| 1. |
- Cheng, Liqin, et al.
(author)
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The protective role of commensal gut microbes and their metabolites against bacterial pathogens
- 2024
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In: Gut microbes. - : Taylor & Francis. - 1949-0976 .- 1949-0984. ; 16:1
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Journal article (peer-reviewed)abstract
- Multidrug-resistant microorganisms have become a major public health concern around the world. The gut microbiome is a gold mine for bioactive compounds that protect the human body from pathogens. We used a multi-omics approach that integrated whole-genome sequencing (WGS) of 74 commensal gut microbiome isolates with metabolome analysis to discover their metabolic interaction with Salmonella and other antibiotic-resistant pathogens. We evaluated differences in the functional potential of these selected isolates based on WGS annotation profiles. Furthermore, the top altered metabolites in co-culture supernatants of selected commensal gut microbiome isolates were identified including a series of dipeptides and examined for their ability to prevent the growth of various antibiotic-resistant bacteria. Our results provide compelling evidence that the gut microbiome produces metabolites, including the compound class of dipeptides that can potentially be applied for anti-infection medication, especially against antibiotic-resistant pathogens. Our established pipeline for the discovery and validation of bioactive metabolites from the gut microbiome as novel candidates for multidrug-resistant infections represents a new avenue for the discovery of antimicrobial lead structures.
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| 2. |
- Gonzales, Lucia, et al.
(author)
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Prevalence, seasonality and severity of disease of pathogenic Escherichia coli in children with diarrhea in Bolivia.
- 2013
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In: Journal of Medical Microbiology. - : Microbiology Society. - 0022-2615 .- 1473-5644. ; 62:11, s. 1687-1695
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Journal article (peer-reviewed)abstract
- The prevalence of infection caused by different categories of diarrhoeagenic E. coli (DEC) strains, including enteroaggregative (EAEC), enteropathogenic (EPEC), enterotoxigenic (ETEC), enteroinvasive (EIEC) and enterohaemorrhagic (EHEC) E. coli, in children who suffered from diarrhoea (n=3943) or did not have diarrhoea (n=1026) were analysed in two areas in Bolivia over a period of 4 years. We also analysed the seasonality of DEC infections and severity of diarrhoea in children with DEC infection and compared antibiotic resistance in DEC strains isolated from children with and without diarrhoea. Stool samples were analysed for the presence of DEC by culturing followed by PCR. The most prevalent DEC categories in samples from the children were: EAEC (11.2%); ETEC (6.6%); EPEC (5.8%); and EIEC and EHEC (<1%). DEC strains were isolated significantly more often from diarrhoea cases (21.6%) than from controls (17.6%; P=0.002). The number of children with diarrhoea associated with EAEC, EPEC and ETEC infections peaked in the Bolivian winter (April–September), although the proportion of DEC-positive stool samples was higher during the warm rainy season (October–March). High levels of antibiotic resistance were detected among the DEC strains. In particular, resistance to tetracycline and sulfamethoxazole–trimethoprim was significantly higher in strains isolated from individuals with diarrhoea than in samples from controls. The severity of disease in children infected with EAEC, EPEC and ETEC varied from mild to severe diarrhoea, although disease severity did not differ significantly between the different DEC categories. ETEC, EPEC and EAEC are commonly found in Bolivia and may cause severe disease in children.
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| 3. |
- Joffre, Enrique, et al.
