Uropathogenic Escherichia coli infection: innate immune disorder, bladder damage, and Tailin Fang II.

Background: Uropathogenic Escherichia coli (UPEC) activates innate immune response upon invading the urinary tract, whereas UPEC can also enter bladder epithelial cells (BECs) through interactions with fusiform vesicles on cell surfaces and subsequently escape from the vesicles into the cytoplasm to establish intracellular bacterial communities, finally evading the host immune system and leading to recurrent urinary tract infection (RUTI). Tailin Fang II (TLF-II) is a Chinese herbal formulation composed of botanicals that has been clinically proven to be effective in treating urinary tract infection (UTI). However, the underlying therapeutic mechanisms remain poorly understood. Methods: Network pharmacology analysis of TLF-II was conducted. Female Balb/C mice were transurethrally inoculated with UPEC CFT073 strain to establish the UTI mouse model. Levofloxacin was used as a positive control. Mice were randomly divided into four groups: negative control, UTI, TLF-II, and levofloxacin. Histopathological changes in bladder tissues were assessed by evaluating the bladder organ index and performing hematoxylin-eosin staining. The bacterial load in the bladder tissue and urine sample of mice was quantified. Activation of the TLR4-NF-κB pathway was investigated through immunohistochemistry and western blotting. The urinary levels of interleukin (IL)-1ß and IL-6 and urine leukocyte counts were monitored. We also determined the protein expressions of markers associated with fusiform vesicles, Rab27b and Galectin-3, and levels of the phosphate transporter protein SLC20A1. Subsequently, the co-localization of Rab27b and SLC20A1 with CFT073 was examined using confocal fluorescence microscopy. Results: Data of network pharmacology analysis suggested that TLF-II could against UTI through multiple targets and pathways associated with innate immunity and inflammation. Additionally, TLF-II significantly attenuated UPEC-induced bladder injury and reduced the bladder bacterial load. Meanwhile, TLF-II inhibited the expression of TLR4 and NF-κB on BECs and decreased the urine levels of IL-1ß and IL-6 and urine leukocyte counts. TLF-II reduced SLC20A1 and Galectin-3 expressions and increased Rab27b expression. The co-localization of SLC20A1 and Rab27b with CFT073 was significantly reduced in the TLF-II group. Conclusion: Collectively, innate immunity and bacterial escape from fusiform vesicles play important roles in UPEC-induced bladder infections. Our findings suggest that TLF-II combats UPEC-induced bladder infections by effectively mitigating bladder inflammation and preventing bacterial escape from fusiform vesicles into the cytoplasm. The findings suggest that TLF-II is a promising option for treating UTI and reducing its recurrence.

Plant phenolics inhibit focal adhesion kinase and suppress host cell invasion by uropathogenic Escherichia coli.

Traditional folk treatments for the prevention and management of urinary tract infections (UTIs) and other infectious diseases often include plants and plant extracts that are rich in phenolic compounds. These have been ascribed a variety of activities, including inhibition of bacterial interactions with host cells. Here, we tested a panel of four well-studied phenolic compounds-caffeic acid phenethyl ester (CAPE), resveratrol, catechin, and epigallocatechin gallate-for the effects on host cell adherence and invasion by uropathogenic Escherichia coli (UPEC). These bacteria, which are the leading cause of UTIs, can bind and subsequently invade bladder epithelial cells via an actin-dependent process. Intracellular UPEC reservoirs within the bladder are often protected from antibiotics and host defenses and likely contribute to the development of chronic and recurrent infections. In cell culture-based assays, only resveratrol had a notable negative effect on UPEC adherence to bladder cells. However, both CAPE and resveratrol significantly inhibited UPEC entry into the host cells, coordinate with attenuated phosphorylation of the host actin regulator Focal Adhesion Kinase (FAK or PTK2) and marked increases in the numbers of focal adhesion structures. We further show that the intravesical delivery of resveratrol inhibits UPEC infiltration of the bladder mucosa in a murine UTI model and that resveratrol and CAPE can disrupt the ability of other invasive pathogens to enter host cells. Together, these results highlight the therapeutic potential of molecules like CAPE and resveratrol, which could be used to augment antibiotic treatments by restricting pathogen access to protective intracellular niches.IMPORTANCEUrinary tract infections (UTIs) are exceptionally common and increasingly difficult to treat due to the ongoing rise and spread of antibiotic-resistant pathogens. Furthermore, the primary cause of UTIs, uropathogenic Escherichia coli (UPEC), can avoid antibiotic exposure and many host defenses by invading the epithelial cells that line the bladder surface. Here, we identified two plant-derived phenolic compounds that disrupt activation of the host machinery needed for UPEC entry into bladder cells. One of these compounds, resveratrol, effectively inhibited UPEC invasion of the bladder mucosa in a mouse UTI model, and both phenolic compounds significantly reduced host cell entry by other invasive pathogens. These findings suggest that select phenolic compounds could be used to supplement existing antibacterial therapeutics by denying uropathogens shelter within host cells and tissues and help explain some of the benefits attributed to traditional plant-based medicines.

Dopamine receptor D2 confers colonization resistance via microbial metabolites.

