Coli K1 Research Articles

Dietary supplementary with ellagic acid improves the intestinal barrier function and flora structure of broiler chicken challenged with E. coli K88

Ellagic acid (EA) contributes to the immunity and anti-oxidant function of body, whereas there are few reports about its effect on the intestinal health and growth performance of broiler chickens. Hence, the present study was arranged to investigate the effect of dietary supplementary with EA on the intestinal barrier function and flora structure of broiler chickens challenged with Escherichia coli K88 (E. coli K88). A total of 216 healthy 1-day-old, Ross 308 broilers with uniform weight were randomly assigned into three treatment groups, six replicates in each group and twelve birds in each replicate. Broilers in the control (CTR) group and E. coli K88 infected group (ETEC) were fed with the basic diet, and 200 mg/kg EA was supplemented into the diet of the E. coli K88 infected group treated with EA (EAETEC). The animal trial had lasted for 42 days, and the outcomes showed that the ADG and ADFI during the animal trial, the jejunal villi height (VH) and the ratio of VH to crypt depth (CD) tended to be decreased with E. coli K88 treated (P< 0.05). Additionally, the level of serum diamine oxidase (DAO) and intestinal malondialdehyde (MDA) were elevated, the activity of intestinal total superoxide dismutase (T-SOD) and glutathione peroxidase (GSH-Px), the mRNA levels in jejunal claudin-1 and occludin were down-regulated with E. coli K88 treated as well as the transcription levels of ileal Mucin-2, aquaporin-3 (AQP-3) and Na+/H+ exchanger proteins-3 (NHE-3) (P< 0.05). In addition, E. coli K88 down-regulated the α-diversity index of cecal flora, the ratio of Bacteroidota to Firmicutes and the relative abundance of Barnesiella were up-regulated and it of Alistipes was descended with E. coli K88 treated (P< 0.05). Beyond that, the content of propionic acid in the cecal chyme was decreased and the amino acid metabolic pathways were inhibited with E. coli K88 challenged (P< 0.05). Additionally, there was a significant positive correlation between the relative abundance of Alistipes and the levels of butyric acid in the caecal chyme and the activity of GSH-Px in the intestine (P< 0.05). Interestingly, dietary supplementary with EA could reshape the intestinal flora structure and alleviate the above negative effects of E. coli K88 on broiler chickens. In conclusion, dietary supplementary with ellagic acid improved the intestinal barrier function and flora structure of broiler chickens challenged with E. coli K88.

Changes in the Distribution of Pathogens Causing Meningoencephalitis Before and During the COVID-19 Pandemic.

Infectious meningoencephalitis (ME) is a major global health concern. Viruses are the most frequently implicated etiologies, whereas bacterial causes, although life-threatening, constitute a lesser proportion of ME cases, together with other pathogens. The strict implementation of COVID-19 mitigation measures led to the decreased viral and non-viral infectious diseases. Therefore, this study aimed to investigate the effect of these mea-sures on ME-causing pathogens by age groups. This retrospective study aimed to determine and compare the rates of pathogen-positive ME cases during the pre-pandemic (P-1) and pandemic (P-2) periods. Molecular diagnostic methods using the cerebrospinal fluid of patients from all age groups were included. The positivity rate difference of the ME-causing pathogens between the two study periods was compared and the distribution pattern of the pathogens among the age groups was determined. The overall positivity rate for at least one ME-causing pathogen during P-1 was 22.0% (503/2,284), which significantly declined to 7.3% (83/1,141) during P-2 (p < 0.001). Particularly, a statistically significant decline in the pathogen positivity was observed in the groups 4 - 6 (≥ 3 years) (p < 0.001, p < 0.001, and p = 0.041, respectively). Specifically, the enterovirus cases decreased significantly, whereas the varicella zoster virus and herpes simplex virus-2 cases increased. Among bacterial causes, the S. agalactiae, S. pneumoniae, and E. coli K1 ME cases significantly increased. Men and women had no significant differences in the positivity rate during either study period. COVID-19 mitigation measures significantly impacted the positivity rates and the distribution of ME-causing agents, especially in the age groups ≥ 3 years, although not uniformly.

