Escherichia coli isolated from food sources and irrigation water : a potential risk for causing intestinal dysfunction?

We have previously shown that diarrheagenic Escherichia coli (DEC) and non-DEC are prevalent in food sources and irrigation water in South Africa. Recent data suggest that an increased relative abundance of faecal Enterobacteriaceae is associated with poorer health outcomes among children in developing countries. Thus, exposure to non-DEC from environmental sources may incur adverse effects, although the mechanisms underlying these effects remain obscure. To further elucidate this phenomenon, we assayed non-DEC strains from environmental sources in South Africa for phenotypes that may be associated with intestinal dysfunction (ID). DEC strains were also used. The strains had previously been isolated from Producer Distributor Bulk Milk (PDBM), irrigated lettuce, street vendor coleslaw and irrigation water. In-vitro assays identified; biofilm formation (n = 38), extracellular polymeric substance (EPS) formation (n = 38), cytotoxic activity (n = 10), disruption of tight junctions and induction of Interleukin 8 (IL-8) on polarized T-84 cells (n = 20). The number of strains tested for each assay differed, depending on prior molecular and phenotypic characterization that signalled potential pathogenicity in-vitro. Subsequently, all strains having data points for all analyses were used to compute Principal Component Analysis (PCA) plot curves to infer potential associations amongst test strains and a standard DEC pathogenic strain (042). Biofilm formation on glass coverslips after strains were grown in nutrient-rich media (LB and DMEM-F12 + 0.5% D-Mannose) at 37 °C varied based on pathotype (DEC and non-DEC) and source of isolation (food, irrigation water, clinical) suggesting that pathotype and source isolation influence persistence within a defined environmental niche. Additionally, DEC isolated from irrigated lettuce had a significantly higher (p ≤ 0.05) propensity for biofilm formation in both media compared to all strains including DEC standard controls. This suggested the propensity for irrigated lettuce as a potential source of persistent pathogenic strains. Furthermore, all strains were able to form EPS suggesting the ability to form mature biofilms under conditions relevant for food processing (20–25 °C). Of the (60%, 6 out of 10) strains that showed cytotoxic activity, most (83%, 5 out of 6 strains) were non-DEC isolated from food sources many of which are consumed with minimal processing. Mean percentage reduction in initial TEER (a measure of intestinal disruption), did not significantly differ (p = 0.05) in all test strains from that observed in the standard DEC. Additionally, IL-8 induction from strains isolated from PDBM (139 pg/mL), irrigation water (231.93 pg/mL) and irrigated lettuce (152.98 pg/mL) was significantly higher (p ≤ 0.05) than in the commensal strain aafa. PCA categorized strains based on sources of isolation showed potential for use in source tracking especially when comparing many strains from various environmental sources. We show that non-DEC strains along the food chain possess characteristics that may lead to ID. Further investigations using a larger collection of strains may provide a clearer link to these reported observations that could be associated with the high diarrheal disease burden within the country, especially among infants.The Department of Research and Innovation, University of Pretoria for a post-graduate travel bursary to Matthew Aijuka to travel to James P. Nataro's laboratory at The University of Virginia. Work in the Nataro lab was supported by US National Institutes of Health grant AI-33096 to JPN.http://www.elsevier.com/locate/foodcont2020-08-01hj2019Food Scienc

A first approach to assess the impact of bottom trawling over Vulnerable Marine Ecosystems on the High Seas of the Southwest Atlantic

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Role of age and comorbidities in mortality of patients with infective endocarditis

[Purpose]: The aim of this study was to analyse the characteristics of patients with IE in three groups of age and to assess the ability of age and the Charlson Comorbidity Index (CCI) to predict mortality. [Methods]: Prospective cohort study of all patients with IE included in the GAMES Spanish database between 2008 and 2015.Patients were stratified into three age groups:<65 years,65 to 80 years,and ≥ 80 years.The area under the receiver-operating characteristic (AUROC) curve was calculated to quantify the diagnostic accuracy of the CCI to predict mortality risk. [Results]: A total of 3120 patients with IE (1327 < 65 years;1291 65-80 years;502 ≥ 80 years) were enrolled.Fever and heart failure were the most common presentations of IE, with no differences among age groups.Patients ≥80 years who underwent surgery were significantly lower compared with other age groups (14.3%,65 years; 20.5%,65-79 years; 31.3%,≥80 years). In-hospital mortality was lower in the <65-year group (20.3%,<65 years;30.1%,65-79 years;34.7%,≥80 years;p < 0.001) as well as 1-year mortality (3.2%, <65 years; 5.5%, 65-80 years;7.6%,≥80 years; p = 0.003).Independent predictors of mortality were age ≥ 80 years (hazard ratio [HR]:2.78;95% confidence interval [CI]:2.32–3.34), CCI ≥ 3 (HR:1.62; 95% CI:1.39–1.88),and non-performed surgery (HR:1.64;95% CI:11.16–1.58).When the three age groups were compared,the AUROC curve for CCI was significantly larger for patients aged <65 years(p < 0.001) for both in-hospital and 1-year mortality. [Conclusion]: There were no differences in the clinical presentation of IE between the groups. Age ≥ 80 years, high comorbidity (measured by CCI),and non-performance of surgery were independent predictors of mortality in patients with IE.CCI could help to identify those patients with IE and surgical indication who present a lower risk of in-hospital and 1-year mortality after surgery, especially in the <65-year group

