Escherichia coli isolated from pyometra and cystitis in the same animal exhibit a wide phenotypic similarity.

AIMS: Pyometra and cystitis caused by Escherichia coli are common diseases identified in canine or feline females. The origin of pyometra infection remains uncertain, and effective prevention strategies for this disease are still unknown. This study aimed to provide a phenotypic characterization, including antimicrobial resistance and virulence profiles, of endometrial pathogenic (EnPEC) and uropathogenic (UPEC) E. coli strains isolated simultaneously from the same animal. METHODS AND RESULTS: Sixteen E. coli strains, from eight different animals, were analyzed in this study. The antimicrobial susceptibility profile of EnPEC and UPEC strains was determined using the disc diffusion method, which showed a similar susceptibility profile among strains (EnPEC and UPEC) from the same animal. The virulence profile of the strains was assessed through biofilm formation, as well as serum resistance abilities. EnPEC and UPEC strains from the same animal exhibited slight variations in their virulence and antimicrobial resistance capabilities. Overall, most of the strain pairs showed a high similarity in their ability to establish biofilms and survive in serum complement activity. CONCLUSIONS: Overall, strains of E. coli isolated from both pyometra and cystitis in the same animal, despite presenting distinct clinical diseases, exhibit a wide phenotypic similarity, suggesting a common origin for the strains.

Redox balance and immunity of piglets pre- and post-E. coli challenge after treatment with hemp or fish oil, and vitamin E.

This study investigated the influence of polyunsaturated fatty acid composition and vitamin E supplementation on oxidative status and immune responses in weanling piglets pre- and post-E. coli challenge. Suckling piglets (n = 24) were randomly selected from two litters for an oral supplementation (1 mL/day) with fish oil or hemp oil and vitamin E supplementation (60 mg natural vitamin E/mL oil) from day 10 to 28 of age. At day 29 and 30 of age, each piglet was orally inoculated with 6.7 × 108 and 3.96 × 108 CFU of F4 and F18 E. coli, respectively. Blood was sampled from all piglets on day 28 before E. coli challenge and on day 35 of age to investigate immunological and oxidative stress markers in plasma. One week after weaning and exposure to E. coli, a general reduction in the α-tocopherol concentration and activity of GPX1 was obtained. Vitamin E supplementation lowered the extent of lipid peroxidation and improved the antioxidative status and immune responses after E. coli challenge. Hemp oil had the greatest effect on antioxidant enzyme activity. Provision of hemp oil and vitamin E to suckling piglets may reduce the incidence of post-weaning diarrhea.

Dietary monoglyceride supplementation to support intestinal integrity and host defenses in health-challenged weanling pigs.

Frequent incidence of postweaning enterotoxigenic Escherichia coli (ETEC) diarrhea in the swine industry contributes to high mortality rates and associated economic losses. In this study, a combination of butyric, caprylic, and capric fatty acid monoglycerides was investigated to promote intestinal integrity and host defenses in weanling pigs infected with ETEC. A total of 160 pigs were allotted to treatment groups based on weight and sex. Throughout the 17-d study, three treatment groups were maintained: sham-inoculated pigs fed a control diet (uninfected control [UC], n = 40), ETEC-inoculated pigs fed the same control diet (infected control [IC], n = 60), and ETEC-inoculated pigs fed the control diet supplemented with monoglycerides included at 0.3% of the diet (infected supplemented [MG], n = 60). After a 7-d acclimation period, pigs were orally inoculated on each of three consecutive days with either 3 mL of a sham-control (saline) or live ETEC culture (3 × 109 colony-forming units/mL). The first day of inoculations was designated as 0 d postinoculation (DPI), and all study outcomes reference this time point. Fecal, tissue, and blood samples were collected from 48 individual pigs (UC, n = 12; IC, n = 18; MG, n = 18) on 5 and 10 DPI for analysis of dry matter (DM), bacterial enumeration, inflammatory markers, and intestinal permeability. ETEC-inoculated pigs in both the IC and MG groups exhibited clear signs of infection including lower (P < 0.05) gain:feed and fecal DM, indicative of excess water in the feces, and elevated (P < 0.05) rectal temperatures, total bacteria, total E. coli, and total F18 ETEC during the peak-infection period (5 DPI). Reduced (P < 0.05) expression of the occludin, tumor necrosis factor α, and vascular endothelial growth factor A genes was observed in both ETEC-inoculated groups at the 5 DPI time point. There were no meaningful differences between treatments for any of the outcomes measured at 10 DPI. Overall, all significant changes were the result of the ETEC infection, not monoglyceride supplementation.

