Topological data analysis of Escherichia coli O157:H7 and non-O157 survival in soils.

Shiga toxin-producing E. coli O157:H7 and non-O157 have been implicated in many foodborne illnesses caused by the consumption of contaminated fresh produce. However, data on their persistence in soils are limited due to the complexity in datasets generated from different environmental variables and bacterial taxa. There is a continuing need to distinguish the various environmental variables and different bacterial groups to understand the relationships among these factors and the pathogen survival. Using an approach called Topological Data Analysis (TDA); we reconstructed the relationship structure of E. coli O157 and non-O157 survival in 32 soils (16 organic and 16 conventionally managed soils) from California (CA) and Arizona (AZ) with a multi-resolution output. In our study, we took a community approach based on total soil microbiome to study community level survival and examining the network of the community as a whole and the relationship between its topology and biological processes. TDA produces a geometric representation of complex data sets. Network analysis showed that Shiga toxin negative strain E. coli O157:H7 4554 survived significantly longer in comparison to E. coli O157:H7 EDL 933, while the survival time of E. coli O157:NM was comparable to that of E. coli O157:H7 EDL 933 in all of the tested soils. Two non-O157 strains, E. coli O26:H11 and E. coli O103:H2 survived much longer than E. coli O91:H21 and the three strains of E. coli O157. We show that there are complex interactions between E. coli strain survival, microbial community structures, and soil parameters

Complete Genome Sequence and Comparative Metabolic Profiling of the Prototypical Enteroaggregative Escherichia coli Strain 042

Background \ud Escherichia coli can experience a multifaceted life, in some cases acting as a commensal while in other cases causing intestinal and/or extraintestinal disease. Several studies suggest enteroaggregative E. coli are the predominant cause of E. coli-mediated diarrhea in the developed world and are second only to Campylobacter sp. as a cause of bacterial-mediated diarrhea. Furthermore, enteroaggregative E. coli are a predominant cause of persistent diarrhea in the developing world where infection has been associated with malnourishment and growth retardation. \ud \ud Methods \ud In this study we determined the complete genomic sequence of E. coli 042, the prototypical member of the enteroaggregative E. coli, which has been shown to cause disease in volunteer studies. We performed genomic and phylogenetic comparisons with other E. coli strains revealing previously uncharacterised virulence factors including a variety of secreted proteins and a capsular polysaccharide biosynthetic locus. In addition, by using Biolog™ Phenotype Microarrays we have provided a full metabolic profiling of E. coli 042 and the non-pathogenic lab strain E. coli K-12. We have highlighted the genetic basis for many of the metabolic differences between E. coli 042 and E. coli K-12. \ud \ud Conclusion \ud This study provides a genetic context for the vast amount of experimental and epidemiological data published thus far and provides a template for future diagnostic and intervention strategies

Characterization of extended-spectrum β-lactamases produced by Escherichia coli isolated from hospitalized and nonhospitalized patients : emergence of CTX-M-15-producing strains causing urinary tract infections

Extended-spectrum β-lactamase-producing Escherichia coli isolates were obtained from hospitalised and non-hospitalised patients in Belgium between August 2006 and November 2007. The antimicrobial susceptibility of these isolates was determined and their ESBL genes were characterized. Clonal relationships between the CTX-M-producing E. coli isolates causing urinary tract infections were also studied. A total of 90 hospital- and 45 community-acquired cephalosporin-resistant E. coli isolates were obtained. Tetracycline, enrofloxacine, gentamicin and trimethoprim-sulfamethaxozole resistance rates were significantly different between the community-onset and hospital-acquired isolates. A high diversity of different ESBLs was observed among the hospital-acquired E. coli isolates whereas CTX-M-15 was dominating among the community-acquired E. coli isolates (n=28). Thirtheen different PFGE profiles were observed in the community-acquired CTX-M-15-producing E. coli indicating that multiple clones have acquired the blaCTX-M-15 gene. All community-acquired CTX-M-15-producing E. coli isolates of phylogroups B2 and D were assigned to the sequence type ST131. The hospital-acquired CTX-M-15-producing E. coli isolates of phylogroups B2, B1, A and D corresponded to ST131, ST617, ST48 and ST405, respectively. In conclusion, CTX-M-type ESBLs have emerged as the predominant class of ESBLs produced by E. coli isolates in the hospital and community in Belgium. Of particular concern is the predominant presence of the CTX-M-15 enzyme in ST131 community-acquired E. coli

Escherichia coli K1 RS218 Interacts with Human Brain Microvascular Endothelial Cells via Type 1 Fimbria Bacteria in the Fimbriated State

Escherichia coli K1 is a major gram-negative organism causing neonatal meningitis. E. coli K1 binding to and invasion of human brain microvascular endothelial cells (HBMEC) are a prerequisite for E. coli penetration into the central nervous system in vivo. In the present study, we showed using DNA microarray analysis that E. coli K1 associated with HBMEC expressed significantly higher levels of the fim genes compared to nonassociated bacteria. We also showed that E. coli K1 binding to and invasion of HBMEC were significantly decreased with its fimH deletion mutant and type 1 fimbria locked-off mutant, while they were significantly increased with its type 1 fimbria locked-on mutant. E. coli K1 strains associated with HBMEC were predominantly type 1 fimbria phase-on (i.e., fimbriated) bacteria. Taken together, we showed for the first time that type 1 fimbriae play an important role in E. coli K1 binding to and invasion of HBMEC and that type 1 fimbria phase-on E. coli is the major population interacting with HBMEC

Herd-level risk factors associated with the presence of Phage type 21/28 E. coli O157 on Scottish cattle farms