The gut microbiome has major roles in modulating host physiology. One such function is colonization resistance, or the ability of the microbial collective to protect the host against enteric pathogens1-3, including enterohaemorrhagic Escherichia coli (EHEC) serotype O157:H7, an attaching and effacing (AE) food-borne pathogen that causes severe gastroenteritis, enterocolitis, bloody diarrhea and acute renal failure4,5 (haemolytic uremic syndrome). Although gut microorganisms can provide colonization resistance by outcompeting some pathogens or modulating host defence provided by the gut barrier and intestinal immune cells6,7, this phenomenon remains poorly understood. Here, we show that activation of the neurotransmitter receptor dopamine receptor D2 (DRD2) in the intestinal epithelium by gut microbial metabolites produced upon dietary supplementation with the essential amino acid L-tryptophan protects the host against Citrobacter rodentium, a mouse AE pathogen that is widely used as a model for EHEC infection8,9. We further find that DRD2 activation by these tryptophan-derived metabolites decreases expression of a host actin regulatory protein involved in C. rodentium and EHEC attachment to the gut epithelium via formation of actin pedestals. Our results reveal a noncanonical colonization resistance pathway against AE pathogens that features an unconventional role for DRD2 outside the nervous system in controlling actin cytoskeletal organization in the gut epithelium. Our findings may inspire prophylactic and therapeutic approaches targeting DRD2 with dietary or pharmacological interventions to improve gut health and treat gastrointestinal infections, which afflict millions globally.

Three different routes of EHEC O157:H7 infection were used to establish EHEC broiler model.

In order to study the prevention and control EHEC disease measures in poultry, the infection process and development of this disease and the pathological changes of various organs were to be observed. In this study, chickens were infected with different doses of enterohemorrhagic Escherichia coli (EHEC) O157:H7 using different routes of administration to establish EHEC broiler model. A total of 195 14-day-old broilers were randomly divided into 13 groups: including control group, Enema-drip groups (1010, 1011, 1012, 1013 CFUs E. coli O157:H7), gavage groups (P.O) (1011, 1012, 1013, 1014 CFUs E. coli O157:H7), and intraperitoneal injection group (I.P.) (108, 109, 1010, 1011 CFUs E. coli O157:H7). Escherichia coli (E. coli) was given using enema-drip, gavage or intraperitoneal infection. Then the feed intake, weight changes, stool and clinical symptoms of the chicks were recorded during the experiment. 7 d after E. coli infection, blood was collected from the jugular vein and serological tests were carried out. The liver, spleen, and colon of the chicks were extracted to get the organ index, bacteria load, and their histopathological changes. After infection with E. coli, some chicks feces were green or red watery stool, sometimes accompanied by foam, and the material to weight ratio of broilers in I.P. group increased significantly (P < 0.05), the 108 CFUs group were 1.3 times as large as control group. Three modeling methods can result in abnormal serum lipid metabolism and liver function indexes (increase of AST, TBA, T-Bil and TC level; decrease of ALB, TG, and TP level). Infection of chicks with O157:H7 by all 3 methods resulted in its detection in the liver, spleen, and colon. Three modeling methods significantly decreased liver index, and inflammatory cell infiltration and hyperemia were observed in liver. The spleen index in E. coli broilers by gavage and enema-drip was significantly decreased, splenic hyperemia and periarteriolar hyalinosis were observed. The spleen was enlarged with purplish-black spheroids in I.P. group broilers, and the spleen histological changes was more serious. The colon villi of broilers in gavage and enema-drip groups were thinner, more prone to rupture, intestinal lamina propria hyperemia, and inflammatory cell infiltration. Moreover, the number of goblet cells in the mucosal epithelium increased. E. coli O157:H7 can induce liver, spleen and intestinal damage and reduce growth performance of chicks. By comparing these 3 methods, we found that chicks infected with O157:H7 by gavage had more severe liver and intestinal damage, the enema-drip method caused most serious intestinal damage, and I.P. method significantly damaged the liver and spleen of chickens.

Comparison of three doses of amikacin on alternate days with a daily dose of meropenem during the same period for the treatment of urinary tract infection with E. coli: a double-blind clinical trial.

OBJECTIVES: Urinary tract infections (UTIs) are very common infections in humans, and Escherichia coli (E. coli) is the commonest pathogen leading to UTIs. The generation of beta-lactamase enzymes in this bacterium results in its resistance against many antibiotics. This study compares three doses of amikacin on alternate days with a daily dose of meropenem in the same period for the treatment of UTIs with E. coli in a double-blind clinical trial. METHODS: The current double-blind clinical trial compares three doses of amikacin on alternate days with a daily dose of meropenem in the same period for the treatment of UTIs with E. coli. The patients were assigned to two groups: Intervention (receiving a single dose of amikacin once a day at 48-h intervals for a week, three doses) and control (receiving meropenem for 1/TDS for a week). RESULTS: The E. coli infection frequency was 61 (21 cases of non-ESBL and 40 cases of ESBL-positive infections) and the frequency of the other infections was 52 (46%). In the patients with ESBL E. coli infection, ciprofloxacin (21; 70%) showed the highest antibiotic resistance, and nitrofurantoin (33; 91.7%) showed the highest sensitivity. The baseline variables between the control and intervention groups indicated no significant difference (p > 0.05). The frequency of signs and symptoms showed no significant difference between the amikacin and meropenem groups in the first 24 h and the first week. In the second week of follow-up, no clinical signs or symptoms were observed in the two groups. CONCLUSION: The results of this study showed that treatment with amikacin, 1 g q48h, for one week (three doses) has the same result as meropenem, 1 g q8h, for one week (21 doses). The results are the same for the treatment of UTIs with ESBL positive and ESBL negative. Amikacin can be used once every 48 h to treat UTIs, is less expensive and can be administered on an outpatient basis. TRIAL REGISTRATION: This study was registered in the Iranian Registry of Clinical Trials (IRCT) with ID number: IRCT20170417033483N2 on the date 2018-02-13.

