that the plasmid genes influence the expression of a number of chromosomal genes in addition to the LEE. This suggests that the genetic diversity of the EAF plasmids could contribute to differences in the global virulence regulons of EPEC isolates. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
    Infection and immunity 08/2015; 83(10). DOI:10.1128/IAI.00769-15 · 3.73 Impact Factor
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    ABSTRACT: In 2011, a Shiga toxin (Stx) type 2a-producing enteroaggregative E. coli (EAEC) strain of serotype O104:H4 caused a large lethal outbreak in Northern Europe. Until recently, the pathogenic mechanisms explaining the high virulence of the strain have remained unclear. Our laboratories have shown that EAEC genes encoded on the pAA virulence plasmid, particularly the AggR-regulated AAF/I fimbriae, enhance inflammation and enable the outbreak strain to both adhere to epithelial cells and translocate Stx2a across the intestinal epithelium, possibly explaining the high incidence of the life threatening post-diarrheal sequelae of hemolytic uremic syndrome. Epidemiologic evidence supports a model of EAEC pathogenesis comprising the concerted action of multiple virulence factors along with induction of inflammation. Here, we suggest a model for the pathogenesis of the O104:H4 outbreak strain that includes contributions from EAEC alone, but incorporating additional injury induced by Stx2a.
    Gut Microbes 06/2015; 6(4). DOI:10.1080/19490976.2015.1054591
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    ABSTRACT: Typical enteropathogenic Escherichia coli (tEPEC) strains were associated with mortality in the Global Enteric Multicenter Study (GEMS). Genetic differences in tEPEC strains could underlie some of the variability in clinical outcome. We produced draft genome sequences of all available tEPEC strains from GEMS lethal infections (LIs) and of closely matched EPEC strains from GEMS subjects with non-lethal symptomatic infections (NSIs) and asymptomatic infections (AIs) to identify gene clusters (potential protein encoding sequences sharing ≥90% nucleotide sequence identity) associated with lethality. Among 14,412 gene clusters identified, the presence or absence of 392 was associated with clinical outcome. As expected, more gene clusters were associated with LI versus AI than LI versus NSI. The gene clusters more prevalent in strains from LI than those from NSI and AI included those encoding proteins involved in O-antigen biogenesis, while clusters encoding type 3 secretion effectors EspJ and OspB were among those more prevalent in strains from non-lethal infections. One gene cluster encoding a variant of an NleG ubiquitin ligase was associated with LI versus AI, while two other nleG clusters had the opposite association. Similar associations were found for two nleG gene clusters in an additional, larger sample of NSI and AI GEMS strains. Particular genes are associated with lethal tEPEC infections. Further study of these factors holds potential to unravel the mechanisms underlying severe disease and to prevent adverse outcomes.
    PLoS Neglected Tropical Diseases 05/2015; 9(5):e0003791. DOI:10.1371/journal.pntd.0003791 · 4.45 Impact Factor
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    ABSTRACT: Pathogens in the gastrointestinal tract exist within a vast population of microbes. We examined associations between pathogens and composition of gut microbiota as they relate to Shigella spp./enteroinvasive Escherichia coli infection. We analyzed 3,035 stool specimens (1,735 nondiarrheal and 1,300 moderate-to-severe diarrheal) from the Global Enteric Multicenter Study for 9 enteropathogens. Diarrheal specimens had a higher number of enteropathogens (diarrheal mean 1.4, nondiarrheal mean 0.95; p<0.0001). Rotavirus showed a negative association with Shigella spp. in cases of diarrhea (odds ratio 0.31, 95% CI 0.17-0.55) and had a large combined effect on moderate-to-severe diarrhea (odds ratio 29, 95% CI 3.8-220). In 4 Lactobacillus taxa identified by 16S rRNA gene sequencing, the association between pathogen and disease was decreased, which is consistent with the possibility that Lactobacillus spp. are protective against Shigella spp.-induced diarrhea. Bacterial diversity of gut microbiota was associated with diarrhea status, not high levels of the Shigella spp. ipaH gene.
