High frequency of multidrug-resistant Escherichia coli from cattle in the Cerrado and Pantanal biomes of Brazil.

The indiscriminate use of antimicrobials has led to the emergence of resistant bacteria, especially pathogenic strains of Escherichia coli, which are associated with diseases in animals and humans. The aim of the present study was to characterize E. coli isolates in calves with regards to the presence of virulence genes and investigate the resistance of the isolates to different antimicrobials. Between 2021 and 2023, 456 fecal samples were collected from calves in the Pantanal and Cerrado biomes of the state of Mato Grosso do Sul, Brazil. All samples were subjected to microbiological analysis and disc diffusion antibiogram testing. The polymerase chain reaction method was used to detect virulence genes. Bacterial growth was found in 451 of the 456 samples and biochemically identified as Escherichia coli. All 451 isolates (100 %) exhibited some phenotypic resistance to antimicrobials and 67.62 % exhibited multidrug resistance. The frequency of multidrug-resistant isolates in the Cerrado biome was significantly higher than that in the Pantanal biome (p = 0.0001). In the Cerrado, the most common pathotype was Shiga toxin-producing Escherichia coli (STEC) (28 %), followed by toxigenic Escherichia coli (ETEC) (11 %), enterohemorrhagic Escherichia coli (EHEC) (8 %) and enteropathogenic Escherichia coli (EPEC) (2 %). In most cases, the concomitant occurrence of pathotypes was more common, the most frequent of which were ETEC + STEC (33 %), ETEC + EHEC (15 %) and ETEC + EPEC (3 %). The STEC pathotype (30 %) was also found more frequently in the Pantanal, followed by EHEC (12 %), ETEC (9 %) and EPEC (6 %). The STEC pathotype had a significantly higher frequency of multidrug resistance (p = 0.0486) compared to the other pathotypes identified. The frequency of resistance was lower in strains from the Pantanal biome compared to those from the Cerrado biome. Although some factors are discussed in this paper, it is necessary to clarify the reasons for this difference and the possible impacts of these findings on both animal and human health in the region.

Acquisition of plasmids from Shiga toxin-producing Escherichia coli strains had low or neutral fitness cost on commensal E. coli.

Although it has been hypothesized that the acquisition of plasmids-especially those bearing virulence factors and antimicrobial resistance genes-increases the energetic burden and reduces the fitness of a bacterium in general, some results have challenged this view, showing little or no effect on fitness after plasmid acquisition, which may lead to change in the view that there are evolutionary barriers for a wide spread of such plasmids among bacteria. Here, to evaluate the fitness impact of plasmid-encoded antibiotic resistance and virulence genes, plasmids from O26:H11, O111:H8, and O118:H16 Shiga toxin-producing Escherichia coli (STEC) human and bovine isolates were transferred to the non-virulent E. coli HS and K-12 MG1655 strains. Sequencing and PCR were used to characterize plasmids, and to identify the presence of antimicrobial resistance and/or virulence genes. The fitness impact of plasmids encoding virulence and antimicrobial resistance upon bacterial hosts was determined by pairwise growth competition. Plasmid profile analysis showed that STEC strains carried one or more high and low molecular weight plasmids belonging to the B/O, F, I, K, P, Q, and/or X incompatibility groups encoding virulence genes (SPATE-encoding genes) and/or antimicrobial resistance genes (aadA1, strAB, tetA, and/or tetB). Competition experiments demonstrated that the biological cost of carriage of these plasmids by the commensal E. coli strain HS or the laboratory strain E. coli K-12 MG1655 was low or non-existent, ranging from - 4.7 to 5.2% per generation. This suggests that there are few biological barriers-or, alternatively, it suggests that there are biological barriers that we were not able to measure in this competition model-against the spread of plasmid encoding virulence and resistance genes from STEC to other, less pathogenic E. coli strains. Thus, our results, in opposition to a common view, suggest that the acquisition of plasmids does not significantly affect the bacteria fitness and, therefore, the theorized plasmid burden would not be a significant barrier for plasmid spread.

Prevalence, Antimicrobial Resistance, and Whole Genome Sequencing Analysis of Shiga Toxin-Producing Escherichia coli (STEC) and Enteropathogenic Escherichia coli (EPEC) from Imported Foods in China during 2015-2021.