(author)
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Allele Variants of Enterotoxigenic Escherichia coli Heat-Labile Toxin Are Globally Transmitted and Associated with Colonization Factors
- 2015
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In: Journal of Bacteriology. - : American Society for Microbiology. - 0021-9193 .- 1098-5530. ; 197:2, s. 392-403
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Journal article (peer-reviewed)abstract
- Enterotoxigenic Escherichia coli (ETEC) is a significant cause of morbidity and mortality in the developing world. ETEC-mediated diarrhea is orchestrated by heat-labile toxin (LT) and heat-stable toxins (STp and STh), acting in concert with a repertoire of more than 25 colonization factors (CFs). LT, the major virulence factor, induces fluid secretion after delivery of a monomeric ADP-ribosylase (LTA) and its pentameric carrier B subunit (LTB). A study of ETEC isolates from humans in Brazil reported the existence of natural LT variants. In the present study, analysis of predicted amino acid sequences showed that the LT amino acid polymorphisms are associated with a geographically and temporally diverse set of 192 clinical ETEC strains and identified 12 novel LT variants. Twenty distinct LT amino acid variants were observed in the globally distributed strains, and phylogenetic analysis showed these to be associated with different CF profiles. Notably, the most prevalent LT1 allele variants were correlated with major ETEC lineages expressing CS1 + CS3 or CS2 + CS3, and the most prevalent LT2 allele variants were correlated with major ETEC lineages expressing CS5 + CS6 or CFA/I. LTB allele variants generally exhibited more-stringent amino acid sequence conservation (2 substitutions identified) than LTA allele variants (22 substitutions identified). The functional impact of LT1 and LT2 polymorphisms on virulence was investigated by measuring total-toxin production, secretion, and stability using GM1-enzyme-linked immunosorbent assays (GM1-ELISA) and in silico protein modeling. Our data show that LT2 strains produce 5-fold more toxin than LT1 strains (P < 0.001), which may suggest greater virulence potential for this genetic variant. Our data suggest that functionally distinct LT-CF variants with increased fitness have persisted during the evolution of ETEC and have spread globally.
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| 4. |
- Joffré, Enrique, et al.
(author)
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Analysis of Growth Phases of Enterotoxigenic Escherichia coli Reveals a Distinct Transition Phase before Entry into Early Stationary Phase with Shifts in Tryptophan, Fucose, and Putrescine Metabolism and Degradation of Neurotransmitter Precursors
- 2022
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In: Microbiology Spectrum. - : American Society for Microbiology. - 2165-0497. ; 10:4
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Journal article (peer-reviewed)abstract
- Enterotoxigenic Escherichia coli (ETEC) is a major cause of diarrhea in children and adults in endemic areas. Gene regulation of ETEC during growth in vitro and in vivo needs to be further evaluated, and here we describe the full transcriptome and metabolome of ETEC during growth from mid-logarithmic growth to early stationary phase in rich medium (LB medium). We identified specific genes and pathways subjected to rapid transient alterations in gene expression and metabolite production during the transition from logarithmic to stationary growth. The transient phase was found to be different from the subsequent induction of early stationary phase-induced genes. The transient phase was characterized by the repression of genes and metabolites involved in organic substance transport. Genes involved in fucose and putrescine metabolism were upregulated, and genes involved in iron transport were repressed. Expression of toxins and colonization factors were not changed, suggesting retained virulence from mid-logarithmic to the start of the stationary phase. Metabolomic analyses showed that the transient phase was characterized by a drop of intracellular amino acids, e.g., l-tyrosine, l-tryptophan, l-phenylalanine, l-leucine, and l-glutamic acid, followed by increased levels at induction of stationary phase. A pathway enrichment analysis of the entire combined transcriptome and metabolome revealed that significant pathways during progression from logarithmic to early stationary phase are involved in the degradation of neurotransmitters aminobutyrate (GABA) and precursors of 5-hydroxytryptamine (serotonin). This work provides a comprehensive framework for further studies on transcriptional and metabolic regulation in pathogenic E. coli.IMPORTANCE We show that E. coli, exemplified by the pathogenic subspecies enterotoxigenic E. coli (ETEC), undergoes a stepwise transcriptional and metabolic transition into the stationary phase. At a specific entry point, E. coli induces activation and repression of specific pathways. This leads to a rapid decrease of intracellular levels of certain amino acids. The resulting metabolic activity leads to an intense but short peak of indole production, suggesting that this is the previously described “indole peak,” rapid decrease of intermediate molecules of bacterial neurotransmitters, increased putrescine and fucose uptake, increased glutathione levels, and decreased iron uptake. This specific transient shift in gene expression and metabolome is short-lived and disappears when bacteria enter the early stationary phase. We suggest that these changes mainly prepare bacteria for ceased growth, but based on the pathways involved, we could suggest that this transient phase substantially influences survival and virulence.