Agave Fructans in Oaxaca&rsquo;s Emblematic Specimens: Agave angustifolia Haw. and Agave potatorum Zucc.

Despite the recognition of Agave tequilana Weber var. Azul as raw material for producing tequila and obtaining prebiotics, there are other highly relevant Agave species in Mexico. Oaxaca contains a startlingly diverse range of Agave species; Agave angustifolia Haw. and Agave potatorum Zucc. are two classic specimens with great commercial potential. In this study, we examined the fructan fluctuation in these two species during their lifetime in the field (from 1 to 6 years old). First, we analyzed their morphological diversity based on vegetative characteristics. Subsequently, fructan extracts were analyzed by TLC, FT-IR, and HPAEC-PAD to identify carbohydrates. Multivariate analyses of the morphological parameters indicated a morphological divergence between the two species. Furthermore, we found that the concentration of simple carbohydrates and fructans, as well as the fructan DP, changed during plant development. Glucose, fructose, and fructooligosaccharides (FOS) were more abundant in A. potatorum, while A. angustifolia showed a greater amount of sucrose and fructans with a high DP. Fructan DP heatmaps were constructed using HPAEC-PAD profiles&mdash;the heatmaps were very helpful for establishing an easy correlation between age and the carbohydrate types present in the fructan extracts. This study is an important contribution to the agave fructan knowledge of the Mexican agave diversity

Enteroaggregative Escherichia coli is the predominant diarrheagenic E. coli pathotype among irrigation water and food sources in South Africa

Diarrheagenic E. coli (DEC) has been implicated in foodborne outbreaks worldwide and have been associated with childhood stunting in the absence of diarrhoea. Infection is extraordinarily common, but the routes of transmission have not been determined. Therefore, determining the most prevalent pathotypes in food and environmental sources may help provide better guidance to various stakeholders in ensuring food safety and public health and advancing understanding of the epidemiology of enteric disease. We characterized 205 E. coli strains previously isolated from producer distributor bulk milk (PDBM)(118), irrigation water (48), irrigated lettuce (29) and street vendor coleslaw (10) in South Africa. Enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), enteroaggregative E. coli (EAEC) and diffusely adherent E. coli (DAEC) were sought. We used PCR and partial gene sequencing for all 205 strains while 46 out of 205 that showed poor resolution were subsequently characterized using cell adherence (HeLa cells). PCR and partial gene sequencing of aatA and/or aaiC genes confirmed EAEC (2%, 5 out of 205) as the only pathotype. Phylogenetic analysis of sequenced EAEC strains with E. coli strains in GenBank showing ≥80% nucleotide sequence similarity based on possession of aaiC and aatA generated distinct clusters of strains separated predominantly based on their source of isolation (food source or human stool) suggesting a potential role of virulence genes in source tracking. EAEC 24%, 11 out of 46 strains (PDBM = 15%, irrigation water = 7%, irrigated lettuce = 2%) was similarly the predominant pathotype followed by strains showing invasiveness to HeLa cells, 4%, 2 out of 46 (PDBM = 2%, irrigated lettuce = 2%), among stains characterized using cell adherence. Therefore, EAEC may be the leading cause of DEC associated food and water-borne enteric infection in South Africa. Additionally, solely using molecular based methods targeting virulence gene determinants may underestimate prevalence, especially among heterogeneous pathogens such as EAEC.The Department of Research and Innovation, University of Pretoria for a post-graduate travel bursary to Matthew Aijuka to travel to James P. Nataro's laboratory at The University of Virginia. Work in the Nataro lab was supported by US National Institutes of Health grant AI-33096 to JPN.http://www.elsevier.com/ locate/ijfoodmicro2019-08-02hj2018Consumer ScienceFood Scienc

