Vitamin C alleviates acute enterocolitis in Campylobacter jejuni infected mice

Human foodborne infections with the zoonotic pathogen Campylobacter jejuni are on the rise and constitute a significant socioeconomic burden worldwide. The health-beneficial, particularly anti-inflammatory effects of vitamin C (ascorbate) are well known. In our preclinical intervention study, we assessed potential anti-pathogenic and immunomodulatory effects of ascorbate in C. jejuni-infected secondary abiotic IL-10-/- mice developing acute campylobacteriosis similar to humans. Starting 4 days prior peroral C. jejuni-infection, mice received synthetic ascorbate via the drinking water until the end of the experiment. At day 6 post-infection, ascorbate-treated mice harbored slightly lower colonic pathogen loads and suffered from less severe C. jejuni-induced enterocolitis as compared to placebo control animals. Ascorbate treatment did not only alleviate macroscopic sequelae of infection, but also dampened apoptotic and inflammatory immune cell responses in the intestines that were accompanied by less pronounced pro-inflammatory cytokine secretion. Remarkably, the anti-inflammatory effects of ascorbate pretreatment in C. jejuni-infected mice were not restricted to the intestinal tract but could also be observed in extra-intestinal compartments including liver, kidneys and lungs. In conclusion, due to the potent anti-inflammatory effects observed in the clinical murine C. jejuni-infection model, ascorbate constitutes a promising novel option for prophylaxis and treatment of acute campylobacteriosis

Carvacrol ameliorates acute campylobacteriosis in a clinical murine infection model

Background: The prevalence of human infections with the zoonotic pathogen Campylobacter jejuni is rising worldwide. Therefore, the identification of compounds with potent anti-pathogenic and anti-inflammatory properties for future therapeutic and/or preventive application to combat campylobacteriosis is of importance for global health. Results of recent studies suggested carvacrol (4-isopropyl-2-methylphenol) as potential candidate molecule for the treatment of campylobacteriosis in humans and for the prevention of Campylobacter colonization in farm animals. Results: To address this in a clinical murine infection model of acute campylobacteriosis, secondary abiotic IL-10-/- mice were subjected to synthetic carvacrol via the drinking water starting 4 days before peroral C. jejuni challenge. Whereas at day 6 post-infection placebo treated mice suffered from acute enterocolitis, mice from the carvacrol cohort not only harbored two log orders of magnitude lower pathogen loads in their intestines, but also displayed significantly reduced disease symptoms. Alleviated campylobacteriosis following carvacrol application was accompanied by less distinct intestinal apoptosis and pro-inflammatory immune responses as well as by higher numbers of proliferating colonic epithelial cells. Remarkably, the inflammation-ameliorating effects of carvacrol treatment were not restricted to the intestinal tract, but could also be observed in extra-intestinal organs such as liver, kidneys and lungs and, strikingly, systemically as indicated by lower IFN-γ, TNF, MCP-1 and IL-6 serum concentrations in carvacrol versus placebo treated mice. Furthermore, carvacrol treatment was associated with less frequent translocation of viable C. jejuni originating from the intestines to extra-intestinal compartments. Conclusion: The lowered C. jejuni loads and alleviated symptoms observed in the here applied clinical murine model for human campylobacteriosis highlight the application of carvacrol as a promising novel option for both, the treatment of campylobacteriosis and hence, for prevention of post-infectious sequelae in humans, and for the reduction of C. jejuni colonization in the intestines of vertebrate lifestock animals

Towards a Standardized Antimicrobial Susceptibility Testing Method for Mycoplasma hyorhinis

Conducting antimicrobial susceptibility testing (AST) in a comparable manner requires the availability of a standardized method. Organizations, such as the Clinical and Laboratory Standards Institute (CLSI) or the European Committee on Antimicrobial Susceptibility Testing (EUCAST), provide standardized protocols for a range of fastidious bacteria but not for Mycoplasma hyorhinis. We developed a broth microdilution method for testing M. hyorhinis in a standardized and harmonized way using a modified Friis broth devoid of antimicrobial or otherwise bacterial growth-inhibiting agents. The type strain M. hyorhinis DSM 25591 was chosen to establish the methodology. The antimicrobial agents of interest were doxycycline, enrofloxacin, erythromycin, florfenicol, gentamicin, marbofloxacin, tetracycline, tiamulin, tilmicosin, tulathromycin, and tylosin, tested by using commercial SensititreTM microtiter plates. In addition, the suitability of the methodology was evaluated via variation of the individual ingredients of the modified Friis broth by either using different batches or choosing other distributors. Despite these alterations, the method provided reliable results. We obtained repeatable minimal inhibitory concentrations for all six tested field isolates and the M. hyorhinis type strain. With this newly proposed method, we aim to provide an improved AST method for diagnostic laboratories and monitoring purposes with better comparability between times and countries. In addition, this new method will allow for an improvement of targeted treatments using antimicrobial agents and thereby reduce the options for resistance development