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| 5. |
- Joffre, Enrique
(author)
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Genetic diversity of the heat labile (LT) and heat stable (ST) toxins of human enterotoxigenic Escherichia coli (ETEC): New insights into polymorphism, regulation, and gene transcription
- 2015
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Doctoral thesis (other academic/artistic)abstract
- Infection with enterotoxigenic Escherichia coli (ETEC) is a leading cause of diarrhea in children in developing countries and travelers to endemic regions. ETEC is a diverse pathogen, with a wide range of virulence factors including enterotoxins and more than 25 identified colonization factors (CFs). ETEC infection causes varying symptoms (mild to profuse, watery, cholera-like diarrhea) as a result of the colonization of the small intestine via CFs, secretion of heat labile (LT) and/or heat stable enterotoxins (STp and STh). To expand the knowledge about the complexity of ETEC pathogenesis we studied the genetic diversity of the LT and ST toxins, using a clinical ETEC strains collection isolated worldwide during three decades. By genomic sequencing we found high diversity in the toxin amino acid sequences, especially in LT where 20 amino acid variants were identified. The LTA subunit was highly polymorphic while the LTB subunit was more conserved. The most common LT variants were LT1 and LT2. ST was less heterogeneous, including 3 ST alleles found in STp and 3 in STh. Phylogenetic analysis of the toxins revealed worldwide distribution of the different variants, and an association with specific CF profiles. The most frequent toxin variants belonged to ETEC linages that have disseminated globally over decades. We also found that main variants differed in ability to produce and secrete the toxins. The STp variant STa5 was linked to disease in adults while the STh variant STa3/4 was associated with disease in children. The gene expression levels of LT (eltAB), and ST (estA) were analyzed by qPCR. We found significantly lower levels of eltAB in presence of glucose in LT1 strains. No polymorphisms were found at the CRP binding sites at eltAB promoter. ST alleles were also significantly downregulated by glucose while bile supplementation favored STp expression. Finally, we performed an RNA-transcriptome study, which showed a dramatic change in global gene expression at the onset of stationary phase. During a specific transient phase we observed up- and down-regulation of genes involved in mechanisms related to virulence, such as biofilm formation, indole induction, iron uptake, fucose catabolism, and the putrescine pathway. The expression levels of the toxins and CFs remained high during this phase. Altogether, this study highlights the diversity within the ETEC population and its virulence factors. We propose that certain combinations of virulence genes influence strain specific responses to host factors that may impact the pathogenesis and severity of ETEC infections.
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| 6. |
- Joffre, Enrique, et al.
(author)
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Identification of new heat-stable (STa) enterotoxin allele variants produced by human enterotoxigenic Escherichia coli (ETEC)
- 2016
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In: International Journal of Medical Microbiology. - : Elsevier BV. - 1438-4221 .- 1618-0607. ; 306:7, s. 586-594
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Journal article (peer-reviewed)abstract
- We describe natural variants of the heat stable toxin (STa) produced by enterotoxigenic Escherichia coli (ETEC) isolates collected worldwide. Previous studies of ETEC isolated from human diarrheal cases have reported the existence of three natural STa gene variants estA1, estA2 and estA3/4 where the first variant encodes STp (porcine, bovine, and human origin) and the two latter ones encode STh (human origin). We identified STa sequences by BLASTn and profiled ST amino acid polymorphisms in a collection of 118 clinical ETEC isolates from children and adults from Asia, Africa and, Latin America that were characterized by whole genome sequencing. Three novel variants of STp and STh were found and designated STa5 and STa6, and STa7, respectively. Presence of glucose significantly decreased the production of STh and STp toxin variants (p < 0.05) as well as downregulated the gene expression (STh: p < 0.001, STp: p < 0.05). We found that the ETEC isolates producing the most common STp variant, STa5, co-expressed coli surface antigen CS6 and was significantly associated with disease in adults in this data set (p < 0.001). Expression of mature STa5 peptide as well as gene expression of tolC, involved in ST secretion, increased in response to bile (p < 0.05). ETEC expressing the common STh variant STa3/4 was associated with disease in children (p < 0.05). The crp gene, that positively regulate estA3/4 encoding STa3/4, and estA3/4 itself had decreased transcriptional levels in presence of bile. Since bile levels in the intestine are lower in children than adults, these results may suggest differences in pathogenicity of ETEC in children and adult populations. (C) 2016 The Author(s). Published by Elsevier GmbH.
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| 7. |
- Joffre, Enrique, et al.