Infection caused by the bacterium known as enterotoxigenic Escherichia coli (ETEC) is a common disruptor of weaned pigs' health, leading to economic losses for the producers. To determine if nutritional supplementation could help protect against these losses, weaned pigs were assigned to one of three treatments: 1) uninfected and fed a standard nursery pig diet, 2) infected with ETEC and fed the same standard diet, or 3) infected with ETEC and fed the standard diet supplemented with a combination of butyric, caprylic, and capric fatty acid monoglycerides. Growth performance was tracked throughout the 17-d study and health outcomes were measured at the peak and resolution of ETEC infection. At the peak-infection time point, pigs that were infected with ETEC had lower fecal moisture content, greater fecal bacterial concentrations, and elevated body temperatures compared with uninfected pigs. Additionally, infection reduced expression of genes related to inflammation, angiogenesis, and the intestinal barrier during the peak-infection period. Overall, all significant changes were the result of the ETEC infection, and there were no meaningful differences observed between the different treatments.

Fatal septicemia in 2 South American camelids with caudal C3-pyloric-duodenal adenocarcinoma.

Gastrointestinal adenocarcinomas are often reported in South American camelids (SAC). We describe here cases of gastroduodenal adenocarcinoma in an adult alpaca (Vicugna pacos) and a llama (Llama glama); both SACs were anorectic and lethargic before death. At autopsy, a prominent and firm caudal C3-pyloric-duodenal junction with stricture and ulceration was present in both animals, as were hemorrhages in various organs and hydrothorax. Microscopically, scattered nests, cords, and tortuous acini of neoplastic epithelial cells were embedded in desmoplastic stroma and invaded the submucosa and muscle layers of the gastroduodenal junction. The mucosa was necrotic, with gram-negative rods in the alpaca and colonies of gram-positive cocci in the llama. No tumor metastases were observed. The neoplastic cells immunolabeled for pancytokeratin. Escherichia coli was isolated from the alpaca and Streptococcus lutetiensis from the llama; septicemia was the cause of death in both animals. Although adenocarcinomas arising from gastric compartments and intestinal segments have been reported in SACs, adenocarcinoma of the caudal C3-pyloric-duodenal junction has not been reported previously in these species, to our knowledge.

Exploration of the immune response of grass carp (Ctenopharyngodon idellus) erythrocytes during bacterial infection.

In teleost blood, red blood cells (RBCs) are the most common type of cell, and they differ from mammalian RBCs in having a nucleus and other organelles. As nucleated cells, teleost RBCs contribute to the immune response against pathogens, but their antibacterial mechanism remains unclear. Here, we utilized RNA-Seq to analyze gene expression patterns of grass carp (Ctenopharyngodon idellus) RBCs (GcRBCs) stimulated by Aeromonas hydrophila, Escherichia coli, and Staphylococcus aureus. Our transcriptomic data showed that bacterial stimulation generated many differentially expressed genes (DEGs). Furthermore, several inflammatory pathways responded to bacterial activation, and the TLR, IL-17, and tumor necrosis factor (TNF) signaling pathways were significantly activated based on Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. Furthermore, the findings of qRT-PCR showed markedly elevated expression of various cytokines, including IL-1ß, IL4, IL6, IL8, IL12, and TNFα, in GcRBCs after incubation with bacteria. Reactive oxygen species (ROS) production in GcRBCs was markedly increased after the cells were stimulated with the three bacteria, and the expression of superoxide dismutase, glutathione peroxidase, and antioxidant enzymes, including catalase, was altered. Flow cytometry analysis showed that the apoptosis rate of GcRBCs was enhanced after stimulation with the three bacteria for different times. In summary, our findings reveal that bacterial stimulation activates the immune response of GcRBCs by regulating ROS release, cytokine expression, and the antioxidant system, leading to apoptosis of GcRBCs.

Genome-wide transcriptional profiling and functional analysis of long noncoding RNAs and mRNAs in chicken macrophages associated with the infection of avian pathogenic E. coli.