Clostridium butyricum Prevents Diarrhea Incidence in Weaned Piglets Induced by Escherichia coli K88 through Rectal Bacteria-Host Metabolic Cross-Talk.

This study aimed to evaluate the effects of Clostridium butyricum (C. butyricum) on the prevention of the diarrhea rates and growth performances of weaned piglets induced by Escherichia coli K88 (E. coli K88). Twenty-four weaned piglets (6.92 ± 0.11 kg) were randomly assigned to one of three treatment groups for a period of 21 days. Each group consisted of eight pigs, with each pig being housed in an individual pen. Group I received the control diet along with normal saline, Group II received the control diet along with E. coli K88, and Group III received the control diet supplemented with 5 × 108 CFU/kg of C. butyricum and E. coli K88. We examined alterations in rectal microbiota and metabolites, analyzed the incidence of diarrhea, and investigated the interactions between microbiota and metabolites through the application of Illumina MiSeq sequencing and liquid chromatography-mass spectrometry. The results showed that, from days 14 to 21, the diarrhea incidence in Group III decreased significantly by 83.29% compared to Group II (p < 0.05). Over the entire experimental duration, the average daily feed intake of Group III decreased significantly by 11.13% compared to Group I (p < 0.05), while the diarrhea incidence in Group III decreased by 71.46% compared to Group II (p < 0.05). The predominant microbial flora in the rectum consisted of Firmicutes (57.32%), Bacteroidetes (41.03%), and Proteobacteria (0.66%). Administering E. coli K88 orally can elevate the relative abundance of Megasphaera (p < 0.05). Conversely, the supplementation of C. butyricum in the diet reduced the relative abundance of Megasphaera (p < 0.05), while increasing the relative abundance of unclassified_f_Lachnospiraceae (p < 0.05). Rectal metabolomics analysis revealed that supplementing C. butyricum in the feed significantly altered the amino acids and fatty acids of the piglets infected with E. coli K88 (p < 0.05). The correlation analysis showed that the occurrence of diarrhea was inversely related to adipic acid (p < 0.05) and positively associated with (5-hydroxyindol-3-YL) acetic acid and L-aspartic acid (p < 0.05). Prevotella_1 exhibited a negative correlation with octadecanoic acid (p < 0.05). Prevotellaceae_UCG-005 showed a negative correlation with (5-hydroxyindol-3-YL) acetic acid (p < 0.05). The findings from this research study aid in probiotic development and the enhancement of healthy growth in weaned piglets.

A live attenuated vaccine to prevent severe neonatal Escherichia coli K1 infections

Preterm birth is currently the leading cause of neonatal morbidity and mortality. Genetic, immunological and infectious causes are suspected. Preterm infants have a higher risk of severe bacterial neonatal infections, most of which are caused by Escherichia coli an in particular E. coli K1strains. Women with history of preterm delivery have a high risk of recurrence and therefore constitute a target population for the development of vaccine against E. coli neonatal infections. Here, we characterize the immunological, microbiological and protective properties of a live attenuated vaccine candidate in adult female mice and their pups against after a challenge by K1 and non-K1 strains of E. coli. Our results show that the E. coli K1 E11 ∆aroA vaccine induces strong immunity, driven by polyclonal bactericidal antibodies. In our model of meningitis, mothers immunized prior to mating transfer maternal antibodies to pups, which protect newborn mice against various K1 and non-K1 strains of E. coli. Given the very high mortality rate and the neurological sequalae associated with neonatal E. coli K1 meningitis, our results constitute preclinical proof of concept for the development of a live attenuated vaccine against severe E. coli infections in women at risk of preterm delivery.

CsiR-Mediated Signal Transduction Pathway in Response to Low Iron Conditions Promotes Escherichia coli K1 Invasion and Penetration of the Blood-Brain Barrier.

Escherichia coli K1 is the leading cause of neonatal gram-negative bacterial meningitis, but the pathogenesis of E coli K1 meningitis remains unclear. Blood-brain barrier (BBB) penetration is a crucial step in E coli meningitis development. Here, we uncovered the crucial role of CsiR, a GntR family regulator, in E coli K1 virulence. During infection, csiR expression was induced due to the derepression by Fur in the blood and human brain microvascular endothelial cells (HBMECs). CsiR positively regulated ilvB expression, which is associated with branched chain amino acid synthesis. Furthermore, we revealed that IlvB activated the FAK/PI3K pathway of HBMECs to induce actin cytoskeleton rearrangements, thereby promoting the bacterial invasion and penetration of the BBB. Overall, this study reveals a CsiR-mediated virulence regulation pathway in E coli K1, which may provide a useful target for the prevention or therapy of E coli meningitis.