Bacillus coagulans prevents the decline in average daily feed intake in young piglets infected with enterotoxigenic Escherichia coli K88 by reducing intestinal injury and regulating the gut microbiota.

Background: Enterotoxigenic Escherichia coli (ETEC), an important intestinal pathogen, poses a significant threat to the intestinal health of piglets. Bacillus coagulans (BC), a potential feed additive, can improve the intestinal function of piglets. However, the effects of BC on growth performance and intestinal function in ETEC-infected piglets are still unclear. In this study, 24 7-day-old piglets were randomly assigned to three treatment groups: control group (fed a basal diet), ETEC group (fed a basal diet and challenged with ETEC K88) and BC+ETEC group (fed a basal diet, orally administered BC, challenged with ETEC K88). During Days 1-6 of the trial, piglets in the BC+ETEC group were orally administered BC (1×108CFU/kg). On Day 5 of the trial, piglets in the ETEC and BC+ETEC groups were orally administered ETEC K88 (5×109CFU/piglet). Blood, intestinal tissue, and content samples were collected from the piglets on Day 7 of the trial. Results: The average daily feed intake in the ETEC group was significantly reduced compared to that of the control group. Further research revealed that ETEC infection significantly damaged the structure of the small intestine. Compared to the control group, the villus height and surface area of the jejunum, the ratio of villus height to crypt depth in the duodenum and jejunum, and the activities of catalase and total superoxide dismutase in the jejunum were significantly reduced. Additionally, the levels of myeloperoxidase in the jejunum, malondialdehyde in the plasma and jejunum, and intestinal epithelial apoptosis were significantly increased in the ETEC group. However, BC supplementation had significantly mitigated these negative effects in the BC+ETEC group by Day 7 of the trial. Moreover, BC supplementation improved the gut microbiota imbalance by reversing the decreased numbers of Enterococcus, Clostridium and Lactobacillus in jejunum and Escherichia coli, Bifidobacterium and Lactobacillus in the colon, as well as the increased number of Escherichia coli in the jejunum induced by ETEC K88. Conclusions: Overall, BC supplementation reduced the decline in average daily feed intake in ETEC K88-infected piglets by attenuating intestinal epithelial apoptosis and oxidative stress and regulating the gut microbiota. This suggests that BC may be used to prevent intestinal infections caused by ETEC in piglets.

The scope of antimicrobial resistance in residential aged care facilities determined through analysis of Escherichia coli and the total wastewater resistome.

IMPORTANCE: Antimicrobial resistance (AMR) is a global threat that imposes a heavy burden on our health and economy. Residential aged care facilities (RACFs), where frequent inappropriate antibiotic use creates a selective environment that promotes the development of bacterial resistance, significantly contribute to this problem. We used wastewater-based epidemiology to provide a holistic whole-facility assessment and comparison of antimicrobial resistance in two RACFs and a retirement village. Resistant Escherichia coli, a common and oftentimes problematic pathogen within RACFs, was isolated from the wastewater, and the phenotypic and genotypic AMR was determined for all isolates. We observed a high prevalence of an international high-risk clone, carrying an extended-spectrum beta-lactamase in one facility. Analysis of the entire resistome also revealed a greater number of mobile resistance genes in this facility. Finally, both facilities displayed high fluoroquinolone resistance rates-a worrying trend seen globally despite measures in place aimed at limiting their use.

Prevalence and characterization of ESBL-producing Escherichia coli in healthy pregnant women and hospital environments in Benin: an approach based on Tricycle.