BACKGROUND: Avian pathogenic E. coli (APEC) can cause localized or systemic infections, collectively known as avian colibacillosis, resulting in huge economic losses to poultry industry globally per year. In addition, increasing evidence indicates that long non-coding RNAs (lncRNAs) play a critical role in regulating host inflammation in response to bacterial infection. However, the role of lncRNAs in the host response to APEC infection remains unclear. RESULTS: Here, we found 816 differentially expressed (DE) lncRNAs and 1,798 DE mRNAs in APEC infected chicken macrophages by RNAseq. The identified DE lncRNA-mRNAs were involved in Toll like receptor signaling pathway, VEGF signaling pathway, fatty acid metabolism, phosphatidylinositol signaling system, and other types of O-glycan biosynthesis. Furthermore, we found the novel lncRNA TCONS_00007391 as an important immune regulator in APEC infection was able to regulate the inflammatory response by directly targeting CD86. CONCLUSION: These findings provided a better understanding of host response to APEC infection and also offered the potential drug targets for therapy development against APEC infection.

Effect of estradiol after bacterial infection on the Wnt/ß-catenin pathway in bovine endometrium epithelial cells and organoids.

Endometritis is a disease caused by a postpartum bacterial infection with a poor prognosis that primarily affects dairy cows. Three-dimensional organoids have been used as a model for endometritis, because they exhibit a structure comparable to that of the endometrium, demonstrating both expansibility and hormone responsiveness. These characteristics render them an ideal platform for in vitro investigations of endometrial diseases. Estradiol (E2) is an endogenous steroid hormone with demonstrated anti-inflammatory properties, and the objective of this study was to determine the mechanism by which E2 modulates the inflammatory response and the Wnt signal transduction pathway in bovine endometrial epithelial cells and organoids following E. coli infection. We present the techniques for isolating and culturing primary bovine endometrial epithelial cells (BEECs), and producing endometrial organoids. For the experiments, the endometrial epithelial cells and organoids were infected with E. coli for 1 h, followed by incubation with E2 for 12 h. The mRNA and protein expressions of the inflammation-related genes, IL-1ß, IL-6, TLR4, and NF-κB, as well as the Wnt pathway-related genes, Wnt4, ß-catenin, c-Myc, and CyclinD1, were assessed using real-time quantitative-PCR and western blotting, respectively. The CCK8 viable cell counting assay was utilized to determine the optimal concentration of the Wnt inhibitor, IWR-1. The mRNA and protein expression of Wnt pathway-related genes was assessed following IWR-1 treatment, while the expression levels of proliferation-associated genes (Ki67, PCNA) and barrier repair genes (occludin, claudin, and Zo-1) in BEECs and organoids were evaluated after E2 treatment. The results of this study show that mRNA expression of the inflammatory genes, IL-1ß, TLR4, and NF-κB (P < 0.05) decreased in BEECs following E2 treatment compared to the E. coli group. The protein expression of the IL-1ß, IL-6, TLR4 and NF-κB genes was also inhibited (P < 0.05). Similar results were observed in tests on the organoids. Our findings demonstrate that E2 significantly upregulates the expression of Wnt-related genes, including ß-catenin and c-Myc, while concurrently downregulating the expression of GSK3ß (P < 0.05). Next, we treated E. coli-infected BEECs and organoids with the Wnt inhibitor, IWR-1. Compared with E. coli and E. coli + E2, the expression of mRNA and protein from Wnt 4, ß-catenin, and CyclinD1 in E. coli + E2 and E. coli + IWR-1 was down-regulated (P < 0.05). The expression of the proliferation genes, Ki67, PCNA, and the tight junction genes, occludin, claudin1, and Zo-1, in organoids was significantly higher than that in BEECs (P < 0.05). In summary, we found strong potential for E2 mitigation of the E. coli-induced inflammatory response in BEECs and organoids, through activation of the Wnt pathway. In addition, the proliferation and repair capacity of organoids was much higher than that of BEECs.

Genes and pathways revealed by whole transcriptome analysis of milk derived bovine mammary epithelial cells after Escherichia coli challenge.

Mastitis, inflammation of the mammary gland, is the costliest disease in dairy cattle and a major animal welfare concern. Mastitis is usually caused by bacteria, of which staphylococci, streptococci and Escherichia coli are most frequently isolated from bovine mastitis. Bacteria activate the mammary immune system in variable ways, thereby influencing the severity of the disease. Escherichia coli is a common cause of mastitis in cattle causing both subclinical and clinical mastitis. Understanding of the molecular mechanisms that activate and regulate the host response would be central to effective prevention of mastitis and breeding of cows more resistant to mastitis. We used primary bovine mammary epithelial cell cultures extracted noninvasively from bovine milk samples to monitor the cellular responses to Escherichia coli challenge. Differences in gene expression between control and challenged cells were studied by total RNA-sequencing at two time points post-challenge. In total, 150 and 440 (Padj < 0.05) differentially expressed genes were identified at 3 h and 24 h post-challenge, respectively. The differentially expressed genes were mostly upregulated at 3 h (141/150) and 24 h (424/440) post-challenge. Our results are in line with known effects of E. coli infection, with a strong early inflammatory response mediated by pathogen receptor families. Among the most significantly enriched early KEGG pathways were the TNF signalling pathway, the cytokine-cytokine receptor interaction, and the NF-kappa B signalling pathway. At 24 h post-challenge, most significantly enriched were the Influenza A, the NOD-like receptor signalling, and the IL-17 signaling pathway.