<p>Background: E. coli O157 is a bacterial pathogen that is shed by cattle and can cause severe disease in humans. Phage type (PT) 21/28 is a subtype of E. coli O157 that is found across Scotland and is associated with particularly severe human morbidity.</p> <p>Methods: A cross-sectional survey of Scottish cattle farms was conducted in the period Feb 2002-Feb 2004 to determine the prevalence of E. coli O157 in cattle herds. Data from 88 farms on which E. coli O157 was present were analysed using generalised linear mixed models to identify risk factors for the presence of PT 21/28 specifically.</p> <p>Results: The analysis identified private water supply, and northerly farm location as risk factors for PT 21/28 presence. There was a significant association between the presence of PT 21/28 and an increased number of E. coli O157 positive pat samples from a farm, and PT 21/28 was significantly associated with larger E. coli O157 counts than non-PT 21/28 E. coli O157.</p> <p>Conclusion: PT 21/28 has significant risk factors that distinguish it from other phage types of E. coli O157. This finding has implications for the control of E. coli O157 as a whole and suggests that control could be tailored to target the locally dominant PT.</p&gt

PRODUCTION AND CHARACTERIZATION OF CRUDE INTRACELLULER PHYTASE FROM RECOMBINANT BACTERIA pEAS1AMP

This research was aimed at producing a crude intracellular phytase characterized from recombinant bacteria. The recombinant bacteria (pEAS1AMP) was produced by way of transforming pET-22b(+) +pEAS1 into competent E. coli BL21 and E. coli BL21(DE3) cells. Crude intracellular phytase production was induced using 1,5 mM Isopropyl-β-D-thiogalactopyranosid (IPTG). Recombinant bacteria product and enzyme activity test followed the Sajidan method. E. coli BL21(+)pEAS1 and E. coli BL21 (DE3)(+)pEAS1 recombinant bacteria showed growth after 20 hours and 10 hours of transformation. Phytase activity of E. coli BL21 (DE3)(+)+pEAS1 showed higher than those of E. coli BL21(+)+pEAS1. Crude intracellular phytase of pEAS1AMP recombinant bacteria has an optimum activity at pH 5, 40o C, incubation period of 60 minutes, substrate concentration of 2%, molecular weight (MW) of 47.3 kDa, Km = 15.91 υM and Vm = 2.41 υM/second. Mg2+ acts as a cofactor but Fe3+ (10-4 M) acts as an inhibitor. Keywords: bacteria recombinant pEAS1AMP, competent cells, crude intracellular phytas

Faecal contamination of lettuce heads after manure application

In recent years, an increasing number of disease outbreaks have been associated with consumption of contaminated vegetables. Thus, it has been speculated to what extent such contamination is associated with application of animal manure as fertilizer, which is particularly practiced in organic vegetable production where conventional fertilizers are prohibited. A field survey was therefore performed aiming to assess the survival and transfer of E. coli from animal manure to lettuces, with E. coli serving as an indicator of bacterial enteric pathogens. Animal manure was applied to 3 Danish fields prior to planting of lettuce seedlings, then 5-8 weeks later at the normal time of harvest, inner and outer leafs of 10 lettuce heads were pooled into one sample unit with a total of 50 pools per field. Additionally, in one field, 15 soil samples were collected weekly until the harvest time. E. coli was enumerated by plating 1 mL of 10-fold serial dilutions of 5 g of homogenized sample material, i.e. manure, soil and lettuce onto PetrifilmTM Select E. coli count plates (3M), which were then incubated 24 h at 44°C. The manure applied to the fields contained 3.0-4.5 Log10 E. coli CFU/g and E. coli was found in 36-54% of the pooled lettuce samples with a detection limit of 10 CFU/g. Numbers of E. coli in 14-20% of pooled lettuce samples exceeded a satisfactory microbiological hygiene criteria level of 100 CFU/g. The highest percentage of faecally contaminated lettuce heads (54%) coincided with the shortest growth period studied indicating that the time gap between application of manure and harvest and the survival of E. coli (and pathogens) influences the contamination of lettuce via manure amended soil. However, at the time of harvest, the numbers of E. coli in 5 of 15 soil samples were reduced below the detection limit and no samples exceeded 100 CFU/g. This is in contrast to the lettuce samples, where 20% of faecally contaminated samples had >100 E. coli/g, which may indicate that faeces contamination of crops could originate from alternative sources, such as contaminated water and wildlife. Comparisons of the genotype of isolated E. coli strains could help to elucidate this

Comparative genomics and transcriptomics of Escherichia coli isolates carrying virulence factors of both enteropathogenic and enterotoxigenic E. coli

AbstractEscherichia coli that are capable of causing human disease are often classified into pathogenic variants (pathovars) based on their virulence gene content. However, disease-associated hybrid E. coli, containing unique combinations of multiple canonical virulence factors have also been described. Such was the case of the E. coli O104:H4 outbreak in 2011, which caused significant morbidity and mortality. Among the pathovars of diarrheagenic E. coli that cause significant human disease are the enteropathogenic E. coli (EPEC) and enterotoxigenic E. coli (ETEC). In the current study we use comparative genomics, transcriptomics, and functional studies to characterize isolates that contain virulence factors of both EPEC and ETEC. Based on phylogenomic analysis, these hybrid isolates are more genomically-related to EPEC, but appear to have acquired ETEC virulence genes. Global transcriptional analysis using RNA sequencing, demonstrated that the EPEC and ETEC virulence genes of these hybrid isolates were differentially-expressed under virulence-inducing laboratory conditions, similar to reference isolates. Immunoblot assays further verified that the virulence gene products were produced and that the T3SS effector EspB of EPEC, and heat-labile toxin of ETEC were secreted. These findings document the existence and virulence potential of an E. coli pathovar hybrid that blurs the distinction between E. coli pathovars.</jats:p