Antibacterial effects of quercetagetin are significantly enhanced upon conjugation with chitosan engineered copper oxide nanoparticles.

The development of antibiotic alternatives that entail distinctive chemistry and modes of action is necessary due to the threat posed by drug resistance. Nanotechnology has gained increasing attention in recent years, as a vehicle to enhance the efficacy of existing antimicrobials. In this study, Chitosan copper oxide nanoparticles (CHI-CuO) were synthesized and were further loaded with Quercetagetin (QTG) to achieve the desired (CHI-CuO-QTG). Size distribution, zeta potential and morphological analysis were accomplished. Next, the developed CHI-CuO-QTG was assessed for synergistic antibacterial properties, as well as cytotoxic attributes. Bactericidal assays revealed that CHI-CuO conjugation showed remarkable effects and enhanced QTG effects against a range of Gram + ve and Gram-ve bacteria. The MIC50 of QTG against S. pyogenes was 107µg/mL while CHI-CuO-QTG reduced it to 9µg/mL. Similar results were observed when tested against S. pneumoniae. Likewise, the MIC50 of QTG against S. enterica was 38µg/mL while CHI-CuO-QTG reduced it to 7µg/mL. For E. coli K1, the MIC50 of QTG was 42µg/mL while with CHI-CuO-QTG it was 23µg/mL. Finally, the MIC50 of QTG against S. marcescens was 98µg/mL while CHI-CuO-QTG reduced it to 10µg/mL. Notably, the CHI-CuO-QTG nano-formulation showed limited damage when tested against human cells using lactate dehydrogenase release assays. Importantly, bacterial-mediated human cell damage was reduced by prior treatment of bacteria using drug nano-formulations. These findings are remarkable and clearly demonstrate that drug-nanoparticle formulations using nanotechnology is an important avenue in developing potential therapeutic interventions against microbial infections.

Pseudomonas fluorescens and Escherichia coli in Fresh Mozzarella Cheese: Effect of Cellobiose Oxidase on Microbiological Stability during Refrigerated Shelf Life.

Background: Mozzarella cheese possesses a high moisture content (50−60%) and a relatively high pH (around 5.5) and is therefore considered a perishable food product characterized by high quality deterioration and the potential risk of microbial contamination. Moreover, it can be spoiled by Pseudomonas spp. and coliform bacteria, which may be involved in different negative phenomena, such as proteolysis, discolorations, pigmentation, and off-flavors. To prevent these, different methods were investigated. In this context, the present study aims to assess the antimicrobial effect of cellobiose oxidase on Pseudomonas fluorescens (5026) and Escherichia coli (k88, k99) in mozzarella cheese during refrigerated shelf life. Methods: microbiological challenge tests were designed by contaminating the mozzarella covering liquid containing different cellobiose oxidase concentrations with P. fluorescens (5026) and E. coli (k88, k99). The behavior of these microorganisms and the variation of hydrogen peroxide concentrations were then tested under refrigerated conditions for 20 days to simulate the mozzarella cheese shelf life. Results and Conclusions: The data obtained demonstrated the effect of cellobiose oxidase on microbial growth. In particular, E. coli (k88, k99) was inhibited over the entire shelf life, while P. fluorescens (5026) was only partially affected after a few days of refrigerated storage.

Effects of glucose oxidase on growth performance, clinical symptoms, serum parameters, and intestinal health in piglets challenged by enterotoxigenic Escherichia coli.