Effect of Saccharomyces cerevisiae Postbiotics and Essential Oil on Growth Performance and Intestinal Health of Weanling Pigs During K88 ETEC Infection.

Enterotoxigenic Escherichia coli (ETEC) is one of the major bacterial infections, causing substantial economic losses globally in the swine industry. This study aimed to investigate the impact of low Saccharomyces cerevisiae fermentation postbiotics (SCFP), high SCFP, essential oil (EO), or their combination on the growth performance and health of weanling pigs during ETEC infection. Forty-eight male weanling pigs were randomly allocated to five groups: 1) control group (CON-basal diet, n = 16); 2) low SCFP group (LSC-basal diet + 1.25 g/kg SCFP, n = 8); 3) high SCFP group (HSC-basal diet + 2 g/kg SCFP, n = 8); 4) essential oil group (EO-basal diet + 0.4 g/kg EO, n = 8); 5) the SCFP and EO combination group (SE-basal diet + 1.25 g/kg SCFP + 0.4 g/kg EO, n = 8). On day 15 of the trial, pigs in CON were divided into positive control (PC) and negative control (NC), and all pigs, except in NC, were challenged with ETEC. Under the normal condition, dietary LSC, HSC, EO, and EO all increased average daily gain (ADG) (P < 0.05), and decreased F:G ratio (P < 0.05) accompanied by decreased malondialdehyde (MDA) and increases in catalase (CAT), total superoxide dismutase (T-SOD), total antioxidant capacity (T-AOC) indicating enhanced anti-oxidative capacity, as well as decreased IL-2, IL-8, INF-γ, indicating mitigated systemic inflammation. During ETEC infection, all treatments alleviated ETEC-induced ADG reduction, diarrhea, damages in intestinal permeability and morphology, and down-regulation of tight junctions (Claudin1, ZO-1, and Occludin), while HSC and EO exhibited additional protections. All treatments increased CAT, T-SOD, and T-AOC, and decreased MDA in serum and jejunal mucosa at similar degrees (P < 0.05). Moreover, all treatments alleviated ETEC-induced inflammation as shown by decreased IL-6, TNF-α, INF-γ, and increased IL-4 and IL-10 in serum or jejunal mucosa (P < 0.05), and enhanced the immunity by increased serum IgG and mucosal sIgA (P < 0.05). HSC and SE further reduced mucosal INF-γ and TNF-α than LSC or EO aligning with their additional protection against diarrhea during ETEC infection. Additionally, the key gut bacteria (e.g., Terrisporobacter) related to the benefits of SCFP and EO were identified. In sum, all treatments enhanced growth performance and protected against ETEC-induced intestinal damage through the regulation of redox and immune homeostasis. HSP and SE offered extra protection during disease for their additional control of inflammation. Our study provided new insight into the use of feed additives in the context of animal health states.

Weanling pigs are vulnerable to a variety of stressors and pathogen infections. Enterotoxigenic Escherichia coli (ETEC) is one of the leading causes of diarrhea and growth retardation in weanling pigs. The postbiotics, Saccharomyces cerevisiae fermentation postbiotics (SCFP), and essential oil (EO, mainly thymol, and cinnamaldehyde) were reported to exert health benefits in different sites of the intestine. However, whether SCFP and EO have dose and synergistic effects on weanling pigs, especially against ETEC infection, is incompletely understood. Our research has revealed that SCFP, EO, and their combination all enhanced the growth performance and intestinal barrier function, and reduced diarrhea of piglets, albeit to varying degrees, under both health conditions and ETEC infection. We further elucidated the disparity in the regulation of redox and immune homeostasis by SCFP, EO, and their combination contributing to their different action in distinct states. This has led to a reevaluation of the function of additives in the context of gut health and disease susceptibility.

Oral phages prophylaxis against mixed Escherichia coli O157:H7 and Salmonella Typhimurium infections in weaned piglets.