Introduction: Extended-Spectrum Beta-Lactamase (ESBL)-producing Enterobacterales are recognized as significant pathogens due to their resistance to multiple antibiotics. This study aimed to determine the prevalence of ESBL-producing Escherichia coli (E. coli) in different settings, including healthy pregnant women, the food chain, and the environment of tertiary hospitals in Benin. Methods: Samples were collected from various sources, including fecal samples from healthy pregnant women, food samples from hospital canteens, and hospital effluents from four tertiary hospitals in southern Benin. Fecal samples were plated on MacConkey agar supplemented with cefotaxime (4 µg/mL), while food and water samples were plated on Tryptone Bile X agar supplemented with cefotaxime (4 µg/mL). Urea indole tests were used for preliminary identification of E. coli colonies, followed by confirmation of ESBL production using the double disk synergy technique. Antibiotic susceptibility testing of ESBL-producing E. coli strains was conducted using the disk diffusion method on MH agar. Polymerase Chain Reaction (PCR) was used to investigate the presence of ESBL-encoding genes. Results: Among the 296 fecal samples collected from four tertiary hospitals, ESBL-producing E. coli was isolated from 22.30% (66) of the samples. All E. coli isolates from hospital effluents exhibited ESBL production, while ESBL-producing E. coli was not detected in food and drinking water samples. The analysis of variable associations showed no significant associations (p > 0.05) for the studied factors. Antibiotic susceptibility testing revealed high resistance rates among the ESBL-Ec isolates against several tested antibiotics, including amoxicillin, aztreonam, ceftriaxone, ciprofloxacin, and trimethoprim-sulfamethoxazole. However, most isolates remained susceptible to ertapenem, amoxicillin-clavulanate, and imipenem. The most prevalent ESBL-encoding genes were blaTEM (37.50%), blaOXA-1 (19.44%), and blaSHV (11.11%), while a smaller proportion of isolates carried blaCTXM-1/blaCTXM-15 (5.55%) and blaCTXM-9. Discussion: This study provides insights into the prevalence of ESBL-producing E. coli carriage in the feces of healthy pregnant women in southern Benin. Additionally, it highlights hospital wastewater as a potential reservoir of ESBL-producing bacteria in the environment. The detection of ESBL-producing E. coli in hospital effluents raises concerns about the dissemination of antibiotic resistance genes into the environment. The high resistance rates observed among ESBL-Ec isolates against commonly used antibiotics emphasize the urgent need for antimicrobial stewardship and infection control measures. The identification of prevalent ESBL-encoding genes contributes to understanding the genetic basis of ESBL resistance in the studied population. Further research is warranted to explore the mechanisms of transmission and potential interventions to mitigate the spread of ESBL-producing Enterobacterales.

An effective antimicrobial strategy of colistin combined with the Chinese herbal medicine shikonin against colistin-resistant Escherichia coli.

IMPORTANCE: Infections caused by multidrug-resistant Escherichia coli (MDR E. coli) have become a major global healthcare problem due to the lack of effective antibiotics today. The emergence of colistin-resistant E. coli strains makes the situation even worse. Therefore, new antimicrobial strategies are urgently needed to combat colistin-resistant E. coli. Combining traditional antibiotics with non-antibacterial drugs has proved to be an effective approach of combating MDR bacteria. This study investigated the combination of colistin and shikonin, a Chinese herbal medicine, against colistin-resistant E. coli. This combination showed good synergistic antibacterial both in vivo and in vitro experiments. Under the background of daily increasing colistin resistance in E. coli, this research points to an effective antimicrobial strategy of using colistin and shikonin in combination against colistin-resistant E. coli.

Fimbria targeting superparamagnetic iron oxide nanoparticles enhance the antimicrobial and antibiofilm activity of ciprofloxacin against quinolone-resistant E. coli.

High quinolone resistance of Escherichia coli limits the therapy options for urinary tract infection (UTI). In response to the urgent need for efficient treatment of multidrug-resistant infections, we designed a fimbriae targeting superparamagnetic iron oxide nanoparticle (SPION) delivering ciprofloxacin to ciprofloxacin-resistant E. coli. Bovine serum albumin (BSA) conjugated poly(acrylic acid) (PAA) coated SPIONs (BSA@PAA@SPION) were developed for encapsulation of ciprofloxacin and the nanoparticles were tagged with 4-aminophenyl-α-D-mannopyrannoside (mannoside, Man) to target E. coli fimbriae. Ciprofloxacin-loaded mannoside tagged nanoparticles (Cip-Man-BSA@PAA@SPION) provided high antibacterial activity (97.1 and 97.5%, respectively) with a dose of 32 µg/mL ciprofloxacin against two ciprofloxacin-resistant E. coli isolates. Furthermore, a strong biofilm inhibition (86.9% and 98.5%, respectively) was achieved in the isolates at a dose 16 and 8 times lower than the minimum biofilm eradication concentration (MBEC) of ciprofloxacin. Weaker growth inhibition was observed with untargeted nanoparticles, Cip-BSA@PAA@SPIONs, confirming that targeting E. coli fimbria with mannoside-tagged nanoparticles increases the ciprofloxacin efficiency to treat ciprofloxacin-resistant E. coli. Enhanced killing activity against ciprofloxacin-resistant E. coli planktonic cells and strong growth inhibition of their biofilms suggest that Cip-Man-BSA@PAA@SPION system might be an alternative and/or complementary therapeutic option for the treatment of quinolone-resistant E. coli infections.

Urinary Tract Infections Caused by Uropathogenic Escherichia coli: Mechanisms of Infection and Treatment Options.