    Emerging infectious diseases 02/2015; 21(2):242-50. DOI:10.3201/eid2101.140795 · 6.75 Impact Factor
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    ABSTRACT: Enteroaggregative Escherichia coli (EAEC) belong to a diarrheagenic pathotype known to cause diarrhea and can be characterized by distinct aggregative adherence (AA) in a stacked brick pattern to cultured epithelial cells. In this study, we investigated 118 EAEC strains isolated from the stools of Danish adults with traveler's diarrhea. We evaluated the presence of the aggregative adherence fimbriae (AAFs) by a multiplex PCR, targeting the four known major subunit variants as well as their usher-encoding genes. Almost half (49/118) of the clinical isolates did not possess any known AAF major fimbrial subunit, despite the presence of other AggR-related loci. Further investigation revealed the presence of AAF-related gene encoding a yet uncharacterized adhesin termed agg5A. The sequence of agg5DCBA gene cluster shared fimbrial accessory genes (usher, chaperone and minor pilin subunit) with AAF/III, as well as the signal peptide present in the beginning of the agg3A encoding gene. The complete agg5DCBA gene cluster from a clinical isolate EAEC strain C338-14 with typical stacked brick binding pattern was cloned and deletion of the cluster was performed. Transformation to a non- adherent E. coli HB101 and complementation of the non-adherent C338-14 mutant with the complete gene cluster restored the AA adhesion. Overall, we found the agg5A encoding gene in 12 % of the 118 strains isolated from Denmark, suggesting that this novel adhesin represents an important variant. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
    Infection and Immunity 01/2015; 83(4). DOI:10.1128/IAI.02820-14 · 3.73 Impact Factor
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    ABSTRACT: Shigellae cause significant diarrheal disease and mortality in humans, as there are approximately 163 million episodes of shigellosis and 1.1 million deaths annually. While significant strides have been made in the understanding of the pathogenesis, few studies on the genomic content of the Shigella species have been completed. The goal of this study was to characterize the genomic diversity of Shigella species through sequencing of 55 isolates representing members of each of the four Shigella species: S. flexneri, S. sonnei, S. boydii, and S. dysenteriae. Phylogeny inferred from 336 available Shigella and E. coli genomes defined exclusive clades of Shigella; conserved genomic markers were then identified that can identify each clade. Polymerase chain reaction (PCR) assays were developed for each clade-specific marker, which was combined with an amplicon for the conserved Shigella invasion antigen, IpaH3, into a multiplex PCR assay. This assay demonstrated high specificity, correctly identifying 218 of 221 presumptive Shigella isolates, and sensitivity, by not identifying any of 151 diverse E. coli isolates incorrectly as Shigella. This new phylogenomic-based PCR assay represents a valuable tool for rapid typing of uncharacterized Shigella isolates and provides a framework that can be utilized for the identification of novel genomic markers from genomic data. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
    Journal of Clinical Microbiology 01/2015; 53(3). DOI:10.1128/JCM.03527-14 · 3.99 Impact Factor
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    ABSTRACT: Diarrhea causes substantial morbidity and mortality in children in low-income countries. Although numerous pathogens cause diarrhea, the etiology of many episodes remains unknown. Serratia marcescens is incriminated in hospital-associated infections, and HIV/AIDS associated diarrhea. We have recently found that Serratia spp. may be found more commonly in the stools of patients with diarrhea than in asymptomatic control children. We therefore investigated the possible enteric pathogenicity of S. marcescens in vitro employing a polarized human colonic epithelial cell (T84) monolayer. Infected monolayers were assayed for bacterial invasion, transepithelial electrical resistance (TEER), cytotoxicity, interleukin-8 (IL-8) release and morphological changes by scanning electron microscopy. We observed significantly greater epithelial cell invasion by S. marcescens compared to Escherichia coli strain HS (p = 0.0038 respectively). Cell invasion was accompanied by reduction in TEER and secretion of IL-8. Lactate dehydrogenase (LDH) extracellular concentration rapidly increased within a few hours of exposure of the monolayer to S. marcescens. Scanning electron microscopy of S. marcescens-infected monolayers demonstrated destruction of microvilli and vacuolization. Our results suggest that S. marcescens interacts with intestinal epithelial cells in culture and induces dramatic alterations similar to those produced by known enteric pathogens.