Escherichia coli and Salmonella Typhimurium are the main pathogens of diarrhea in weaned piglets. The prevention of bacterial diarrhea in weaned piglets by phage is rarely reported. We conducted this study to evaluate the preventive effect of phages on mixed Escherichia coli and Salmonella Typhimurium infections in weaned piglets. A novel phage named NJ12 was isolated by using Salmonella Typhimurium SM022 as host bacteria and characterized by electron microscopy, genomic analysis and in vitro bacteriostatic activity. Phage NJ12 and a previously reported phage EP01 were microencapsulated with sodium alginate to make phage cocktail. Microencapsulated phage cocktail and PBS (Phosphate buffer solution) were used to piglets the phage and phage-free group through oral administration before bacterial infection 2 h, respectively. Piglets of the phage and phage-free group were consumed with feed contaminated with 6 mL (108CFU/mL) Escherichia coli O157:H7 GN07 (GXEC-N07) and 6 mL (108CFU/mL) SM022 every day for seven consecutive days. The results showed that piglets in the phage-free group had more severe diarrhea, larger decreased average weight gain and higher levels of neutrophils compared with piglets in phage group. Meanwhile, piglets in the phage-free group had higher load of SM022 and GN07 in jejunal tissue and more severe intestinal damage compared with piglets in group phage in vivo. In addition, oral administration phage can significant decreased the relative abundance of Enterobacteriaceae but hardly repaired the changes of diversity and composition of gut microbiota caused by the mixed infection of SM022 and GN07. This implies that phage used as a feed additive have a marvelous preventive effect on bacterial diarrhea during weaning of piglets.

Immunomodulatory effect of probiotic exopolysaccharides in a porcine in vitro co-culture model mimicking the intestinal environment on ETEC infection.

The aim of this study was to evaluate the immunomodulatory effect of EPS-L26 isolated from the probiotic strain Lactobacillus (Limosilactobacillus) reuteri L26 Biocenol™, in a model of infection with an enterotoxigenic E. coli (ETEC) by establishing monocultures consisting of the IPEC-J2 cell line or monocyte-derived dendritic cells (moDCs) and creating a 3D model of cell co-cultures established with IPEC-J2 cells and moDCs. The immunomodulatory and immunoprotective potential of used EPS-L26 was confirmed in monocultures in an experimental group of pretreated cells, where our study showed that pretreatment of cells with EPS-L26 and subsequent exposure to infection resulted in significantly down-regulated mRNA levels of genes encoding inflammatory cytokines compared to ETEC challenge in single cell cultures (in IPEC-J2, decreased mRNA levels for TNF-α, IL-6, IL-1ß, IL-12p35; in moDCs, decreased mRNA levels for IL-1ß). Similar to monocultures, we also demonstrated the immunostimulatory potential of the ETEC strain in the co-culture model on directly treated IPEC-J2 cells cultivated on insert chambers (apical compartment) and also on indirectly treated moDCs cultivated in the lower chamber (basolateral compartment), however in the co-culture model the expression of inflammatory cytokines was attenuated at the mRNA level compared to monocultures. Pretreatment of the cells on the insert chambers pointed to the immunoprotective properties of EPS-L26, manifested by decreased mRNA levels in both cell lines compared to ETEC challenge (in IPEC-J2 decreased mRNA levels for IL-12p35; in moDCs decreased mRNA levels for IL-1ß, IL-6). Our results suggest intercellular communication via humoral signals derived from IPEC-J2 cells by influencing the gene expression of indirectly treated moDC cells located in the basolateral compartment.

In situ hybridization to detect Escherichia coli in canine granulomatous colitis.

Canine granulomatous colitis (histiocytic ulcerative colitis) is an uncommon disease, predominantly of young French Bulldogs and Boxer dogs, that manifests from a dysregulated immune response, primarily to adherent-invasive Escherichia coli (AIEC). In conjunction with histopathology and periodic acid-Schiff staining, the diagnosis of granulomatous colitis currently relies on fluorescence in situ hybridization (ISH) or immunohistochemistry to identify and localize AIEC organisms within macrophages in the mucosa and/or submucosa. We investigated the utility of ISH for E. coli using formalin-fixed, paraffin-embedded specimens collected from 29 cases of suspected granulomatous colitis. Most confirmed cases of granulomatous colitis were in French Bulldogs (12 of 20; 60%) and Boxers (3 of 20; 15%), and the mean age was 25 ± 6 mo with no sex predilection. E. coli ISH signal localized bacterial genetic material within the mucosa in 20 of 29 (69%) cases, supporting the diagnosis. ISH signal was limited to the lumen in 8 of 29 (28%) cases, which did not support the identification of these organisms as AIEC. The remaining case had no hybridization signal, and the diagnosis of granulomatous colitis was not supported. Our results revealed that ISH is a quick and specific detection method that can effectively confirm the diagnosis of canine granulomatous colitis.