Shiga toxin-producing Escherichia coli (STEC) and enteropathogenic Escherichia coli (EPEC) are foodborne pathogens that cause hemolytic uremic syndrome and fatal infant diarrhea, respectively, but the characterization of these bacteria from imported food in China are unknown. A total of 1577 food samples from various countries during 2015-2021 were screened for STEC and EPEC, and the obtained isolates were tested for antimicrobial resistance and whole genome sequencing analysis was performed. The prevalence of STEC and EPEC was 1.01% (16/1577) and 0.51% (8/1577), respectively. Antimicrobial resistances to tetracycline (8%), chloramphenicol (8%), ampicillin (4%), ceftazidime (4%), cefotaxime (4%), and trimethoprim-sulfamethoxazole (4%) were observed. The antimicrobial resistance phenotypes corresponded with genotypes for most strains, and some resistance genes were related to mobile genetic elements. All 16 STEC isolates were eae negative, two solely contained stx1 (stx1a or stx1c), 12 merely carried stx2 (stx2a, stx2d, or stx2e), and two had both stx1 and stx2 (stx1c + stx2b, stx1a + stx2a + stx2c). Although they were eae negative, several STEC isolates carried other adherence factors, such as iha (5/16), sab (1/16), and lpfA (8/16), and belonged to serotypes (O130:H11, O8:H19, and O100:H30) or STs (ST297, ST360), which have caused human infections. All the eight EPEC isolates were atypical EPEC; six serotypes and seven STs were found, and clinically relevant EPEC serotypes O26:H11, O103:H2, and O145:H28 were identified. Two STEC/ETEC (enterotoxigenic E. coli) hybrids and one EPEC/ETEC hybrid were observed, since they harbored sta1 and/or stb. The results revealed that food can act as a reservoir of STEC/EPEC with pathogenic potential, and had the potential ability to transfer antibiotic resistance and virulence genes.

Large-Scale Phylogenetic Analysis Reveals a New Genetic Clade among Escherichia coli O26 Strains.

Shiga toxin-producing Escherichia coli (STEC) O26 is the predominant non-O157 serogroup causing hemolytic uremic syndrome worldwide. Moreover, the serogroup is highly dynamic and harbors several pathogenic clones. Here, we investigated the phylogenetic relationship of STEC O26 at a global level based on 1,367 strains from 20 countries deposited in NCBI and Enterobase databases. The whole-genome-based analysis identified a new genetic clade, called ST29C4. The new clade was unique in terms of multilocus sequence type (ST29), CRISPR (group Ia), and dominant plasmid gene profile (ehxA+/katP-/espP-/etpD-). Moreover, the combination of multiple typing methods (core genome single nucleotide polymorphism [SNP] typing, CRISPR typing, and virulence genes analysis) demonstrated that this new lineage ST29C4 was in the intermediate phylogenetic position between ST29C3 and other non-ST29C3 strains. Besides, we observed that ST29C4 harbored extraintestinal pathogenic E. coli (ExPEC)-related virulence gene (VG), tsh, and STEC-associated VG, stx2a, suggesting the emergence of a hybrid pathogen. The ST29C4 strains also exhibited high similarity in stx2a-prophage and integrase with the O104:H4 strain, further demonstrating its potential risk to human health. Collectively, the large-scale phylogenetic analysis extends the understanding of the clonal structure of O26 strains and provides new insights for O26 strain microevolution. IMPORTANCE Shiga toxin-producing Escherichia coli (STEC) O26 is the second prevalent STEC serogroup only to O157, which can cause a series of diseases ranging from mild diarrhea to life-threatening hemolytic uremic syndrome (HUS). The serogroup is highly diverse and multiple clones are characterized, including ST29C1-C3 and ST21C1-C2. However, the phylogenetic relationship of these clones remains fully unclear. In this study, we revealed a new genetic clade among O26 strains, ST29C4, which was unique in terms of CRISPR, multilocus sequence type (MLST), and plasmid gene profile (PGP). Moreover, the combination of multiple typing methods demonstrated that this new clone was located in the intermediate phylogenetic position between ST29C3 and other non-ST29C3 strains (i.e., ST29C1-C2 and ST21C1-C2). Overall, the large-scale phylogenetic analysis extends our current understanding of O26 microevolution.

Prevalence of genotypic antimicrobial resistance in clinical Shiga toxin-producing Escherichia coli in Norway, 2018 to 2020.

Introduction. Shiga toxin-producing Escherichia coli (STEC) can cause severe to fatal disease in humans. Antimicrobial treatment is sometimes necessary, but contraindicated due to undesirable clinical outcome. However, recent studies have shown promising outcomes following antimicrobial treatment. Before the establishment of a possible antimicrobial treatment strategy for STEC infections, the prevalence of antimicrobial resistance in STEC needs to be determined.Gap Statement. The resistance status of Norwegian clinical STEC is not known and should be assessed.Aim. We aim to characterize genotypic antimicrobial resistance determinants in clinical STEC in Norway, and determine the prevalence of genotypic resistance in order to inform possible antimicrobial treatment options for STEC infections.Methodology. We included all clinical STEC submitted to the Norwegian Reference Laboratory from March 2018 to April 2020. All samples were whole-genome sequenced and screened for genotypic antimicrobial resistance,virulence determinants and plasmid incompatibility groups. We performed phylogenetic clustering of STEC by core-genome multi-locus sequence typing, and statistical association analyses between isolate characteristics and genotypic resistance.Results. A total of 459 STEC were analysed. For 385 (83.9 %) STEC we did not identify any antimicrobial resistance determinants. Seventy-four STEC (16.1 %) harboured antimicrobial resistance determinants against one or more antimicrobial classes. The most frequent genotypic resistance was identified against aminoglycosides (10.5 %). Thirty-nine STEC (8.5 %) had a multi-drug resistance (MDR) genotype. Genotypic resistance was more prevalent in non-O157 than O157 STEC (P=0.02). A positive association was seen between genotypic resistance and the low-virulent STEC O117:H7 phylogenetic cluster (no. 14) (P<0.001). Genotypic resistance was not significantly associated to high-virulent STEC. STEC O146:H28 and isolates harbouring the plasmid replicon type IncQ1 were positively associated with MDR.Conclusion. The overall prevalence of genotypic resistance in clinical STEC in Norway is low (16.1 %). Genotypic resistance is more prevalent in non-O157 strains compared to O157 strains, and not significantly associated to high-virulent STEC. Resistance to antimicrobials suggested for treatment, especially azithromycin is low and may present an empiric treatment alternative for severe STEC infections.