Urinary tract infections (UTIs) are common bacterial infections that represent a severe public health problem. They are often caused by Escherichia coli (E. coli), Klebsiella pneumoniae (K. pneumonia), Proteus mirabilis (P. mirabilis), Enterococcus faecalis (E. faecalis), and Staphylococcus saprophyticus (S. saprophyticus). Among these, uropathogenic E. coli (UPEC) are the most common causative agent in both uncomplicated and complicated UTIs. The adaptive evolution of UPEC has been observed in several ways, including changes in colonization, attachment, invasion, and intracellular replication to invade the urothelium and survive intracellularly. While antibiotic therapy has historically been very successful in controlling UTIs, high recurrence rates and increasing antimicrobial resistance among uropathogens threaten to greatly reduce the efficacy of these treatments. Furthermore, the gradual global emergence of multidrug-resistant UPEC has highlighted the need to further explore its pathogenesis and seek alternative therapeutic and preventative strategies. Therefore, a thorough understanding of the clinical status and pathogenesis of UTIs and the advantages and disadvantages of antibiotics as a conventional treatment option could spark a surge in the search for alternative treatment options, especially vaccines and medicinal plants. Such options targeting multiple pathogenic mechanisms of UPEC are expected to be a focus of UTI management in the future to help combat antibiotic resistance.

Isolation and Characterization of a Novel Phage Collection against Avian-Pathogenic Escherichia coli.

The increase in antibiotic-resistant avian-pathogenic Escherichia coli (APEC), the causative agent of colibacillosis in poultry, warrants urgent research and the development of alternative therapies. This study describes the isolation and characterization of 19 genetically diverse, lytic coliphages, 8 of which were tested in combination for their efficacy in controlling in ovo APEC infections. Genome homology analysis revealed that the phages belong to nine different genera, one of them being a novel genus (Nouzillyvirus). One phage, REC, was derived from a recombination event between two Phapecoctavirus phages (ESCO5 and ESCO37) isolated in this study. Twenty-six of the 30 APEC strains tested were lysed by at least one phage. Phages exhibited varying infectious capacities, with narrow to broad host ranges. The broad host range of some phages could be partially explained by the presence of receptor-binding protein carrying a polysaccharidase domain. To demonstrate their therapeutic potential, a phage cocktail consisting of eight phages belonging to eight different genera was tested against BEN4358, an APEC O2 strain. In vitro, this phage cocktail fully inhibited the growth of BEN4358. In a chicken lethality embryo assay, the phage cocktail enabled 90% of phage-treated embryos to survive infection with BEN4358, compared with 0% of nontreated embryos, indicating that these novel phages are good candidates to successfully treat colibacillosis in poultry. IMPORTANCE Colibacillosis, the most common bacterial disease affecting poultry, is mainly treated by antibiotics. Due to the increased prevalence of multidrug-resistant avian-pathogenic Escherichia coli, there is an urgent need to assess the efficacy of alternatives to antibiotherapy, such as phage therapy. Here, we have isolated and characterized 19 coliphages that belong to nine phage genera. We showed that a combination of 8 of these phages was efficacious in vitro to control the growth of a clinical isolate of E. coli. Used in ovo, this phage combination allowed embryos to survive APEC infection. Thus, this phage combination represents a promising treatment for avian colibacillosis.

Uropathogenic Escherichia coli endeavors: an insight into the characteristic features, resistance mechanism, and treatment choice.

Uropathogenic Escherichia coli (UPEC) are the strains diverted from the intestinal status and account mainly for uropathogenicity. This pathotype has gained specifications in structure and virulence to turn into a competent uropathogenic organism. Biofilm formation and antibiotic resistance play an important role in the organism's persistence in the urinary tract. Increased consumption of carbapenem prescribed for multidrug-resistant (MDR) and Extended-spectrum-beta lactamase (ESBL)-producing UPECs, has added to the expansion of resistance. The World Health Organization (WHO) and Centre for Disease Control (CDC) placed the Carbapenem-resistant Enterobacteriaceae (CRE) on their treatment priority lists. Understanding both patterns of pathogenicity, and multiple drug resistance may provide guidance for the rational use of anti-bacterial agents in the clinic. Developing an effective vaccine, adherence-inhibiting compounds, cranberry juice, and probiotics are non-antibiotical approaches proposed for the treatment of drug-resistant UTIs. We aimed to review the distinguishing characteristics, current therapeutic options and promising non-antibiotical approaches against ESBL-producing and CRE UPECs.

Dietary berberine supplementation improves growth performance and alleviates gut injury in weaned piglets by modulating ileal microbiota and metabolites.

This study investigated the effects of dietary berberine (BBR) supplementation on the growth performance, intestinal health, and ileal microbiome and metabolomic profile in weaned piglets challenged with enterotoxigenic Escherichia coli (ETEC). Dietary BBR supplementation significantly attenuated the reduced average daily gain (ADG) and attenuated the increased feed to gain ratio (F/G) and the incidence of diarrhea induced by ETEC K88 (P < 0.05). Dietary BBR supplementation significantly increased the villus height and the villus height to crypt depth ratio in the ileum (P < 0.05). Moreover, the mRNA expression of ZO-1 and occludin as well as aquaporins (AQP1, AQP3, AQP4, AQP7, and AQP10) and Na+/H+ exchanger 3 (NHE3) in ileal mucosa was significantly upregulated by BBR treatment (P < 0.05). Additionally, BBR treatment significantly inhibited the increase of interleukin-1ß (IL-1ß) in jejunal mucosa caused by ETEC and reduced the levels of tumor necrosis factor-α (TNF-α) and IL-1ß and increased interleukin-10 (IL-10) in colonic mucosa (P < 0.05). Dietary BBR treatment significantly increased the Observed_species, Chao 1, abundance based coverage estimators (ACE), and PD_whole tree in the ileal digesta of weaned piglets challenged with ETEC. At the genus level, the relative abundance of unidentified Clostridiales was decreased, while Weissella, Alloprevotella, unidentified Prevotellaceae, and Catenibacterium were increased in the BBR + ETEC group when compared to the ETEC group (P < 0.05). Spearman correlation analysis showed that the relative abundance of unidentified Clostridiales (genus) was negatively correlated with the ileal villus height but negatively correlated with diarrhea and intestinal IL-1ß and TNF-α concentrations (P < 0.05). The ileal metabolome analysis showed that the metabolic pathways including primary and secondary bile acid biosynthesis and bile secretion were significantly enriched by BBR treatment. Collectively, dietary BBR supplementation effectively improved the growth performance and alleviated the diarrhea and intestinal injury induced by ETEC K88 in weaned piglets, which might closely involve the modulation of ileal microbiota and metabolites.