Andrographolide loaded montmorillonite attenuated enterotoxigenic Escherichia coli induced intestinal barrier injury and inflammation in a mouse model.

Montmorillonite (MMT), a natural absorbent agent, has widely been accepted for its antidiarrhea function in human and farm animals; however, its specific physicochemical property limits its biological function in practical use. In the current study, raw MMT was loaded by andrographolide, namely andrographolide loaded montmorillonite (AGP-MMT). The microstructure of AGP-MMT was observed by scanning electron microscope (SEM) and X-ray diffraction (XRD). The effect of AGP-MMT on the growth performance, intestinal barrier and inflammation was investigated in an enterotoxigenic Escherichia coli (ETEC) challenged mice model. The results show that the microstructure of MMT was obviously changed after andrographolide modification: AGP-MMT exhibited a large number of spheroid particles, and floccule aggregates, but lower interplanar spacing compared with MMT. ETEC infection induced body weight losses and intestinal barrier function injury, as indicated by a lower villus height and ratio of villus height/crypt depth, whereas the serum levels of diamine oxidase (DAO), D-xylose and ETEC shedding were higher in the ETEC group compared with the CON group. Mice pretreated with AGP-MMT showed alleviated body weight losses and the intestinal barrier function injury induced by ETEC challenge. The villus height and the ratio of villus height/crypt depth, were higher in mice pretreated with AGP-MMT than those pretreated with equal levels of MMT. Pretreatment with AGP-MMT also alleviated the increased concentration of serum tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß), and the corresponding genes in the jejunum induced by ETEC infection in mice. The protein and mRNA levels of IL-1ß were lower in mice pretreated with AGP-MMT than those with equal levels of MMT. The results indicate that AGP-MMT was more effective in alleviating intestinal barrier injury and inflammation in mice with ETEC challenge than MMT.

Dihydroartemisinin inhibits plasmid transfer in drug-resistant Escherichia coli via limiting energy supply.

Conjugative transfer of antibiotic resistance genes (ARGs) by plasmids is an important route for ARG dissemination. An increasing number of antibiotic and nonantibiotic compounds have been reported to aid the spread of ARGs, highlighting potential challenges for controlling this type of horizontal transfer. Development of conjugation inhibitors that block or delay the transfer of ARG-bearing plasmids is a promising strategy to control the propagation of antibiotic resistance. Although such inhibitors are rare, they typically exhibit relatively high toxicity and low efficacy in vivo and their mechanisms of action are inadequately understood. Here, we studied the effects of dihydroartemisinin (DHA), an artemisinin derivative used to treat malaria, on conjugation. DHA inhibited the conjugation of the IncI2 and IncX4 plasmids carrying the mobile colistin resistance gene ( mcr-1) by more than 160-fold in vitro in Escherichia coli, and more than two-fold (IncI2 plasmid) in vivo in a mouse model. It also suppressed the transfer of the IncX3 plasmid carrying the carbapenem resistance gene bla NDM-5 by more than two-fold in vitro. Detection of intracellular adenosine triphosphate (ATP) and proton motive force (PMF), in combination with transcriptomic and metabolomic analyses, revealed that DHA impaired the function of the electron transport chain (ETC) by inhibiting the tricarboxylic acid (TCA) cycle pathway, thereby disrupting PMF and limiting the availability of intracellular ATP for plasmid conjugative transfer. Furthermore, expression levels of genes related to conjugation and pilus generation were significantly down-regulated during DHA exposure, indicating that the transfer apparatus for conjugation may be inhibited. Our findings provide new insights into the control of antibiotic resistance and the potential use of DHA.

The therapeutic potential for targeting CSE/H2S signaling in macrophages against Escherichia coli infection.