Genetic and antimicrobial resistance profiles of non-O157 Shiga toxin-producing Escherichia coli from different sources in Egypt.

BACKGROUND: The Shiga toxin-producing Escherichia coli (STEC) represented a great risk to public health. In this study, 60 STEC strains recovered from broiler and duck fecal samples, cow's milk, cattle beef, human urine, and ear discharge were screened for 12 virulence genes, phenotypic and genotypic antimicrobial resistance, and multiple-locus variable-number tandem-repeat analysis (MLVA). RESULTS: The majority of strains harbored Shiga toxin 1 (stx1) and stx1d, stx2 and stx2e, and ehxA genes, while a minority harbored stx2c subtype and eaeA. We identified 10 stx gene combinations; most of strains 31/60 (51.7%) exhibited four copies of stx genes, namely the stx1, stx1d, stx2, and stx2e, and the strains exhibited a high range of multiple antimicrobial resistance indices. The resistance genes blaCTX-M-1 and blaTEM were detected. For the oxytetracycline resistance genes, most of strains contained tetA, tetB, tetE, and tetG while the tetC was present at low frequency. MLVA genotyping resolved 26 unique genotypes; genotype 21 was highly prevalent. The six highly discriminatory loci DI = 0.9138 are suitable for the preliminary genotyping of STEC from animals and humans. CONCLUSIONS: The STEC isolated from animals are virulent, resistant to antimicrobials, and genetically diverse, thus demands greater attention for the potential risk to human.

Multidrug resistance IncC plasmid carrying blaCMY-97 in Shiga toxin-producing Escherichia coli ST215-H54 of ovine origin.

CMY-type ß-lactamases are the most reported plasmid-mediated AmpC (pAmpC), with the CMY-2-like group being the most clinically relevant described in Escherichia coli at human-animal-environment interface. Shiga toxin-producing E. coli (STEC) lineages are zoonotic pathogens commonly reported causing serious clinical conditions in humans, including severe diarrheagenic diseases. Therefore, this study aimed to investigate a multidrug-resistant (MDR) STEC isolate (A313) recovered from a healthy sheep and carrying mobile blaCMY-97, that encodes a pAmpC belonging to the CMY-2-like group. The A313 isolate exhibited a MDR profile to clinically relevant antimicrobials (i.e., cephalosporins, aminoglycosides, and fluoroquinolones), but reduced susceptibility to extended-spectrum cephalosporins and aztreonam. Besides, virulence genes (stx2, gad and iutA) were detected in A313, which belonged to ST215/CC10 and phylogenetic group A, whereas the fimH54 was identified. The blaCMY-97 gene and other antimicrobial resistance determinants [aph(6)-Id, aph(3″)-Ib, aac(3)-IId, aadA5, floR, tetA, sul1, and sul2], as well as genes encoding tolerance to mercury (merRTPCADE), were harbored by an IncC plasmid (named pA313-CMY-97, ~ 176 kb). A novel genetic context of blaCMY-2-like, in which a 208-bp ISEcp1 was truncated by an IS26 in the opposite orientation upstream of the blaCMY-97 gene (IS26-∆ISEcp1-blaCMY-97-blc-sugE-encR), was also identified in pA313-CMY-97. To the best of our knowledge, this is the first report on the acquisition of blaCMY-97 into a plasmid. Therefore, we reported ovine as reservoir of clinically relevant MDR bacteria carrying mobile blaCMY-97 with potential for zoonotic transmission.

Detection of novel sequence types and zoonotic transmission potentiality among strains of Shiga toxigenic Escherichia coli (STEC) from dairy calves, animal handlers and associated environments.

Shiga toxigenic Escherichia coli (STEC) is one of the most important food-borne zoonotic bacterial pathogens responsible for causing gastrointestinal infections, haemorrhagic colitis and haemolytic uremic syndrome. The present study was aimed to isolate and characterize STEC from neonatal dairy calves, animal handlers and their surrounding environment and to establish the genetic relationship among isolates by multilocus sequence typing (MLST). A total number of 115 samples were collected and processed for the isolation of E. coli. The occurrence rate of E. coli was 92.2% (106/115), of which, 18 were typed as STEC. Antibacterial susceptibility analysis revealed 11 (61.1%) strains as multiple drug-resistant (MDR). MLST analysis has delineated 16 sequence types (STs) including nine novel STs. Among STs, ST58 dominated with three strains and was recovered from the environment and neonatal calves. Strains from neonatal calves and humans showed genetic relatedness with significant bootstrap support values indicative of zoonotic transmission potentiality. Analysis of 211 global isolates belonging to 61 STs indicated predominant STs (ST 21, ST 33 and ST 3416) that can be either host-specific (ST 33 and ST 3416) or can be shared among human and bovine hosts (ST 21). The MLST analysis indicates genetic relatedness among isolates and the results predispose inter-host transmission and zoonotic spread.