Monitoring fluoroquinolone resistance among ESBL-positive and ESBL-negative Escherichia coli strains isolated from urinary tract infections: An alert for empirical treatment.

BACKGROUND: Bacterial resistance to extended-spectrum beta-lactamases (ESBL) is present worldwide. Empirical antibiotic therapy is often needed, and the use of fluoroquinolones, such as ciprofloxacin and norfloxacin, is common. This study aimed to analyze the urine cultures from 2,680 outpatients in January 2019, 2020, 2021, and 2022, with bacterial counts above 100,000 CFU/mL in which Escherichia coli was the etiological agent. METHODS: We monitored the resistance of ESBL-positive and ESBL-negative strains to ciprofloxacin and norfloxacin and evaluated resistance rates. RESULTS: Significantly higher fluoroquinolone resistance rates were observed among ESBL-positive strains in all years studied. Furthermore, a significant increase in the rate of fluoroquinolone resistance was observed between 2021 and 2022 in ESBL-positive and -negative strains, as well as from 2020 to 2021 among the ESBL-positive strains. CONCLUSIONS: The data obtained in the present study showed a tendency towards an increase in fluoroquinolone resistance among ESBL-positive and -negative E. coli strains isolated from urine cultures in Brazil. Since empirical antibiotic therapy with fluoroquinolones is commonly used to treat diverse types of infections, such as community-acquired urinary tract infections, this work highlights the need for continuous monitoring of fluoroquinolone resistance among E. coli strains circulating in the community, which can mitigate the frequency of therapeutic failures and development of widespread multidrug-resistant strains.

High prevalence of Escherichia coli co-harboring conjugative plasmids with colistin- and carbapenem resistance genes in a wastewater treatment plant in China.

Emergence and dissemination of resistance to last-resort antibiotics such as carbapenem and colistin is a growing, global health concern. Wastewater treatment plants (WWTPs) link human activities and the environment, can act as reservoirs and sources for emerging antibiotic resistance, and likely play a large role in antibiotic resistance transmission. The aim of this study was to investigate occurrence and characteristics of colistin- and carbapenem-resistant Escherichia coli (CCREC) in wastewater and sludge samples collected over a one-year period from different functional areas of an urban WWTP in Jinan city, Shandong, China. A total of 8 CCREC were isolated from 168 samples with selective agar and PCR, corresponding to high prevalence of 4.8%, co-harboring carbapenem resistance genes (blaNDM) and colistin resistance gene (mcr-1) and subsequently whole-genome sequenced. Additionally, all isolates were multidrug-resistant by antimicrobial susceptibility testing and carried a variety of antibiotic resistance genes. Two isolates carrying virulence genes associated with avian pathogenic E. coli were identified, one belonging to the high-risk clone O101:H9-ST167. Southern blotting was used to characterize CCREC isolates and plasmids carrying blaNDM-genes or mcr-1 could be transferred to a recipient strain E. coli J53 by in vitro conjugation assays. Resistance to other antibiotic classes were sporadically co-transferred to the transconjugant. Transposition of and mcr-1-carrying element from a transferable IncHI2-plasmid was observed among two CCREC clones isolated within 4 days of each other. The occurrence of multidrug-resistant CCREC capable of transferring their antibiotic resistance genotypes via conjugative plasmids is alarming. WWTPs bring bacteria from different sources together, providing opportunities for horizontal exchange of DNA among compatible hosts. Further dissemination of the colistin-, carbapenem-, or both colistin- and carbapenem resistant E. coli could lead to a serious threat to public health.

Efficacy of zinc glycinate reducing zinc oxide on intestinal health and growth of nursery pigs challenged with F18+ Escherichia coli.