    Gut Microbes 11/2014; 5(6). DOI:10.4161/19490976.2014.972223
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    ABSTRACT: Background Group A Rotaviruses are a major etiologic agent of gastroenteritis in infants and young children (<5 years) worldwide. Although rotavirus vaccines have been successfully administered in many countries, in India the introduction of rotavirus vaccine in national immunization program was approved in 2014. Since high disease burden and large number of genetic variants have been reported from low income countries including India, monitoring of rotavirus was initiated prior to implementation of the vaccine in the region. Methods A total number of 3,582 stool samples were collected from an urban slum community in Kolkata, among which 1,568 samples were obtained from children of ≤5 years of age, with moderate to severe diarrhoea and 2,014 samples were collected from age-sex matched healthy neighbourhood controls. Rotavirus positive samples were typed by multiplex semi-nested PCR and nucleotide sequencing. Circulating strains were phylogenetically analyzed. Results Among 1,568 children with diarrhoea, 395 (25.2%), and among 2,014 asymptomatic children, 42 (2%) were rotavirus positive. G1P[8] was identified as the most common strain (32%) followed by G9P[8] (16.9%), G2P[4] (13.5%) and G9P[4] (10.75%). G12 strains with combinations of P[4], P[6] and P[8] comprised 11.9% of total positive strains. The rest (<10%) were rare and uncommon strains like G1P[4], G1P[6], G2P[8] and animal-like strains G4P[6], G6P[14] and G11P[25]. The 42 rotavirus positive samples from asymptomatic children revealed common genotypes like G1, G2 and G9. Conclusion This community based case-control study showed increased predominance of genotype G9 in Kolkata. It also confirmed co-circulation of a large number of genetic variants in the community. Asymptomatic rotavirus positive children though low in number can also be a source of dispersal of infection in the community. This study provides background information to the policy makers for implementation of rotavirus vaccines in this region.
    PLoS ONE 11/2014; 9(11):e112970. DOI:10.1371/journal.pone.0112970 · 3.23 Impact Factor
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    ABSTRACT: The Global Enteric Multicenter Study (GEMS) is a prospective collaborative study to determine the pathogen specific, diarrhea associated attributable disease burden of diarrhea in Africa and Asia in children less than 5 years of age. Using samples from children that participated in this case-control study and were seen at in the GEMS clinics in The Gambia and Kenya, we conducted a host gene candidate study that examined the association of the host DNA single nucleotide polymorphisms (SNPs) and specific bacterial enteropathogens causing diarrhea. Cases of diarrhea were matched with healthy controls from the same area. Stool was collected and examined comprehensively for the presence of enteropathogens and host DNA polymorphisms. We studied de-identified fecal DNA for the presence of SNPs (N=144) in 26 genes that code for host proteins involved in pathogen attachment, inflammation, innate and acquired immune responses to enteropathogens. We analyzed the distribution of host genotypes and compared cases vs. controls according to enteropathogens identified. Comparisons were made using SNPSTATs software following an additive model. A mixed logistic regression model was used to adjust for potential confounding factors including site, age, weight for height and weight for age scores. Microbiological and genotype data were available in 1,164 subjects. Diarrhea was associated with 6 SNPs located in SELPLG, CD55, LPLUNC, IL12 B, and CORO1C. In The Gambia, diarrhea due to Shigella was associated with a SNP in SELPLG, diarrhea from enterotoxigenic E. coli was associated with SNPs in DAF (CD55), LPLUNC, SELPLG, diarrhea from enteroaggregative E. coli was associated with SNPs in CORO1C. In Kenya, diarrhea from enteroaggregative E.coli was associated with SNPs in IL12B and CORO1C Distinct SNPs were associated with pathogen specific diarrhea in children under 5 years of age living in two African countries.