Psychoactive Drugs Induce the SOS Response and Shiga Toxin Production in Escherichia coli.

Several classes of non-antibiotic drugs, including psychoactive drugs, proton-pump inhibitors (PPIs), non-steroidal anti-inflammatory drugs (NSAIDs), and others, appear to have strong antimicrobial properties. We considered whether psychoactive drugs induce the SOS response in E. coli bacteria and, consequently, induce Shiga toxins in Shiga-toxigenic E. coli (STEC). We measured the induction of an SOS response using a recA-lacZ E. coli reporter strain, as RecA is an early, reliable, and quantifiable marker for activation of the SOS stress response pathway. We also measured the production and release of Shiga toxin 2 (Stx2) from a classic E. coli O157:H7 strain, derived from a food-borne outbreak due to spinach. Some, but not all, serotonin selective reuptake inhibitors (SSRIs) and antipsychotic drugs induced an SOS response. The use of SSRIs is widespread and increasing; thus, the use of these antidepressants could account for some cases of hemolytic-uremic syndrome due to STEC and is not attributable to antibiotic administration. SSRIs could have detrimental effects on the normal intestinal microbiome in humans. In addition, as SSRIs are resistant to environmental breakdown, they could have effects on microbial communities, including aquatic ecosystems, long after they have left the human body.

Multidrug-Resistant Extended-Spectrum ß-Lactamase-Producing Escherichia coli Pathotypes in North Eastern Region of India: Backyard Small Ruminants-Human-Water Interface.

A total of 648 diarrheagenic Escherichia coli (DEC) were isolated from calves (n = 219), lambs (n = 87), kids (n = 103), human (n = 193), and water (n = 46) samples. The presence of enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), and shigatoxigenic E. coli (STEC) was confirmed by PCR-based detection of the Shiga toxin, intimin, hemolysin, and enterotoxin genes. All the isolates were tested for antimicrobial resistance (AMR) by disc diffusion assay. Extended-spectrum ß-lactamase (ESBL), carbapenemase, and metallo-beta-lactamase production were determined by double-disk synergy test, modified Hodge test, and combined disk test assays. AMR genes (blaTEM, blaSHV, blaCTX-M, blaCMY-2, blaNDM, blaKPC, blaVIM, and blaIMP) were detected by PCR using specific primers. Majority of the isolates from human and water exhibited resistance (>80%) against amoxicillin, ampicillin, aztreonam, cefotaxime, cefixime, gentamicin, ceftazidime, and cefalexin, and against imipenem (70.98%), doripenem (70.47%), and ertapenem (60.62%). Bovine isolates were sensitive to carbapenems. Many isolates (5.75-24.35%) from human, water, calves, kids, and lambs were multidrug resistant (MDR), with resistance against three or more classes of antimicrobials. A total of 170/648 (26.23%) isolates were classified as STEC (9.88%), EPEC (4.32%), and ETEC (12.04%). The AMR genes, including blaTEM, blaCMY2, blaCTX-M, and blaSHV were detected in the E. coli from all sources. but blaNDM and blaKPC were detected only in the isolates from human and water. Three STEC isolates from human origin possessed multiple ESBLs, carbapenemase and metallo-beta-lactamase genes reported for the first time. ESBLs producing EPEC and ETEC in lambs and kids are also reported under this study. Presence of MDR-DEC in domestic animals and common potable water poses public health concern in this region.

Whole-genome sequencing analysis of uncommon Shiga toxin-producing Escherichia coli from cattle: Virulence gene profiles, antimicrobial resistance predictions, and identification of novel O-serogroups.

Shiga toxin-producing E. coli (STEC) are major foodborne pathogens. While many studies have focused on the "top-7 STEC", little is known for minor serogroups. A total of 284 non-top-7 STEC strains isolated from cattle feces were subjected to whole-genome sequencing (WGS) to determine the serotypes, the presence of virulence genes and antimicrobial resistance (AMR) determinants. Nineteen typeable and three non-typeable serotypes with novel O-antigen loci were identified. Twenty-one AMR genes and point mutations in another six genes that conferred resistance to 10 antimicrobial classes were detected, as well as 46 virulence genes. The distribution of 33 virulence genes and 15 AMR determinants exhibited significant differences among serotypes (p < 0.05). Among all strains, 81.7% (n = 232) and 14.1% (n = 40) carried stx2 and stx1 only, respectively; only 4.2% (n = 12) carried both. Subtypes stx1a, stx1c, stx2a, stx2c, stx2d, and stx2g were identified. Forty-six strains carried eae and stx2a and therefore had the potential cause severe diseases; 47 strains were genetically related to human clinical strains inferred from a pan-genome phylogenetic tree. We were able to demonstrate the utility of WGS as a surveillance tool to characterize the novel serotypes, as well as AMR and virulence profiles of uncommon STEC that could potentially cause human illness.