Glucose oxidase (GOD) could benefit intestinal health and growth performance in animals. However, it is unknown whether GOD can protect piglets against bacterial challenge. This study aimed to evaluate the protective effects of GOD on growth performance, clinical symptoms, serum parameters, and intestinal health in piglets challenged by enterotoxigenic Escherichia coli (ETEC). A total of 44 male weaned piglets around 38 days old were divided into four groups (11 replicates/group): negative control (NC), positive control (PC), CS group (PC piglets +40 g/t colistin sulfate), and GOD group (PC piglets +200 g/t GOD). All piglets except those in NC were challenged with ETEC (E. coli K88) on the 11th day of the experiment. Parameter analysis was performed on the 21st day of the experiment. The results showed that the ETEC challenge elevated (p < 0.05) the rectal temperature and fecal score of piglets at certain time-points post-challenge, reduced (p < 0.05) serum glucose and IgG levels but increased (p < 0.05) serum alanine aminotransferase activity, as well as caused (p < 0.05) intestinal morphology impairment and inflammation. Supplemental GOD could replace CS to reverse (p < 0.05) the above changes and tended to increase (p = 0.099) average daily gain during the ETEC challenge. Besides, GOD addition reversed ETEC-induced losses (p < 0.05) in several beneficial bacteria (e.g., Lactobacillus salivarius) along with increases (p < 0.05) in certain harmful bacteria (e.g., Enterobacteriaceae and Escherichia/Shigella). Functional prediction of gut microbiota revealed that ETEC-induced upregulations (p < 0.05) of certain pathogenicity-related pathways (e.g., bacterial invasion of epithelial cells and shigellosis) were blocked by GOD addition, which also normalized the observed downregulations (p < 0.05) of bacterial pathways related to the metabolism of sugars, functional amino acids, nucleobases, and bile acids in challenged piglets. Collectively, GOD could be used as a potential antibiotic alternative to improve growth and serum parameters, as well as attenuate clinical symptoms and intestinal disruption in ETEC-challenged piglets, which could be associated with its ability to mitigate gut microbiota dysbiosis. Our findings provided evidence for the usage of GOD as an approach to restrict ETEC infection in pigs.

P117. Development of an in vitro model to study intestinal integrity during an E. coli K88 challenge

P117. Development of an in vitro model to study intestinal integrity during an E. coli K88 challenge

The immunomodulatory function of the porcine β-defensin 129: Alleviate inflammatory response induced by LPS in IPEC-J2 cells

β-defensin family plays a critical role in host defense against infections. In this study, we found that pBD129 are widely expressed in porcine tissues such as the intestine, liver, and spleen. Interestingly, the expression level of pBD129 in most tissues was higher in Tibetan pigs than in DLY (Duroc × Landrace × Yorkshire) pigs (P < 0.05), and was significantly upregulated upon E. coli K88 infection (P < 0.05). The pBD129 protein was successfully expressed in E. coli and the molecule weight was estimated by SDS-PAGE to be 37.2 kDa. Mass spectrometry verified the protein as a pBD129. The protein showed antibacterial activities against Streptococcus and E. coli DH5α with a minimal inhibitory concentration (MIC) of 32 μg/mL. Hemolytic and cytotoxicity assays indicated that pBD129 had no detrimental effect on cell viability. Importantly, pBD129 significantly reduced the apoptosis of porcine intestinal epithelial cells exposure to bacterial endotoxins, which was associated with down-regulation of inflammatory cytokines such as the IL-1β, IL-6 and TNFα (P < 0.05), and down-regulation of apoptosis-related genes such as the caspase-3, caspase-8, and caspase-9 (P < 0.05). These results suggested that pBD129 is a novel modulator of innate immunity involved in mammalian inflammatory responses.

Effects of dietary lysozyme supplementation on growth performance, nutrient digestibility, intestinal microbiota, and blood profiles of weanling pigs challenged with Escherichia coli.