    American Society of Tropical Medicine and Hygiene; 11/2014
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    ABSTRACT: Stunting is common in young children in developing countries, and is associated with increased morbidity, developmental delays, and mortality. Its complex pathogenesis likely involves poor intrauterine and postnatal nutrition, exposure to microbes, and the metabolic consequences of repeated infections. Acquired enteropathy affecting both gut structure and function likely plays a significant role in this outcome, especially in the first few months of life, and serve as a precursor to later interactions of infection and malnutrition. However, the lack of validated clinical diagnostic criteria has limited the ability to study its role, identify causative factors, and determine cost-effective interventions. This review addresses these issues through a historical approach, and provides recommendations to define and validate a working clinical diagnosis and to guide critical research in this area to effectively proceed. Prevention of early gut functional changes and inflammation may preclude or mitigate the later adverse vicious cycle of malnutrition and infection.
    Clinical Infectious Diseases 11/2014; 59(suppl 4):S207-S212. DOI:10.1093/cid/ciu485 · 8.89 Impact Factor
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    ABSTRACT: We have encountered an Escherichia coli strain isolated from a child with acute diarrhea. This strain harbored eae and elt genes encoding for E. coli attaching and effacing property and heat-labile enterotoxin of EPEC and ETEC, respectively. Due to the presence of these distinct virulence factors, we named this uncommon strain as EPEC/ETEC hybrid. The elt gene was identified in a conjugally transferable plasmid of the EPEC/ETEC hybrid. In addition, several virulence genes in the locus of enterocyte effacement have been identified, which confirms that the EPEC/ETEC has an EPEC genetic background. The hybrid nature of this strain was further confirmed by using tissue culture assays. In the multi locus sequence typing (MLST) analysis, the EPEC/ETEC belonged to the sequence type ST328 and was belonging to ST278 Cplx. Sequence analysis of the plasmid DNA revealed presence of six large contigs with several insertion sequences. A phage integrase gene and the prophages of gp48 and gp49 have been found in the upstream of eltAB. In the downstream of elt, an urovirulence loci adhesion encoding (pap) cluster containing papG, and papC were also identified. Similar to other reports, we have identified a heterogenic virulence in a diarrheagenic E. coli but with different combination of genes. Copyright © 2014 Elsevier GmbH. All rights reserved.
    International journal of medical microbiology: IJMM 10/2014; 305(1). DOI:10.1016/j.ijmm.2014.10.006 · 3.61 Impact Factor
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    ABSTRACT: Background: Group A streptococcus (GAS) pharyngitis is associated with high rates of rheumatic heart disease in developing countries. We sought to identify guidelines for empiric treatment of pharyngitis in low-resource settings. To inform the design of GAS vaccines, we determined the emm types associated with pharyngitis among African schoolchildren. Methods: Surveillance for pharyngitis was conducted among children 5-16 years of age attending schools in Bamako, Mali. Students were encouraged to visit a study clinician when they had a sore throat. Enrollees underwent evaluation and throat swab for isolation of GAS. Strains were emm typed by standard methods. Results: GAS was isolated from 449 (25.5%) of the 1,759 sore throat episodes. Painful cervical adenopathy was identified in 403 children (89.8%) with GAS infection and was absent in 369 uninfected children (28.2%). Emm type was determined in 396 (88.2%) of the 449 culture-positive children; 70 types were represented and 14 types accounted for 49% of isolates. Based on the proportion of the 449 isolates bearing emm types included in the 30-valent vaccine (31.0%) plus nonvaccine types previously shown to react to vaccine-induced bactericidal antibodies (44.1%), the vaccine could protect against almost 75% of GAS infections among Bamako schoolchildren. Conclusions: Two promising strategies could reduce rheumatic heart disease in low-resource settings. Administering antibiotics to children with sore throat and tender cervical adenopathy could treat most GAS-positive children while reducing use of unnecessary antibiotics for uninfected children. Broad coverage against M types associated with pharyngitis in Bamako schoolchildren might be achieved with the 30-valent GAS vaccine under development.