The objective of this study was to investigate effects of zinc glycinate (ZnGly) supplementation reducing zinc oxide (ZnO) in feeds on intestinal health and growth of nursery pigs challenged with F18+Escherichia coli (E. coli). In total, 72 nursery pigs (BW 6.5 ± 0.5 kg) were allotted in a randomized complete block design to nine treatments: (1) NC: no challenge/no supplement; (2) PC: E. coli challenge/no-supplement; (3) E. coli challenge/ZnO at 2,500 mg/kg; (4, 5, and 6) E. coli challenge/ZnGly at 400, 800, and 1,200 mg/kg; and (7, 8, and 9) E. coli challenge/ZnGly at 400 mg/kg and ZnO at 700, 1,400, and 2,357 mg/kg. Pigs were fed for 28 d based on two phases (phase 1: 14 d and phase 2: 14 d). On day 7, challenged groups were orally inoculated with F18+E. coli at 6 × 109 CFU/mL whereas NC received saline solution. The PC showed reduced ADG (P = 0.076) and G:F (P = 0.055) during phase 1 and increased fecal score (P < 0.05) during the first week of postchallenge when compared with NC, whereas supplementation of ZnGly from 0 to 1,200 mg/kg linearly increased (P = 0.092) G:F and decreased (P < 0.05) the fecal score of the pigs challenged with F18+E. coli. Supplementation of ZnGly from 0 to 1,200 mg/kg had quadratic effects on TNF-α (P = 0.065; minimum 1.13 pg/mg at 850 mg/kg ZnGly), IL-8 (P = 0.093; minimum 0.53 ng/mg at 494 mg/kg), and protein carbonyl (P = 0.054; minimum 2.30 pg/mg at 675 mg/kg) and linearly increased mRNA expressions of ZIP4 (P = 0.057) and ZnT5 (P = 0.075) in the jejunum of the pigs. Supplementation of ZnGly from 0 to 1,200 mg/kg linearly increased (P < 0.05) the relative abundance of Actinobacteria and had quadratic effects on Cyanobacteria (minimum 0.67% at 625 mg/kg ZnO) and Proteobacteria (maximum 45.6 g/d at 735 mg/kg) at the phylum level, with linearly decreased (P < 0.05) Enterobacteriaceae at the family level in the jejunal mucosa-associated microbiota of the pigs. There was no difference in growth performance during the overall period, although pigs fed with ZnO at 2,500 mg/kg had greater (P < 0.05) ADG than pigs fed with ZnGly at 400 mg/kg during the first week of the post challenge period. In conclusion, ZnGly could be an alternative to the pharmaceutical use of ZnO without negatively affecting the growth of nursery pigs by enhancing intestinal Zn absorption, reducing intestinal inflammation and oxidative stress, and providing positive changes in jejunal mucosa-associated microbiota.

Enterotoxigenic Escherichia coli (E. coli) strains such as F18+ and F4+E. coli are prominent harmful bacteria that cause impaired intestinal health to nursery pigs with postweaning diarrhea, leading to significant economic losses in the swine industry. In previous years, swine producers used excessively high levels of zinc oxide (ZnO) in feeds for therapeutic purpose as a growth promoter with diarrhea prevention. However, the pharmaceutical use of ZnO is prohibited by the legislation of several countries due to potential health and environmental concerns. Considering the biological functions of zinc (Zn) and the biological efficacy of the organic mineral, zinc glycinate (ZnGly) could effectively enhance the intestinal health of nursery pigs, leading to a decrease in the use of ZnO in feeds. This study aimed to investigate the effects of ZnGly supplementation, allowing the reduced use of ZnO for growth performance and intestinal health of nursery pigs challenged with F18+E. coli. Supplementation of ZnGly at a range of 400 to 675 mg/kg would reduce the negative impacts of F18+E. coli by reducing intestinal inflammation and oxidative stress with positive changes in jejunal microbiota, leading to improved growth performance and reduced diarrhea. In particular, ZnGly supplementation at 400 mg/kg could effectively reduce the use of ZnO in nursery feeds by having similar effects to the pharmaceutical use of ZnO.

Design of a chimeric protein composed of FimH, FyuA and CNF-1 virulence factors from uropathogenic Escherichia coli and evaluation its biological activity and immunogenicity in vitro and in vivo.

Urinary tract infections (UTIs) caused by Uropathogenic Escherichia coli (UPEC) are among the most prevalent bacterial infections in humans. Antibiotic resistance among UPEC isolates is increasing, and designing an effective vaccine can prevent or reduce these infections. FimH adhesin, iron scavenger receptor FyuA, and cytotoxic necrotizing factor -1 (CNF-1) are among the most important virulence factors of UPEC strains. Thus, a novel multi-epitope protein composed of FimH, FyuA, and CNF-1 was designed to evaluate its biological activity and immunogenicity in vitro and in vivo, respectively. The final vaccine design had seven domains, including the N-terminal domain of FimH, four domains of FyuA, and two domains of CNF-1, as determined by immunoinformatics analysis. The results of tertiary structure prediction showed that the chimeric protein had a C-score of -0.25 and Z-score of -1.94. Molecular docking indicated that thirty six ligand residues of the chimeric protein interacted with 53 receptor residues of TLR-4 by hydrogen bonds and hydrophobic interactions. Analysis of protein expression by SDS-PAGE showed an approximately 44 kDa band with different concentrations of IPTG which were confirmed by Western blot. According to ELISA results, the level of IL-8 produced by stimulated Ht29 cells with the chimeric protein was significantly higher than the stimulated Ht29 cells with CNF-1 alone and un-stimulated Ht29 cells. Rabbits subcutaneously immunized with the chimeric protein admixed with Freund adjuvant induced higher level of serum IgG on day 14 after the first vaccination than control rabbits. Furthermore, the booster dose of the chimeric protein significantly enhanced the IgG levels as compared to day 14 and also controls. As, the chimeric protein has suitable B-cell epitopes and MHC-I and MHC-II binding epitopes to stimulate humoral and cellular immunity, it could be a promising vaccine candidate against UTIs caused by UPEC. Evaluating the multi-epitope protein in inducing humoral and cellular immune responses, as well as protection, is ongoing in the mice models.