Macrophages play a pivotal role in the inflammatory response to the zoonotic pathogen E. coli, responsible for causing enteric infections. While considerable research has been conducted to comprehend the pathogenesis of this disease, scant attention devoted to host-derived H2S. Herein, we reported that E. coli infection enhanced the expression of CSE in macrophages, accompanied by a significantly increased inflammatory response. This process may be mediated by the involvement of excessive autophagy. Inhibition of AMPK or autophagy with pharmacological inhibitors could alleviate the inflammation. Additionally, cell model showed that the mRNA expression of classic inflammatory factors (Il-1ß, Il-6), macrophage polarization markers (iNOS, Arg1) and ROS production was significantly down-regulated after employing CSE specific inhibitor PAG. And PAG is capable of inhibiting excessive autophagy through the LKB1-AMPK-ULK1 axis. Interestingly, exogenous H2S could suppress inflammation response. Our study emphasizes the importance of CSE in regulating the macrophage-mediated response to E. coli. Increased CSE in macrophages leads to excessive inflammation, which should be considered a new target for drug development to treat intestinal infection.

Detection of Antibiotic Resistance in Feline-Origin ESBL Escherichia coli from Different Areas of China and the Resistance Elimination of Garlic Oil to Cefquinome on ESBL E. coli.

The development of drug-resistance in the opportunistic pathogen Escherichia coli has become a global public health concern. Due to the share of similar flora between pets and their owners, the detection of pet-origin antibiotic-resistant E. coli is necessary. This study aimed to detect the prevalence of feline-origin ESBL E. coli in China and to explore the resistance elimination effect of garlic oil to cefquinome on ESBL E. coli. Cat fecal samples were collected from animal hospitals. The E. coli isolates were separated and purified by indicator media and polymerase chain reaction (PCR). ESBL genes were detected by PCR and Sanger sequencing. The MICs were determined. The synergistic effect of garlic oil and cefquinome against ESBL E. coli was investigated by checkerboard assays, time-kill and growth curves, drug-resistance curves, PI and NPN staining, and a scanning electronic microscope. A total of 80 E. coli strains were isolated from 101 fecal samples. The rate of ESBL E. coli was 52.5% (42/80). The prevailing ESBL genotypes in China were CTX-M-1, CTX-M-14, and TEM-116. In ESBL E. coli, garlic oil increased the susceptibility to cefquinome with FICIs from 0.2 to 0.7 and enhanced the killing effect of cefquinome with membrane destruction. Resistance to cefquinome decreased with treatment of garlic oil after 15 generations. Our study indicates that ESBL E. coli has been detected in cats kept as pets. The sensitivity of ESBL E. coli to cefquinome was enhanced by garlic oil, indicating that garlic oil may be a potential antibiotic enhancer.

Escherichia coli-associated follicular cystitis in dogs: Clinical and pathologic characterization.

BACKGROUND: Follicular cystitis is an uncommon inflammatory change in the urinary bladder wall characterized by the formation of tertiary lymphoid structures (TLSs) in the submucosa. OBJECTIVES: To characterize clinical and pathologic features of follicular cystitis in dogs and to explore in situ distribution and possible role of Escherichia coli as an associated cause. ANIMALS: Eight dogs diagnosed with follicular cystitis and 2 control dogs. METHODS: Retrospective descriptive study. Dogs diagnosed with follicular cystitis (macroscopic follicular lesions in the urinary bladder mucosa and histopathologic detection of TLSs in bladder wall biopsies) were identified from medical records. Paraffin embedded bladder wall biopsies were subject to in situ hybridization for E. coli 16SrRNA identification. RESULTS: Follicular cystitis was diagnosed in large breed (median weight 24.9 kg, interquartile range [IQR] 18.8-35.4 kg) female dogs with a history of chronic recurrent urinary tract infections (UTIs; median duration of clinical signs 7 months, IQR 3-17 months; median number of previous UTIs 5, IQR 4-6). Positive E. coli 16SrRNA signal was detected within developing, immature and mature TLSs in 7/8 dogs, through submucosal stroma in 8/8 dogs and within the urothelium in 3/8 dogs. CONCLUSIONS AND CLINICAL IMPORTANCE: Chronic inflammation associated with an intramural E. coli infection in the urinary bladder wall represents a possible triggering factor for the development of follicular cystitis.

Effect of different ratios of phytogenic feed additives on growth performance, nutrient digestibility, intestinal barrier integrity, and immune response in weaned pigs challenged with a pathogenic Escherichia coli.