    The Pediatric Infectious Disease Journal 10/2014; 34(5). DOI:10.1097/INF.0000000000000608 · 2.72 Impact Factor
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    ABSTRACT: Live attenuated bacteria hold great promise as multivalent mucosal vaccines against a variety of pathogens. A major challenge of this approach has been the successful delivery of sufficient amounts of vaccine antigens to adequately prime the immune system without over-attenuating the live vaccine. Here we have used a live attenuated Salmonella enterica serovar Typhi strain to create a bivalent mucosal plague vaccine that produces both the protective F1 capsular antigen of Yersinia pestis as well as the LcrV protein required for secretion of virulence effector proteins. To reduce metabolic burden associated with the co-expression of F1 and LcrV within the live vector, we balanced expression of both antigens by combining plasmid-based expression of F1 with chromosomal expression of LcrV from three independent loci. The immunogenicity and protective efficacy of this novel vaccine were assessed in mice using a heterologous prime-boost immunization strategy, and compared to a conventional strain in which F1 and LcrV were expressed from a single low copy number plasmid. The serum antibody responses to LPS induced by the optimized bivalent vaccine were indistinguishable from those elicited by the parent strain, suggesting adequate immunogenic capacity maintained through preservation of bacterial fitness; by contrast, LPS titers were 10-fold lower in mice immunized with the conventional vaccine strain. Importantly, mice receiving the optimized bivalent vaccine were fully protected against lethal pulmonary challenge. These results demonstrate the feasibility of distributing foreign antigen expression across both chromosomal and plasmid locations within a single vaccine organism for induction of protective immunity.
    Infection and Immunity 10/2014; 83(1). DOI:10.1128/IAI.02443-14 · 3.73 Impact Factor
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    ABSTRACT: Background: The Global Enteric Multicenter Study (GEMS) is a prospective case control study to determine pathogen specific diarrheal disease burden among children <5 years of age in Africa and Asia. Using samples from children in The Gambia and Kenya, we conducted a gene candidate study that examined the association of the host DNA single nucleotide polymorphisms (SNPs) and specific viral enteropathogens rotavirus and norovirus causing diarrhea. Methods: Stool specimens from moderate to severe diarrhea cases and their age, sex and area matched controls were examined for the presence of enteropathogens and host DNA SNPs (N=144) in 26 genes that code for host proteins involved in pathogen attachment, inflammation, innate and acquired immune responses to enteropathogens. We analyzed the distribution of SNPs and compared cases vs. controls according to enteropathogens identified. Comparisons were made using SNPSTATs software following the dominant model. Logistic regression model was used to adjust for potential confounders including site and age. Results: Microbiological and genotype data were available in 1,164 subjects. In The Gambia, diarrhea due to norovirus was associated with SNP in C3orf23 (69% vs. 36%, OR=4.0, CI=1.1.5-13.7, P=0.03), SNPs in IL12B (67% vs. 32%, OR=4.2, CI=1.19-15, P=0.025) and a SNP in MBL (29% vs. 58%, OR=0.3, CI=0.08-0.9, P=0.045) and diarrhea from rotavirus was associated with SNPs in CD180 (88% vs. 49%, OR=8, CI=1.66-35, P=0.003). In Kenya, diarrhea from norovirus was associated with a SNP in C3orf23 (71% vs. 23%, OR=8, CI=1.4-42, P=0.013), and SNPs in LPLUNC (50% vs.15%, OR=6, CI=1.5-21, P=0.012) and diarrhea from rotavirus was associated with LPLUNC (61% vs. 40%, OR=2.5, CI=1.2-5.3, P=0.011). Conclusion: Distinct SNPs were associated with pathogen specific viral diarrhea in children under 5 years of age living in two African countries.