Antibiotic-mediated expression analysis of Shiga toxin 1 and 2 in multi-drug-resistant Shiga toxigenic Escherichia coli.

Shiga toxin-producing Escherichia coli (STEC) is an important foodborne pathogens, known to cause enteric infections especially diarrhea, mainly attributed to Shiga toxins (Stxs). The use of certain antibiotics for treating this infection is controversial, owing to an increased risk for producing Stxs (Stx 1 and Stx 2). Increased antibiotic resistance is also thought to be involved in the pathogenesis of STEC diseases. The purpose of this study was to analyze the effects of antibiotics on induction of Stx 1 and Stx 2 in clinical STEC isolates and to investigate the relationships between increased resistance and Stx production. Fifteen clinical isolates were treated with sub minimum inhibitory concentrations (Sub MIC) of clinically used antibiotics (ciprofloxacin, fosfomycin, tigecycline, and meropenem), and the changes in expression levels of stx1 and stx2 genes were estimated using qRT-PCR. The expressions of Shiga toxins were found to be increased up to 6.5- and eightfold under ciprofloxacin and tigecycline Sub MIC, respectively. Fosfomycin had weak induction effect of up to twofold, whereas meropenem had the weakest influence on such expression. Resistant isolates were found to be more prone to increased expression of toxins.

Natural and synthetic antimicrobials reduce adherence of enteroaggregative and enterohemorrhagic Escherichia coli to epithelial cells.

Adherence of bacteria to the human intestinal mucosa can facilitate their internalization and the development of pathological processes. Escherichia coli O104:H4 is considered a hybrid strain (enteroaggregative hemorrhagic E. coli [EAHEC]), sharing virulence factors found in enterohemorrhagic (EHEC), and enteroaggregative (EAEC) E. coli pathotypes. The objective of this study was to analyze the effects of natural and synthetic antimicrobials (carvacrol, oregano extract, brazilin, palo de Brasil extract, and rifaximin) on the adherence of EHEC O157:H7, EAEC 042, and EAHEC O104:H4 to HEp-2 cells and to assess the expression of various genes involved in this process. Two concentrations of each antimicrobial that did not affect (p≤0.05) bacterial viability or damage the bacterial membrane integrity were used. Assays were conducted to determine whether the antimicrobials alter adhesion by affecting bacteria and/or alter adhesion by affecting the HEp-2 cells, whether the antimicrobials could detach bacteria previously adhered to HEp-2 cells, and whether the antimicrobials could modify the adherence ability exhibited by the bacteria for several cycles of adhesion assays. Giemsa stain and qPCR were used to assess the adhesion pattern and gene expression, respectively. The results showed that the antimicrobials affected the adherence abilities of the bacteria, with carvacrol, oregano extract, and rifaximin reducing up to 65% (p≤0.05) of E. coli adhered to HEp-2 cells. Carvacrol (10 mg/ml) was the most active compound against EHAEC O104:H4, even altering its aggregative adhesion pattern. There were changes in the expression of adhesion-related genes (aggR, pic, aap, aggA, and eae) in the bacteria and oxidative stress-related genes (SOD1, SOD2, CAT, and GPx) in the HEp-2 cells. In general, we demonstrated that carvacrol, oregano extract, and rifaximin at sub-minimal bactericidal concentrations interfere with target sites in E. coli, reducing the adhesion efficiency.

Characterization of antimicrobial susceptibility, extended-spectrum ß-lactamase genes and phylogenetic groups of Shigatoxin producing Escherichia coli isolated from patients with diarrhea in Iran.

BACKGROUND: Shiga toxin-producing Escherichia coli (STEC) are among common foodborne bacterial pathogens and healthy livestock are the main source of this bacterium. Severe diseases attribute to two types of cytotoxin Stx1 and Stx2, which are also called Shiga toxin (Stx). Infection of humans with STEC may result in Acute diarrhea with or without bleeding, hemorrhagic colitis (HC) and the hemolytic uremic syndrome (HUS). As antibiotic resistance is increasingly being reported among STEC isolates obtained from livestock and patients worldwide, in this study the pattern of antibiotic resistance in clinical isolates was determined. METHODS: Stool samples were collected from patients with diarrhea. All samples were cultured and identified by biochemical and molecular tests. Antimicrobial susceptibility test and assessment of extended-spectrum ß-lactamase (ESBL)-related genes were conducted. Moreover, phylogenetic groups were analyzed using quadruplex PCR, and DNA analysis assessed multi-locus sequence types (MLST). RESULTS: Out of 340 E. coli samples, 174 were identified as STEC by PCR. Antimicrobial susceptibility test results showed that, 99.4%, 96% and 93.1% of isolates were susceptible to imipenem/ertapenem, piperacillin-tazobactam and amikacin, respectively. The highest resistance was towards ampicillin (68.4%), followed by trimethoprim-sulfamethoxazole (59.8%), and tetracycline (57.5%). A total of 106 (60.9%) isolates were multidrug resistance (MDR) and 40.8% of isolates were determined to be extended spectrum ß-lactamase producers. In 94.4% of isolates, genes responsible for ESBL production could be detected, and blaTEM was the most prevalent, followed by blaCTX-M9. Furthermore, phylogenetic grouping revealed that majority of STEC strains belonged to Group C, followed by Groups E, B2 and A. MLST unveiled diverse ST types. CONCLUSION: A periodical surveillance studies and thorough understanding of antibiotic resistant profiles in STEC isolates could help select effective antibiotic treatment for patients and develop strategies to effectively manage food contamination and human infections.