The aim of this was evaluate the efficacy of lysozyme on growth performance, nutrient digestibility, excreta microflora population, and blood profiles of weanling pigs under Escherichia coli (E. coli) challenge. A total of 30 piglets weaned at 25 days, 7.46 kg body weight, were assigned to three dietary treatments, composed of five replications, two piglets per replication, for 7 days. The dietary treatment groups were negative control (NC; without antibiotics and lysozyme), positive control (PC; NC + antibiotics), lysozyme (NC + 0.1% lysozyme). All piglets were challenged orally with 6 ml suspension, containing E. coli K88 (2 × 109 CFU/mL). Dietary supplementation with lysozyme and PC resulted in no significant differences in average daily gain and gain to feed efficiency. Weanling pigs fed with E. coli challenge with lysozyme and PC treatments had significantly enhanced nutrient retentions of dry matter and energy (p < 0.05); however, there was a tendency to increase nitrogen digestibility. Furthermore, dietary inclusion of lysozyme and antibiotics treatment groups had a beneficial effect on excreta, ileal, and cecal of the fecal microbial population as decreased E. coli (p < 0.05) counts, without effects on lactobacillus counts. A significant effect were observed on a white blood cells, epinephrine and cortisol concentrations were reduced in piglets fed diets containing E. coli challenge with lysozyme and antibiotics supplementation comparison with the NC group. Therefore, the present data indicate that lysozyme in diet could ameliorate the experimental stress response induced by E. coli in piglets by decreasing intestinal E. coli, white blood cells and stress hormones and improving nutrient digestibility.

Effect of Replacing in-Feed Antibiotic Growth Promoters with a Combination of Egg Immunoglobulins and Phytomolecules on the Performance, Serum Immunity, and Intestinal Health of Weaned Pigs Challenged with Escherichia coli K88.

Simple SummaryPost-weaning diarrhea (PWD) in pigs caused by Escherichia coli (E. coli) is a global problem which results in substantial economic losses, due to decreased performance and a high incidence of mortality and morbidity. Due to the banning of antibiotic growth promoters (AGPs) by many countries, it would be valuable to find environmentally friendly and non-antibiotic alternatives to AGPs and to evaluate their effectiveness. Both immunoglobulins and phytomolecules are separately reported as benefiting animal growth, but the efficiency of combinations of immunoglobulins and phytomolecules as AGP alternatives is largely unknown. In this study, the results showed that a mixture of immunoglobulin and phytomolecule administration had positive effects on feed efficiency, diarrhea reduction, intestinal morphology, and coliform control. Combinations of immunoglobulins and phytomolecules can be used as a potential alternative to AGPs in weanling piglets.The study was conducted to investigate the effects of replacing antibiotic growth promoters (AGPs) with an egg immunoglobulin (IgY) combined with phytomolecules (PM) on the growth rate, serum immunity, and intestinal health of weaned pigs challenged with Escherichia coli K88 (E. coli K88). A total of 192 piglets were weaned at 28 days old with an average weight of 7.29 (± 0.04) kg. They were randomly divided into four treatments containing eight replicates with six piglets per replicate. The treatment groups were NC and PC fed a basal diet, AGP fed a basal diet supplemented with 75 mg/kg chlortetracycline, 50 mg/kg oxytetracycline calcium, and 40 mg/kg zinc bacitracin, IPM fed a basal diet supplemented with IgY at dose of 2.5 g/kg and 1.0 g/kg and PM at dose of 300 mg/kg and 150 mg/kg during days 1 to 17 and 18 to 42, respectively. On days 7 to 9 of the experiment, piglets in the PC, AGP, and IPM groups were orally challenged with 20 mL E. coli K88 (109 CFU/mL), while piglets in the NC group were challenged with 20 mL medium without E. coli K88. The E. coli K88 challenge model was successful as the incidence of post-weaning diarrhea (PWD) of piglets challenged with E. coli K88 was significantly higher than that of those unchallenged piglets during the challenge time (days 7 to 9) and days 1 to 7 of post-challenge (p < 0.05). A diet with combinations of IgY and PM and AGPs significantly decreased the incidence of PWD during the challenge time and days 1 to 7 of post-challenge (p < 0.05) compared to the PC group and significantly improved the ratio of feed to weight gain (F:G) during days 1 to 17 of the experiment compared to the NC and PC groups (p < 0.05). In comparison with the PC group, piglets in the IPM group had significantly higher serum levels of IgA, IgG, and IgM (p < 0.05), but lower serum IL-1β on day 17 of experiement (p < 0.05). Besides, diet supplementation with AGP significantly decreased serum IL-1β, IL-6, and TNF-α on days 17 and 42 (p < 0.05) with comparison to the PC group. Piglets in the IPM group showed a significantly lower level of fecal coliforms (p < 0.05), but a higher villus height of jejunum and ileum and higher ratio of villus height to crypt depth of duodenum and jejunum (p < 0.05) than those piglets in the PC group. In summary, diet supplementation with a mixture of IgY and PM decreased the incidence of PWD and coliforms, increased feed conversion ratio, and improved intestinal histology and immune function.