    IDWeek 2014 Meeting of the Infectious Diseases Society of America; 10/2014
  • James Nataro ·

    IDWeek 2014 Meeting of the Infectious Diseases Society of America; 10/2014
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    ABSTRACT: Autotransporter (AT) proteins provide a diverse array of important virulence functions to Gram-negative bacterial pathogens, and have also been adapted for protein surface display applications. The “autotransporter” moniker refers to early models that depicted these proteins facilitating their own translocation across the bacterial outer membrane. Although translocation is less autonomous than originally proposed, AT protein segments upstream of the C-terminal transmembrane β-barrel have nevertheless consistently been found to contribute to efficient translocation and/or folding of the N-terminal virulence region (the “passenger”). However, defining the precise secretion functions of these AT regions has been complicated by the use of multiple overlapping and ambiguous terms to define AT sequence, structural, and functional features, including “autochaperone”, “linker” and “junction”. Moreover, the precise definitions and boundaries of these features vary among ATs and even among research groups, leading to an overall murky picture of the contributions of specific features to translocation. Here we propose a unified, unambiguous nomenclature for AT structural, functional and conserved sequence features, based on explicit criteria. Applied to 16 well studied AT proteins, this nomenclature reveals new commonalities for translocation but also highlights that the autochaperone function is less closely coupled with a conserved sequence element than previously believed.
    Molecular Microbiology 10/2014; 95(1). DOI:10.1111/mmi.12838 · 4.42 Impact Factor
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    ABSTRACT: Enterotoxigenic Escherichia coli (ETEC) is an important cause of childhood diarrhea in resource-limited regions. It is also an important cause of diarrhea in travellers to these areas. To evaluate the protective efficacy of new ETEC vaccines that are under development, there is a need to increase the capacity to undertake Phase IIB (human challenge) clinical trials and to develop suitable challenge models. An in-hospital study was performed where fasting adult volunteers were experimentally infected with 1 × 106 to 1 × 109 colony forming units (CFUs) of the wild-type ETEC strain TW10598, which had been isolated from a child with diarrhea in West Africa in 1997. We recorded symptoms and physical signs and measured serum immune response to the TW10598 bacterium. We included 30 volunteers with mean age 22.8 (range 19.8, 27.4) years. The most common symptoms were diarrhea (77%), abdominal pain (67%), nausea (63%), and abdominal cramping (53%). Seven subjects (23%) experienced fever, none were hypotensive. Most of the volunteers responded with a substantial rise in the level of serum IgA antibodies against the challenge strain. We established the capacity and methods for safely undertaking challenge studies to measure the efficacy of ETEC vaccine candidates in a hospital ward. Strain TW10598 elicited both clinical symptoms and an immune response across the doses given.
    BMC Infectious Diseases 09/2014; 14(1):482. DOI:10.1186/1471-2334-14-482 · 2.61 Impact Factor
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    ABSTRACT: Enteroaggregative Escherichia coli (EAEC) is a leading cause of acute and persistent diarrhea worldwide. A recently emerged Shiga-toxin-producing strain of EAEC resulted in significant mortality and morbidity due to progressive development of hemolytic-uremic syndrome. The attachment of EAEC to the human intestinal mucosa is mediated by aggregative adherence fimbria (AAF). Using X-ray crystallography and NMR structures, we present new atomic resolution insight into the structure of AAF variant I from the strain that caused the deadly outbreak in Germany in 2011, and AAF variant II from archetype strain 042, and propose a mechanism for AAF-mediated adhesion and biofilm formation. Our work shows that major subunits of AAF assemble into linear polymers by donor strand complementation where a single minor subunit is inserted at the tip of the polymer by accepting the donor strand from the terminal major subunit. Whereas the minor subunits of AAF have a distinct conserved structure, AAF major subunits display large structural differences, affecting the overall pilus architecture. These structures suggest a mechanism for AAF-mediated adhesion and biofilm formation. Binding experiments using wild type and mutant subunits (NMR and SPR) and bacteria (ELISA) revealed that despite the structural differences AAF recognize a common receptor, fibronectin, by employing clusters of basic residues at the junction between subunits in the pilus. We show that AAF-fibronectin attachment is based primarily on electrostatic interactions, a mechanism not reported previously for bacterial adhesion to biotic surfaces.