Prevalence, characterization and antibiotic resistance of Shiga toxigenic Escherichia coli serogroups isolated from fresh beef and locally processed ready-to-eat meat products in Lagos, Nigeria.

Fresh beef and meat products have been implicated in outbreaks of Shiga toxin-producing Escherichia coli (STEC) worldwide. This study investigated the prevalence of E. coli O157: H7 and non-O157 STEC serogroups in fresh beef in the open market and street vended meat products (n = 180) in Lagos metropolis, Nigeria. A combination of culture media and immunomagnetic separation followed by typing for associated virulence factors and serotypes was performed. Antimicrobial susceptibility testing was performed on the isolated STEC serotypes using the disk diffusion method. A total of 72 STEC serogroup isolates were detected from 61 out of 180 samples. The O157 STEC serotypes were detected in fresh beef, suya, minced meat and tsire with prevalence of 20.8% while non-O157 STEC serogroups were detected in all the samples. Molecular typing revealed 25% (n = 18) of the STEC serogroups showed presence of all the stx1, stx2, eaeA, fliCH7 and rfbEO157 virulence factors while 54.2% (n = 39) possessed a combination of two virulence genes. Multidrug resistance was discovered in 23.6% (n = 17) of the total STEC serogroups. Locally processed ready-to-eat meat products in Lagos metropolis, Nigeria harbour potentially pathogenic multi-drug resistant STEC serogroups that can constitute public health hazard.

Sensitivity of wild-type and rifampicin-resistant O157 and non-O157 Shiga toxin-producing Escherichia coli to elevated hydrostatic pressure and lactic acid in ground meat and meat homogenate.

Various serogroups of Shiga toxin-producing Escherichia coli have been epidemiologically associated with foodborne disease episodes in the United States and around the globe, with E. coli O157: H7 as the dominant serogroup of public health concern. Serogroups other than O157 are currently associated with about 60% of Shiga toxin-producing E. coli related foodborne illness episodes. Current study evaluated sensitivity of the O157 and epidemiologically important non-O157 serogroups of the pathogen to elevated hydrostatic pressure and 1% lactic acid. Pressure intensity of 250 to 650 MPa were applied for 0 to 7 min for inactivation of strain mixtures of wild-type and rifampicin-resistant E. coli O157, as well as O26, O45, O103, O111, O121, and O145 serogroups and ATCC® 43895™ strain in ground meat and 10% meat homogenate. E. coli O157 were reduced (p < 0.05) from 6.86 ± 0.2 to 4.56 ± 0.1 log CFU/g when exposed to pressure of 650 MPa for 7 min. Corresponding reductions (p < 0.05) for non-O157 E. coli were from 6.98 ± 0.3 to 4.72 ± 0.1. The D-values at 650 MPa were 3.71 and 3.47 min for O157 and non-O157 serogroups, respectively. Presence of 1% lactic acid to a great extent augmented (p < 0.05) decontamination efficacy of the treatment in meat homogenate resulting in up to 5.6 and 6.0 log CFU/mL reductions for O157 and non-O157 serogroups, respectively. Among the tested serogroups, the wild-type and rifampicin-resistant phenotypes exhibited (p ≥ 0.05) comparable pressure sensitivity. Thus, these two phenotypes could be used interchangeably in validation studies. Our results also illustrate that, application of elevated hydrostatic pressure could be utilized for assuring safety of ground and non-intact meat products against various serogroups of Shiga toxin-producing E. coli. Addition of 1% lactic acid additionally provided industrially appreciable augmentation in efficacy of the pressure-based treatments.

Shiga Toxin-Producing Escherichia coli in the Long-Term Survival Phase Exhibit Higher Chlorine Tolerance and Less Sublethal Injury Following Chlorine Treatment of Romaine Lettuce.