Development of a chitosan\u2010modified PLGA nanoparticle vaccine for protection against Escherichia coli K1 caused meningitis in mice

BackgroundEscherichia coli K1 (E. coli K1) caused neonatal meningitis remains a problem, which rises the urgent need for an effective vaccine. Previously, we rationally designed and produced the recombinant protein OmpAVac (Vo), which elicited protective immunity against E. coli K1 infection. However, Vo has limited stability, which hinders its future industrial application.MethodChitosan-modified poly (lactic-co-glycolic acid) (PLGA) nanoparticles were prepared and used as carried for the recombinant Vo. And the safety, stability and immunogenicity of Vo delivered by chitosan-modified PLGA nanoparticles were tested in vitro and in a mouse model of bacteremia.ResultsWe successfully generated chitosan-modified PLGA nanoparticles for the delivery of recombinant Vo (VoNP). In addition, we found that a freeze-drying procedure increases the stability of the VoNPs without changing the shape, size distribution and encapsulation of the Vo protein. Unlike aluminum adjuvant, the nanoparticles that delivered Vo were immunoprotective in mice even after storage for as long as 180 days.ConclusionsWe identified an effective strategy to improve the stability of Vo to maintain its immunogenicity, which will contribute to the future development of vaccines against E. coli K1.

Transcriptome analysis of Escherichia coli K1 after therapy with hesperidin conjugated with silver nanoparticles

BackgroundsEscherichia coli K1 causes neonatal meningitis. Transcriptome studies are indispensable to comprehend the pathology and biology of these bacteria. Recently, we showed that nanoparticles loaded with Hesperidin are potential novel antibacterial agents against E. coli K1. Here, bacteria were treated with and without Hesperidin conjugated with silver nanoparticles, and silver alone, and 50% minimum inhibitory concentration was determined. Differential gene expression analysis using RNA-seq, was performed using Degust software and a set of genes involved in cell stress response and metabolism were selected for the study.Results50% minimum inhibitory concentration with silver-conjugated Hesperidin was achieved with 0.5 μg/ml of Hesperidin conjugated with silver nanoparticles at 1 h. Differential genetic analysis revealed the expression of 122 genes (≥ 2-log FC, P< 0.01) in both E. coli K1 treated with Hesperidin conjugated silver nanoparticles and E. coli K1 treated with silver alone, compared to untreated E. coli K1. Of note, the expression levels of cation efflux genes (cusA and copA) and translocation of ions, across the membrane genes (rsxB) were found to increase 2.6, 3.1, and 3.3- log FC, respectively. Significant regulation was observed for metabolic genes and several genes involved in the coordination of flagella.ConclusionsThe antibacterial mechanism of nanoparticles maybe due to disruption of the cell membrane, oxidative stress, and metabolism in E. coli K1. Further studies will lead to a better understanding of the genetic mechanisms underlying treatment with nanoparticles and identification of much needed novel antimicrobial drug candidates.

Incidence and Distribution of the Pathogens Causing Central Nervous System Infections at the University Hospital of Korea.

The pathogens involved in central nervous system (CNS) infections are various, such as viruses, bacteria, and fungi, so a syndromic approach can be required. In addition, since their rapid and accurate detection is very crucial, molecular diagnostics using cerebrospinal fluid is becoming the emerging standard method. The study was conducted retrospectively to identify the incidence and distribution patterns of the pathogens according to gender, age, season, and month and to analyze their codetection from August 2017 to July 2020. It was also conducted to investigate turn-around times (TATs) according to the detection method. The detection methods were FilmArray® Meningitis/Encephalitis (M/E) method (FilmArray), Cepheid® Xpert EV assay (Xpert), and Multiplex PCR method for five species of bacteria. The overall incidence for at least one pathogen was 13.9% (346/2,496). The highest incidence was shown in age group 4 (3 - 6 years), with 27.4%. The detection rates by FilmArray, Xpert, and Multiplex PCR method were 39.8%, 41.7%, and 0.4%, respectively. Enterovirus (EV) showed the highest incidence rate, which accounted for 37.0%. The distribution of the pathogens according to the age groups were the highest in age group 4, with 47.5% (168/354), followed by 27.4% (97/354) in age group 5. Of the ten cases in which bacteria were detected, S. agalactiae accounted for 60.0% (6/10), most of which occurred in age group 1. E. coli K1, L. monocytogenes, and N. meningitidis were not detected. In the viral distribution, EV accounted for the highest proportion in all age groups. The overall proportion of EV accounted for 87.6% (310/354), followed by human parechovirus with 2.8% (10/354). The most commonly detected season was summer, comprising 75.1%. A total of eight cases of co-detection with two pathogens accounted for 1.6% (8/507) in FilmArray. In FilmArray, all TATs were found to be shorter than Xpert. The information on the incidence and distribution patterns of the pathogens causing CNS infections and their rapid detection are critically important to clinicians in the management of immunocompromised patients, elderly, and children. The expeditious molecular diagnostics for these pathogens would be valuable in medical decisions by clinicians.