    PLoS Pathogens 09/2014; 10(9):e1004404. DOI:10.1371/journal.ppat.1004404 · 7.56 Impact Factor
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    ABSTRACT: Adherence to the intestinal epithelia is a key feature in enteroaggregative Escherichia coli (EAEC) infection. The aggregative adherence fimbriae (AAFs) are involved in EAEC interaction with receptors at the surface of intestinal cells. We and others have demonstrated that fibronectin is a receptor for AAF/II fimbriae. Considering that the major cellular receptor of fibronectin is integrin α5β1, in this study we evaluated the participation of this receptor in the fibronectin-mediated adherence of EAEC strain 042 to intestinal cells. We found that EAEC strain 042 has the ability to bind directly and indirectly to integrin α5β1; direct binding was not mediated by AAF/II fimbriae and indirect binding was mediated by AAF/II and fibronectin. Coimmunoprecipitation assays confirmed the formation of the complex AafA/fibronectin/integrin α5β1. To evaluate EAEC adherence to intestinal cells, T84 cells were incubated with fibronectin and an antibody that blocks the interaction region of integrin α5β1 to fibronectin, the RGD site. Under these conditions, we found the number of adherent bacteria to epithelial cells significantly reduced. Additionally, fibronectin-mediated adherence of EAEC strain 042 was abolished in HEp-2 cells transfected with integrin α5 shRNA. Altogether, our data support the involvement of integrin α5β1 in the fibronectin-mediated EAEC binding to intestinal cells.
    BioMed Research International 08/2014; 2014:781246. DOI:10.1155/2014/781246 · 1.58 Impact Factor

Publication Stats

18k Citations
1,257.28 Total Impact Points


  • 2000-2015
    • University of Virginia
      • Division of Pediatrics
      Charlottesville, Virginia, United States
  • 1985-2014
    • University of Maryland, Baltimore
      • • Center for Vaccine Development
      • • Department of Pediatrics
      • • Department of Microbiology and Immunology
      • • Department of Medicine
      • • Division of Infectious Diseases
      Baltimore, Maryland, United States
  • 2008-2012
    • Statens Serum Institut
      • Department of Microbiology and Infection Control
      København, Capital Region, Denmark
  • 2010-2011
    • University of Bergen
      • • Centre for International Health
      • • Department of Molecular Biology
      Bergen, Hordaland, Norway
      Akra, Greater Accra, Ghana
  • 2009-2011
    • Kansas State University
      • Department of Diagnostic Medicine/Pathobiology
      Kansas, United States
  • 2007-2009
    • University of Birmingham
      • School of Immunity and Infection
      Birmingham, England, United Kingdom
  • 2006
    • Malaria Vaccine Development Program
      New Dilli, NCT, India
  • 2005
    • Universidade Federal de São Paulo
      San Paulo, São Paulo, Brazil
  • 2003
    • University of Cambridge
      Cambridge, England, United Kingdom
  • 2002
    • University of Zurich
      Zürich, Zurich, Switzerland
    • University of Kansas
      Lawrence, Kansas, United States
  • 2001-2002
    • Johns Hopkins University
      • Department of Medicine
      Baltimore, Maryland, United States
    • Pasteur Institute of Iran (IPI)
      • Molecular Biology Department
      Teheran, Tehrān, Iran
    • Queen's University Belfast
      Béal Feirste, Northern Ireland, United Kingdom
  • 1999
    • Pennsylvania State University
      • Department of Biology
      University Park, Maryland, United States
  • 1998
    • University of Cincinnati
      Cincinnati, Ohio, United States
    • Imperial College London
      Londinium, England, United Kingdom
  • 1995-1997
    • Universidade Federal do Ceará
      Ceará, Ceará, Brazil
  • 1987
    • University of Santiago, Chile
      CiudadSantiago, Santiago Metropolitan, Chile