This study was conducted to investigate the effects of supplementing different ratios of phytogenic feed additives (PFA) to weaned pigs challenged with pathogenic Escherichia coli on growth performance, nutrient digestibility, intestinal barrier integrity, and immune response, and to determine the optimal mixing ratio for post-weaning diarrhea (PWD) prevention. A total of 48 4-wk-old weaned pigs with initial body weight of 8.01 ± 0.39 kg were placed in individual metabolic cages, and then randomly assigned to eight treatment groups. The eight treatments were as follows: a basal diet without E. coli challenge (negative control, NC), a basal diet with E. coli challenge (positive control, PC), PC with supplementing 0.1% mixture of 20% bitter citrus extract (BCE), 10% microencapsulated blend of thymol and carvacrol (MEO), and 70% excipient (T1), PC with supplementing 0.1% mixture of 10% MEO, 20% premixture of grape seed and grape marc extract, green tea, and hops (PGE), and 60% excipient (T2), PC with supplementing 0.1% mixture of 10% BCE, 10% MEO, 10% PGE, and 70% excipient (T3), PC with supplementing 0.1% mixture of 20% BCE, 20% MEO, and 60% excipient (T4), PC with supplementing 0.1% mixture of 20% MEO, 20% PGE, and 60% excipient (T5), and PC with supplementing 0.1% mixture of 10% BCE, 20% MEO, 10% PGE, and 60% excipient (T6). The experiments progressed in 16 days, including 5 days before and 11 days after the first E. coli challenge (day 0). In the E. coli challenge treatments, all pigs were orally inoculated by dividing a total of 10 mL of E. coli F 18 for three consecutive days from day 0 postinoculation (PI). Compared with the PC group, the PFA2 and PFA6 groups significantly increased (P < 0.05) feed efficiency and decreased (P < 0.05) diarrhea during the entire period. At day 11 PI, the PFA6 group significantly improved (P < 0.05) gross energy digestibility compared to the PFA1 group. The PFA6 group significantly decreased (P < 0.05) tumor necrosis factor α (TNF-α) and interleukin-6 in serum and increased (P < 0.05) the villus height to crypt depth ratio (VH:CD). The PFA2 significantly decreased (P < 0.05) the relative protein expression of calprotectin in the ileum. In conclusion, improvements in growth performance, diarrhea reduction, and immunity enhancement are demonstrated when 10% BCE, 20% MEO, 10% PGE, and 60% excipient are mixed.

Phytogenic feed additives (PFA) include various herbs and spices, such as essential oils and polyphenols. Flavonoids and polyphenols contained in PFA are generally known to have antioxidant and antibacterial actions and based on this, PFA is considered an alternative to antibiotics in the swine industry. Pathogenic Escherichia coli infection is one of the most important causes of post-weaning diarrhea (PWD) in pigs. PWD causes intestinal damage, which leads to severe diarrhea, reduced growth performance, and mortality in weaned pigs, resulting in significant financial loss to the swine industry. Therefore, this study was conducted to investigate the effects of supplementing different ratios of PFA to weaned pigs challenged with E. coli and determine the optimal mixing ratio for PWD prevention. Our study results showed that growth performance was improved when supplementing a mixture of 10% bitter citrus extract (BCE), 20% microencapsulated blend of thymol and carvacrol (MEO), 10% premixture of grape seed and grape marc extract, green tea, and hops (PGE), and 60% excipient. Also, the effect of improving the immune response and intestinal morphology was shown. In conclusion, a mixture of 10% BCE, 20% MEO, 10% PGE, and 60% excipients is considered the optimal mixing ratio.

Analysis of alternative splicing in chicken macrophages infected with avian pathogenic E. coli (APEC).

Colibacillosis is a complex disease that caused by avian pathogenic Escherichia coli (APEC), resulting in huge economic loss to the global poultry industry and threatening to human health. Alternative splicing (AS) is a universal post-transcriptional regulatory mechanism, which can simultaneously produce many proteins from a single gene to involve in various diseases and individual development. Herein, we characterized genome-wide AS events in wild type macrophages (WT) and APEC infected macrophages (APEC) by high-throughput RNA sequencing technology. A total of 751 differentially expressed (DE) AS genes were identified in the comparison of APEC vs. WT, including 587 of SE, 114 of MXE, 25 of RI, 17 of A3 and 8 of A5 event. Functional analysis showed that these identified DE AS genes were involved in 'Endocytosis', 'p53 signaling pathway', 'MAPK signaling pathway', 'NOD-like receptor signaling pathway', 'Ubiquitin mediated proteolysis' and 'Focal adhesion' immune related pathways. In summary, we comprehensively investigate AS events during APEC infection. This study has expanded our understanding of the process of APEC infection and provided new insights for further treatment options for APEC infection.

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.