Escherichia coli K1 ibeA-mediated virulence enhances biofilm-associated adhesion to human brain microvascular endothelial cells

Background: Previous studies have confirmed that ibeA is a key virulence gene of E.coli K1 which plays an important role in the pathogenesis of E.coli K1. The adhesion of E.coli K1 to human brain microvascular endothelial cells is one of the key issues in the pathogenesis of bacterial neonatal meningitis. Biofilm is associated with bacterial adhesion. In this study, the biofilm formation was observed in the IbeA gene deletion mutant, confirming that IbeA is involved in biofilm formation. Methods and materials: The bacteria were divided into four groups: (1) DH5α, (2) E. coli K1, the IbeA knockout strain and the IbeA complemented strain. The biofilm formation was detected by crystal violet staining. The adhesion was measured specifically due to IbeA by comparing the adhesive properties of the IbeA complemented strain to those of the IbeA knockout mutant strain. Results: The formation of biofilm in the wildtype E. coli K1 group was significantly higher than that in the knockout group and in the negative group (p < 0.05). The biofilm formation of the IbeA complemented group was significantly higher than that in IbeA knockout group (p < 0.05). Besides, the adhesion of E. coli K1 was significantly higher than that in the knockout group and the negative group (p < 0.05). The adhesion rate of the IbeA complemented group was significantly higher than that of the IbeA knockout group (p < 0.05). Conclusion: The biofilm formation of E. coli K1 strain is induced by ibeA which is a key virulence gene in the pathogenesis, and ibeA-mediated virulence enhances biofilm-associated adhesion to human brain microvascular endothelial cells in vitro. (Acknowledgements: *Corresponding author: Hong Cao, The study was supported by Undergraduate Training Program for Innovation and Entrepreneurship of SMU, China, No. 201912121275 and Scientific Research Enlightenment Plan of SMU, China,2019)

An ArcA-Modulated Small RNA in Pathogenic Escherichia coli K1

Escherichia coli K1 is the leading cause of meningitis in newborns. Understanding the molecular basis of E. coli K1 pathogenicity will help develop treatment of meningitis and prevent neurological sequelae. E. coli K1 replicates in host blood and forms a high level of bacteremia to cause meningitis in human. However, the mechanisms that E. coli K1 employs to sense niche signals for survival in host blood are poorly understood. We identified one intergenic region in E. coli K1 genome that encodes a novel small RNA, sRNA-17. The expression of sRNA-17 was downregulated by ArcA in microaerophilic blood. The ΔsRNA-17 strain grew better in blood than did the wild-type strain and enhanced invasion frequency in human brain microvascular endothelial cells. Transcriptome analyses revealed that sRNA-17 regulates tens of differentially expressed genes. These data indicate that ArcA downregulates the sRNA-17 expression to benefit bacterial survival in blood and penetration of the blood–brain barrier. Our findings reveal a signaling mechanism in E. coli K1 for host adaptation.