Fructosyltransferases in plants: Structure, function and application: A review

Fructosyltransferases (FTs) are an important group of enzymes involved in the biosynthesis of fructans in some plants and in a large number of microorganisms. Several studies have shown that FTs play a key role in regulating the levels of fructan molecules during abiotic stress tolerance of certain plant species. Because of their diverse range of functional characteristics, efforts have increased to improve fructans applications in food quality and in added health benefits of plants. For these reasons, interest on the search for more direct and cost-effective routes to producing fructans have also increased. In this paper, we review recent research on plant FTs and summarize their role in the regulation of fructan biosynthesis and their use in producing fructans at an industrial scale. We also present the challenges and opportunities that FTs will present in the future

Novel Aggregative Adherence Fimbria Variant of Enteroaggregative Escherichia coli

Enteroaggregative Escherichia coli (EAEC) organisms 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 one-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 an AAF-related gene encoding a yet-uncharacterized adhesin, termed agg5A. The sequence of the agg5DCBA gene cluster shared fimbrial accessory genes (usher, chaperone, and minor pilin subunit genes) with AAF/III, as well as the signal peptide present in the beginning of the agg3A gene. The complete agg5DCBA gene cluster from a clinical isolate, EAEC strain C338-14, with the typical stacked-brick binding pattern was cloned, and deletion of the cluster was performed. Transformation to a nonadherent E. coli HB101 and complementation of the nonadherent C338-14 mutant with the complete gene cluster restored the AA adhesion. Overall, we found the agg5A gene in 12% of the 118 strains isolated from Denmark, suggesting that this novel adhesin represents an important variant

Characterization of a novel AraC/XylS-regulated family of N-acyltransferases in pathogens of the order Enterobacterales.

Enteroaggregative Escherichia coli (EAEC) is a diarrheagenic pathotype associated with traveler's diarrhea, foodborne outbreaks and sporadic diarrhea in industrialized and developing countries. Regulation of virulence in EAEC is mediated by AggR and its negative regulator Aar. Together, they control the expression of at least 210 genes. On the other hand, we observed that about one third of Aar-regulated genes are related to metabolism and transport. In this study we show the AggR/Aar duo controls the metabolism of lipids. Accordingly, we show that AatD, encoded in the AggR-regulated aat operon (aatPABCD) is an N-acyltransferase structurally similar to the essential Apolipoprotein N-acyltransferase Lnt and is required for the acylation of Aap (anti-aggregation protein). Deletion of aatD impairs post-translational modification of Aap and causes its accumulation in the bacterial periplasm. trans-complementation of 042aatD mutant with the AatD homolog of ETEC or with the N-acyltransferase Lnt reestablished translocation of Aap. Site-directed mutagenesis of the E207 residue in the putative acyltransferase catalytic triad disrupted the activity of AatD and caused accumulation of Aap in the periplasm due to reduced translocation of Aap at the bacterial surface. Furthermore, Mass spectroscopy revealed that Aap is acylated in a putative lipobox at the N-terminal of the mature protein, implying that Aap is a lipoprotein. Lastly, deletion of aatD impairs bacterial colonization of the streptomycin-treated mouse model. Our findings unveiled a novel N-acyltransferase family associated with bacterial virulence, and that is tightly regulated by AraC/XylS regulators in the order Enterobacterales

Broad Spectrum Activity of a Lectin-Like Bacterial Serine Protease Family on Human Leukocytes

<div><p>The serine protease autotransporter from Enterobacteriaceae (SPATE) family, which number more than 25 proteases with apparent diverse functions, have been phylogenetically divided into two distinct classes, designated 1 and 2. We recently demonstrated that Pic and Tsh, two members of the class-2 SPATE family produced by intestinal and extraintestinal pathogenic <i>E. coli</i>, were able to cleave a number of O-glycosylated proteins on neutrophils and lymphocytes resulting in impaired leukocyte functions. Here we show that most members of the class-2 SPATE family have lectin-like properties and exhibit differential protease activity reliant on glycoprotein type and cell lineage. Protease activity was seen in virtually all tested O-glycosylated proteins including CD34, CD55, CD164, TIM1, TIM3, TIM4 and C1-INH. We also show that although SPATE proteins bound and cleaved glycoproteins more efficiently on granulocytes and monocytes, they also targeted glycoproteins on B, T and natural killer lymphocytes. Finally, we found that the characteristic domain-2 of class-2 SPATEs is not required for glycoprotease activity, but single amino acid mutations in Pic domain-1 to those residues naturally occurring in domain-1 of SepA, were sufficient to hamper Pic glycoprotease activity. This study shows that most class-2 SPATEs have redundant activities and suggest that they may function as immunomodulators at several levels of the immune system.</p></div