The extent of chlorine inactivation and sublethal injury of stationary-phase (STAT) and long-term survival-phase (LTS) cells of Shiga toxin-producing Escherichia coli (STEC) in vitro and in a lettuce postharvest wash model was investigated. Four STEC strains were cultured in tryptic soy broth supplemented with 0.6% (w/v) yeast extract (TSBYE; 35°C) for 24 h and 21 d to obtain STAT and LTS cells, respectively. Minimum bactericidal concentration (MBC) and dose-response assays were performed to determine chlorine's antibacterial efficacy against STAT and LTS cells. Chlorine solutions (pH 6.5) and romaine lettuce were each inoculated with STAT and LTS cells to obtain initial populations of ∼7.8 log colony-forming units (CFU)/mL. Survivors in chlorine solutions were determined after 30 s. Inoculated lettuce samples were held at 22°C ± 1°C for 2 h or 20 h and then exposed to chlorine (10-40 ppm) for 60 s. Survivors were enumerated on nonselective and selective agar media following incubation (35°C, 48 h). The MBC for STAT and LTS cells was 0.04 and 0.08 ppm, respectively. Following exposure (30 s) to chlorine at 2.5, 5.0, and 10 ppm, STAT cells were reduced to <1.0 log CFU/mL, whereas LTS survivors were at 5.10 (2.5 ppm), 3.71 (5.0 ppm), and 2.55 (10 ppm) log CFU/mL. At 20 and 40 ppm chlorine, greater log CFU reductions of STAT cells (1.64 and 1.85) were observed compared with LTS cells (0.94 and 0.83) after 2 h of cell contact with lettuce (p < 0.05), but not after 20 h. Sublethal injury in STEC after chlorine (40 ppm) treatment was lower in LTS compared with STAT survivors (p < 0.05). Compared with STAT cells, LTS cells of STEC seem to have higher chlorine tolerance as planktonic cells and as attached cells depending on cell contact time on lettuce. In addition, a higher percentage of LTS cells, compared with STAT cells, survive in a noninjured state after chlorine (40 ppm) treatment of lettuce.

Incidence of Shiga toxin-producing Escherichia coli in diarrheic calves and its susceptibility profile to antimicrobials and Eugenia uniflora L.

The aim of this study was to evaluate the occurrence of Shiga toxin (stx)-producing Escherichia coli (STEC) in diarrheic newborn calves, as well as the resistance profile of this microorganism against antimicrobials routinely used in veterinary therapy. The antimicrobial profile of Eugenia uniflora against E. coli clinical isolates was also analyzed. Specimens from the recto-anal junction mucosa were investigated by using chromogenic medium and identification of E. coli was done using microbiological methods (Gram staining, indole test, methyl red test, Voges-Proskauer test, citrate test, urease test, and hydrogen sulfide test). The stx1 and stx2 genes corresponding to the STEC pathotype were evaluated by using polymerase chain reaction and electrophoresis. The susceptibility profile to antimicrobial agents commonly used in veterinary therapeutic practice and the antimicrobial effect of lyophilized hydroalcoholic extract of E. uniflora L. leaves against E. coli clinical isolates were evaluated by disk diffusion and microdilution methods. Shiga toxin-positive E. coli was identified in 45% of diarrheic newborn calves (stx1 = 23.2%, stx2 = 4.0%, stx1 + stx2 = 18.2%). The frequency of stx-positive E. coli in the bacterial population was equal to 17.0% (168/990 clinical isolates): 97 (9.8%) stx1-positive E. coli, 12 (1.2%) stx2-positive E. coli, and 59 (6.0%) stx1 + stx2-positive E. coli isolates. All stx-positive E. coli analyzed showed resistance to multiple drugs, that is, from 4 to 10 antimicrobials per clinical isolate (streptomycin, tetracycline, cephalothin, ampicillin, sulfamethoxazole + trimethoprim, nitrofurantoin and nalidixic acid, ciprofloxacin, gentamicin, and chloramphenicol). Effective management measures should be implemented, including clinical and laboratory monitoring, in order to promote animal and worker health and welfare, prevent and control the spread of diseases, and ensure effective treatment of infectious diseases. The E. uniflora L. leaves showed inhibition of microbial growth based on the diameter of halos, ranging from 7.9 to 8.0 mm and 9.9 to 10.1 mm for concentrations of 50 and 150 mg/mL, respectively. This plant displayed bacteriostatic action and a minimum inhibitory concentration of 12.5 mg/mL for all clinical isolates. Its clinical or synergistic effects with antimicrobial agents must be determined from clinical and preclinical trials.