(author)
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The LT1 and LT2 variants of the enterotoxigenic Escherichia coli (ETEC) heat-labile toxin (LT) are associated with major ETEC lineages
- 2016
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In: Gut microbes. - : Informa UK Limited. - 1949-0976 .- 1949-0984. ; 7:1, s. 75-81
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Journal article (peer-reviewed)abstract
- The heat-labile toxin (LT) is one of the major virulence factors of enterotoxigenic Escherichia coli (ETEC). We recently described that 20 polymorphic LT variants are present in ETEC strains isolated globally. Two of the variants, LT1 and LT2, are particularly common and we found that they were associated with clonal ETEC lineages that express the colonization factors (CFs), CFA/I, CS1+CS3, CS2+CS3, and CS5+CS6. ETEC expressing these CFs are frequently found among ETEC strains isolated from cases with diarrhea. ETEC expressing the colonization factors CS1+CS3, and CS2+CS3 are found in 2 discrete clonal lineages and express the LT1 variant and heat stable toxin (STh). Although they clearly are virulent they neither produce, nor secrete, high amounts of LT toxin. On the other hand ETEC strains expressing LT, STh, CFA/I and LT, STh, CS5+CS6, carry the LT2 variant and produce and secrete significantly more LT toxin. Despite differences in toxin production, LT1 and LT2 are found in ETEC lineages that have managed to spread globally confirming that these variants are important for ETEC virulence. © 2016 The Author(s). Published with license by Taylor & Francis Group, LLC.
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| 8. |
- Toledo, Carla Calderon, et al.
(author)
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Circulation of enterotoxigenic Escherichia coli (ETEC) isolates expressing CS23 from the environment to clinical settings
- 2023
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In: mSystems. - 2379-5077. ; 8:5
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Journal article (peer-reviewed)abstract
- Enterotoxigenic Escherichia coli (ETEC) is one of the leading causes of infant diarrhea in low- and middle-income countries (LMICs). Diarrheal pathogens are transmitted through environmental reservoirs; however, the bacterial clones that spread across the human-environment interface remain unexplored. We aimed to determine the relationship and clonal dissemination of ETEC between children with diarrhea and polluted water samples from a local river in La Paz, Bolivia. By using WGS and the PhenePlates phenotypic system to analyze ETEC strains, we showed that ST218 and ST410 LT+STh ETEC expressing the colonization factor (CF) CS23 were found with high frequency in both samples. The CS23 ETEC isolates were found within several STs, E. coli phylogroups, and across ETEC lineages. Comparative genomic evaluation and PhenePlate screening of globally distributed clinical ETEC strains suggest that the CS23 gene is likely carried on plasmids acquired independently of the bacterial chromosomal background. Clinical strains were more often multidrug-resistant (MDR) than environmental isolates and harbored the class 1 integron-integrase gene intI1 next to the MDR cassettes. Retrospective analysis of antibiotic resistance in ETEC revealed a high frequency of MDR in clinical isolates. The LT+STh CS23 environmental ETEC isolates, showed an increased biofilmability at environmental temperature, equal cytotoxicity, and significantlylower adherence to human epithelial cells compared to ETEC expressing other CFs. Together, we suggest that CS23 is more prevalent in ETEC than previously estimated, and the Choqueyapu River is a reservoir for LT+STh CS23 ETEC containing strains capable of causing diarrheal cases in children.
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| 9. |
- von Mentzer, Astrid, 1983, et al.
(author)
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Identification of enterotoxigenic Escherichia coli (ETEC) clades with long-term global distribution
- 2014
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In: Nature Genetics. - : Springer Science and Business Media LLC. - 1061-4036 .- 1546-1718. ; 46:12, s. 1321-1326
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Journal article (peer-reviewed)abstract
- Enterotoxigenic Escherichia coil (ETEC), a major cause of infectious diarrhea, produce heat-stable and/or heat-labile enterotoxins and at least 25 different colonization factors that target the intestinal mucosa. The genes encoding the enterotoxins and most of the colonization factors are located on plasmids found across diverse E. coli serogroups. Whole-genome sequencing of a representative collection of ETEC isolated between 1980 and 2011 identified globally distributed lineages characterized by distinct colonization factor and enterotoxin profiles. Contrary to current notions, these relatively recently emerged lineages might harbor chromosome and plasmid combinations that optimize fitness and transmissibility. These data have implications for understanding, tracking and possibly preventing ETEC disease.
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