Diagnostic Yield of Multiplex PCR Method in Cerebrospinal Fluid for the Diagnosis of Purulent Meningitis in Children

Abstract Objective To evaluate the diagnostic yield of the multiplex polymerase chain reaction (PCR) method in cerebrospinal fluid (CSF) for the diagnosis of purulent meningitis (PM) in children. Methods PM was diagnosed according to the European Society for Clinical Microbiology and Infectious Diseases guideline (2016). Patients with PM between May 2015 and October 2018 were included. The multiplex PCR method was used to detect eight common identified bacteria in PM. Its sensitivity and specificity were compared with bacteria culture. Results A total of 106 cases were enrolled. Pathogenic bacteria were identified in 27 (25.5%) cases by culture and in 37 (34.9%) cases by multiplex PCR assay. The top three bacteria were Streptococcus pneumoniae, Escherichia coli K1, and Streptococcus agalactiae. When using culture as the gold standard, the multiplex PCR assay showed a sensitivity of 100, 88.9, and 75.0% for S. agalactiae, S. pneumoniae, and E. coli K1, respectively, and a specificity of more than 91.3% for all three bacteria. For detectable bacteria, the positive rate of the multiplex PCR assay (36.6%, 37/101) was significantly higher than that of the bacteria culture (21.8%, 22/101). When combining the two methods, etiology was identified in 42.5% (45/106) of the patients. Conclusion Streptococcus pneumoniae, E. coli K1, and S. agalactiae were the predominant pathogens causing pediatric PM. As a rapid method with high sensitivity and specificity, the multiplex PCR assay in CSF could be used as an adjunctive approach with bacteria culture for the pathogen identification of PM.

Lactobacillus plantarum BSGP201683 Improves the Intestinal Barrier of Giant Panda Microbiota-Associated Mouse Infected by Enterotoxigenic Escherichia coli K88.

Giant pandas often suffered from gastrointestinal disease, especially the captive sub-adult one. Our study aims to investigate whether L. plantarum G83, a good panda-derived probiotic, can improve the intestinal barrier against the enterotoxigenic Escherichia coli K88 (E. coli K88) infection in giant panda microbiota-associated mice (GPAM). We treated SPF mice with antibiotics cocktail and transplanted the giant panda intestinal microbiota to set up a GPAM. Our results demonstrated that the microbiota of GPAM changed over time and was relatively stable in the short-term experiment (2-4weeks). Whereafter, the GPAM pretreated with L. plantarum G83 for 15days and infected with enterotoxigenic E. coli K88. The result indicated that the number of Bifidobacteria spp. increased in GPAM-G and GPAM-GE groups; the Lactobacillus spp. only increased in the GPAM-G group. Although the abundance of Enterobacteriaceae spp. only decreased in the GPAM-G group, the copy number of Escherichia coli in the GPAM-E group was significantly lower than that in the other groups. Meanwhile, the L. plantarum G83-induced alteration of microbiota could increase the mRNA expression of Claudin-1, Zo-1, and Occludin-1 in the GPAM-G group in the ileum; only Occludin-1 was increased in the GPAM-GE group. The sIgA in the ileum showed a positive response, also the result of body weight and histology in both the GPAM-G and GPAM-GE group. These results indicated that the L. plantarum G83 could improve the intestinal barrier to defense the enterotoxigenic E. coli K88 invasion.

Enterotoxigenic Escherichia coli infection promotes enteric defensin expression via FOXO6-METTL3-m6A-GPR161 signalling axis

ABSTRACT The production of natural antimicrobial peptides has emerged as an important mechanism of innate immunity in animals. Defensins, members of a large family of antimicrobial peptides, have been suggested as effector molecules in host defence against bacteria, fungi, protozoa and enveloped viruses. However, the molecular mechanism underlying defensin upregulation in bacterial infection remains poorly understood. The modification of mRNA by N6-adenosine methylation (m6A) on internal bases influences gene expression in eukaryotes. Here, we show that β-defensin production triggered by Enterotoxigenic Escherichia coli K88 (E. coli K88) infection is controlled by the cellular m6A methyltransferase METTL3. Adding back with METTL3 robustly stimulated the re-expression of defensin, which further supports the conclusion. Furthermore, using a MeRIP-seq approach, we identified a functional connection between m6A dependent GPR161 signalling and the expression of defensins. Mechanistically, we found that the transcription factor FOXO6 interacted with METTL3 to trigger the transcription of GPR161 and the subsequent regulation of β-defensin expression. The study has shed light on the mechanisms by which enterotoxigenic Escherichia coli infection promotes enteric defensin expression.