Synergistic effect of β-thujaplicin and tigecycline against tet(X4)-positive Escherichia coli in vitro / 生物工程学报

The widespread of tigecycline resistance gene tet(X4) has a serious impact on the clinical efficacy of tigecycline. The development of effective antibiotic adjuvants to combat the looming tigecycline resistance is needed. The synergistic activity between the natural compound β-thujaplicin and tigecycline in vitro was determined by the checkerboard broth microdilution assay and time-dependent killing curve. The mechanism underlining the synergistic effect between β-thujaplicin and tigecycline against tet(X4)-positive Escherichia coli was investigated by determining cell membrane permeability, bacterial intracellular reactive oxygen species (ROS) content, iron content, and tigecycline content. β-thujaplicin exhibited potentiation effect on tigecycline against tet(X4)-positive E. coli in vitro, and presented no significant hemolysis and cytotoxicity within the range of antibacterial concentrations. Mechanistic studies demonstrated that β-thujaplicin significantly increased the permeability of bacterial cell membranes, chelated bacterial intracellular iron, disrupted the iron homeostasis and significantly increased intracellular ROS level. The synergistic effect of β-thujaplicin and tigecycline was identified to be related to interfere with bacterial iron metabolism and facilitate bacterial cell membrane permeability. Our studies provided theoretical and practical data for the application of combined β-thujaplicin with tigecycline in the treatment of tet(X4)-positive E. coli infection.

Therapeutic effects of orally administration of viable and inactivated probiotic strains against murine urinary tract infection.

Urinary tract infections (UTIs) are highly prevalent bacterial infections that pose significant health risks. Specific probiotic strains have been recommended for UTI control and management of antibiotic resistance. Otherwise, para-probiotics, defined as inactivated probiotic cells, offer potential advantages by minimizing risks associated with live microorganisms. However, the effectiveness of heat-killed probiotic strains against UTIs remains uncertain. Additionally, lactoferrin (LF), an iron-binding glycoprotein, exhibits immunomodulatory, antimicrobial, and anti-inflammatory properties. Recently, we had developed recombinant LF-expression probiotics, which can display considerate antibacterial activities against select food-borne pathogens in vitro. Thus, the present study aimed to evaluate the antibacterial activities of heat-killed natural and recombinant LF-expressing probiotics against UTIs in vitro and in vivo. Firstly, using in vitro assays, we assessed the antibacterial activity of heat-killed natural and recombinant LF-expressing probiotics against uropathogenic Escherichia coli and Klebsiella pneumoniae. Among the tested probiotics, 10 heat-killed LF-expressing strains displayed superior antibacterial efficacy compared to 12 natural probiotics. Based on their potent in vitro activity, selected probiotics were formulated into three probiotic mixtures: viable probiotic mixture (LAB), heat-killed probiotic mixture (HK-LAB), and heat-killed LF-expressing probiotic mixture (HK-LAB/LF). To further evaluate the therapeutic potential of these probiotic mixtures in vivo, we established a murine model of UTIs by intraurethral administration of E. coli to 40 female C57BL/6JNarl mice on day 0. Subsequently, mice received oral gavage of placebo, LAB, HK-LAB, or HK-LAB/LF for 21 consecutive days (n = 8 per group). An additional control group (n = 8) received ampicillin treatment for 7 days. To assess protective effects against re-infection or UTI relapse, all mice were challenged with E. coli on day 22 and E. coli plus K. pneumoniae on day 25. Results from the murine UTI model demonstrated that placebo administration did not reduce bacteriuria throughout the experiment. Conversely, supplementation with ampicillin, HK-LAB/LF, HK-LAB, or LAB significantly (p < 0.05) reduced daily bacteriuria by 103 to 104-fold on days 1, 3, 5, and 14, respectively. Furthermore, all four therapeutic treatments improved the bacteriological cure rate (BCR) with varying levels of efficacy. For the 7-day treatment course, the BCR was 25% (placebo), 62.5% (ampicillin), 37.5% (LAB), 37.5% (HK-LAB), and 62.5% (HK-LAB/LF). For the 21-day treatment course, the BCR was 25% (placebo), 75% (ampicillin), 37.5% (LAB), 37.5% (HK-LAB), and 75% (HK-LAB/LF). Notably, HK-LAB and HK-LAB/LF demonstrated superior therapeutic efficacy compared to viable LAB in treating UTIs. Overall, regarding BCR, the three probiotic mixtures can provide benefits against UTI in mice, but ampicillin therapy remains the most efficient among the four treatments. Furthermore, there was no significant difference between pre- and post-challenge courses for the two instances of re-challenging uropathogens in all mice groups, as bacteriuria levels remained below 103 CFU/mL, implying that adaptive responses of mice may help reduce the risk of recurrent UTIs. In conclusion, our results provide new evidence that oral administration of heat-killed probiotic mixtures can confer significant therapeutic efficacy against UTIs in a murine model.