Le but de cette étude était d'évaluer la présence d'Escherichia coli (STEC) productrices de Shiga toxine (stx) chez les veaux nouveaunés diarrhéiques, ainsi que le profil de résistance de ce microorganisme aux antimicrobiens couramment utilisés en thérapie vétérinaire. Le profil antimicrobien d'Eugenia uniflora contre les isolats cliniques d'E. coli a également été analysé. Des échantillons de la muqueuse de la jonction recto-anale ont été étudiés en utilisant un milieu chromogène et l'identification d'E. coli a été effectuée à l'aide de méthodes microbiologiques (coloration de Gram, test à l'indole, test au rouge de méthyle, test de Voges-Proskauer, test de citrate, test d'uréase et production de sulfure d'hydrogène). Les gènes stx1 et stx2 correspondant au pathotype STEC ont été évalués en utilisant la réaction en chaîne par polymérase et l'électrophorèse. Le profil de sensibilité aux agents antimicrobiens couramment utilisés dans la pratique thérapeutique vétérinaire et l'effet antimicrobien de l'extrait hydroalcoolique lyophilisé de feuilles d'E. uniflora L. contre les isolats cliniques d'E. coli ont été évalués par des méthodes de diffusion en disque et de microdilution. Des E. coli positifs à la Shiga toxine ont été identifiés chez 45 % des veaux nouveau-nés diarrhéiques (stx1 = 23,2 %, stx2 = 4,0 %, stx1 + stx2 = 18,2 %). La fréquence des E. coli stx-positifs dans la population bactérienne était égale à 17,0 % (168/990 isolats cliniques) : 97 (9,8 %) E. coli positifs pour stx1, 12 (1,2 %) E. coli positifs pour stx2, et 59 isolats d'E. coli positifs pour stx1 + stx2 (6,0 %). Tous les E. coli stx positifs analysés ont montré une résistance à plusieurs médicaments, à savoir de 4 à 10 antimicrobiens par isolat clinique (streptomycine, tétracycline, céphalothine, ampicilline, sulfaméthoxazole + triméthoprime, nitrofurantoïne et acide nalidixique, ciprofloxacine, gentamicine et chloramphénicol). Des mesures de gestion efficaces devraient être mises en oeuvre, y compris une surveillance clinique et de laboratoire, afin de promouvoir la santé et le bien-être des animaux et des travailleurs, de prévenir et de contrôler la propagation des maladies et de garantir un traitement efficace des maladies infectieuses. Les feuilles d'E. uniflora L. ont montré une inhibition de la croissance microbienne basée sur le diamètre des zones, allant de 7,9 à 8,0 mm et de 9,9 à 10,1 mm pour des concentrations de 50 et 150 mg/mL, respectivement. Cette plante a montré une action bactériostatique et une concentration inhibitrice minimale de 12,5 mg/mL pour tous les isolats cliniques. Ses effets cliniques ou synergiques avec les agents antimicrobiens doivent être déterminés à partir d'essais cliniques et précliniques.(Traduit par Docteur Serge Messier).

Modeling the effect of simultaneous use of allyl isothiocyanate and cinnamaldehyde on high hydrostatic pressure inactivation of Uropathogenic and Shiga toxin-producing Escherichia coli in ground chicken.

BACKGROUND: A combination of high-pressure processing (HPP) and antimicrobials is a well-known approach for enhancing the microbiological safety of foods. However, few studies have applied multiple antimicrobials simultaneously with HPP, which could be an additional hurdle for microbial inactivation. The present study applied a full factorial design to investigate the impact of HPP (225-325 MPa; 10-20 min), allyl isothiocyanate (AITC) (0.3-0.9 g kg-1 ) and trans-cinnamaldehyde (tCinn) (1.0-2.0 g kg-1 ) on the inactivation of Shiga toxin-producing Escherichia coli (STEC) O157:H7 and uropathogenic E. coli (UPEC) in ground chicken meat. RESULTS: The regulatory requirement of 5-log reduction was achieved at 305 MPa, 18 min, 0.8 g kg-1 AITC and 1.7 g kg-1 tCinn for STEC O157:H7 and at 293 MPa, 16 min, 0.6 g kg-1 AITC and 1.6 g kg-1 tCinn for UPEC, as specified by response surface analysis and verified via experiments. The surviving population was eliminated by post-treatment storage of 9 days at 10 °C. The developed linear regression models showed r2 > 0.9 for the E. coli inactivation. The developed dimensionless non-linear regression models covered a factorial range slightly wider than the original experimental limit, with probability Pr > F (< 0.0001). CONCLUSION: Simultaneous use of AITC and tCinn reduced not only the necessary concentration of each compound, but also the intensity of high-pressure treatments, at the same time achieving a similar level of microbial inactivation. STEC O157:H7 was found to be more resistant than UPEC to the HPP-AITC-tCinn stress. The developed models may be applied in commercial application to enhance the microbiological safety of ground chicken meat. Published 2020. This article is a U.S. Government work and is in the public domain in the USA.

Efficacy of antimicrobial interventions in reducing Salmonella enterica, Shiga toxin-producing Escherichia coli, Campylobacter, and Escherichia coli biotype I surrogates on non-chilled and chilled, skin-on and skinless pork.

Effect of various antimicrobial interventions on pork carcass cuts - skin-on and skinless, non-chilled and chilled - was studied. Carcass pieces were inoculated with Salmonella enterica, Shiga toxin-producing Escherichia coli (STEC), Escherichia coli pathogen surrogates or Campylobacter spp. Inoculated pieces were assigned to one of the following antimicrobial treatments: 2.5% and 5.0% room temperature lactic acid, 2.5% and 5.0% warm (55 °C) lactic acid, 400 ppm (0.4 mg/mL) room temperature peroxyacetic acid (PAA) or warm (55 °C) water. Treated pieces were sampled before antimicrobial treatment of non-chilled pork tissue, then at 30 m and 24 h post-treatment. For chilled pork, samples were collected after 24 h chilling and 30 m post-treatment. Lactic acid and PAA treatments reduced (P < 0.05) pathogenic and surrogate bacteria; warm water did not produce similar results. Objective and sensory color evaluations on treated pork indicated minimal negative impacts on pork color. Various antimicrobial interventions were effective in reducing surrogates on pork without diminishing quality.