Whole-Genome Sequence of the Mycoplasma (Mesomycoplasma) hyorhinis DSM 25591 Type Strain

The whole-genome sequence of the type strain Mycoplasma (Mesomycoplasma) hyorhinis DSM 25591 is reported and compared to the available sequences of the corresponding type strains from other strain collections to ascertain conformity. Knowledge of the identity of type strains is of importance for their application in standardized test systems

Antimicrobial effect of a drinking water additive comprising four organic acids on Campylobacter load in broilers and monitoring of bacterial susceptibility

Application of organic acids via feed or drinking water is under discussion as a possible intervention strategy to reduce Campylobacter (C.) load in primary poultry production. A previous in vitro study showed that reduced concentrations of sorbic acid, benzoic acid, propionic acid, and acetic acid were required for antibacterial activity against Campylobacter when using a mixture of these 4 acids compared to when using the single acids. The present study aimed at determining the antibacterial efficiency of this combination in vivo as a drinking water additive for reducing shedding and intestinal C. jejuni colonization in broilers. Furthermore, we assessed whether the inoculated C. jejuni strain BfR-CA-14430 adapted in vivo to the applied organic acids. Results of this study showed that adding the organic acids consistently reduced Campylobacter loads in cloacal swabs. While significant reductions were observed within the entire study period, a maximum 2 log reduction occurred at an age of 18 d. However, after dissection at the end of the trial, no significant differences were detected in Campylobacter loads of cecal and colon contents compared to the control group. Susceptibility testing of re-isolates from cloacal swabs and cecal content revealed equal minimum inhibitory concentration (MIC) values compared to the inoculated test strain, suggesting that C. jejuni remained susceptible throughout the trial

Wild Boars Carry Extended-Spectrum β-Lactamase- and AmpC-Producing Escherichia coli

Extended-spectrum β-lactamase (ESBL)-producing Escherichia coli and methicillin-resistant Staphylococcus aureus (MRSA) represent major healthcare concerns. The role of wildlife in the epidemiology of these bacteria is unclear. The purpose of this study was to determine their prevalence in wild boars in Germany and to characterize individual isolates. A total of 375 fecal samples and 439 nasal swabs were screened for the presence of ESBL-/AmpC-E. coli and MRSA, respectively. The associations of seven demographic and anthropogenic variables with the occurrence of ESBL-/AmpC-E. coli were statistically evaluated. Collected isolates were subjected to antimicrobial susceptibility testing, molecular typing methods, and gene detection by PCR and genome sequencing. ESBL-/AmpC-E. coli were detected in 22 fecal samples (5.9%) whereas no MRSA were detected. The occurrence of ESBL-/AmpC-E. coli in wild boars was significantly and positively associated with human population density. Of the 22 E. coli, 19 were confirmed as ESBL-producers and carried genes belonging to blaCTX-M group 1 or blaSHV-12. The remaining three isolates carried the AmpC-β-lactamase gene blaCMY-2. Several isolates showed additional antimicrobial resistances. All four major phylogenetic groups were represented with group B1 being the most common. This study demonstrates that wild boars can serve as a reservoir for ESBL-/AmpC-producing and multidrug-resistant E. coli

Characteristics of methicillin-resistant Staphylococcus aureus from broiler farms in Germany are rather lineage- than source-specific

Methicillin-resistant Staphylococcus aureus (MRSA) are a major concern for public health, and broiler farms are a potential source of MRSA isolates. In this study, a total of 56 MRSA isolates from 15 broiler farms from 4 different counties in Germany were characterised phenotypically and genotypically. Spa types, dru types, SCCmec types, and virulence genes as well as resistance genes were determined by using a DNA microarray or specific PCR assays. In addition, PFGE profiles of isolates were used for analysis of their epidemiological relatedness. While half of the isolates belonged to spa type t011, the other half was of spa types t1430 and t034. On 3 farms, more than 1 spa type was found. The most common dru type was dt10a (n = 19), followed by dt11a (n = 17). Susceptibility testing of all isolates by broth microdilution revealed 21 different resistance phenotypes and a wide range of resistance genes was present among the isolates. Up to 10 different resistance phenotypes were found on individual farms. Resistance to tetracyclines (n = 53), MLSB antibiotics (n = 49), trimethoprim (n = 38), and elevated MICs of tiamulin (n = 29) were most commonly observed. Microarray analysis detected genes for leucocidin (lukF/S), haemolysin gamma (hlgA), and other haemolysines in all isolates. In all t1430 isolates, the egc cluster comprising of genes encoding enterotoxin G, I, M, N, O, U, and/or Y was found. The splitstree analysis based on microarray and PCR gene profiles revealed that all CC9/SCCmec IV/t1430/dt10a isolates clustered apart from the other isolates. These findings confirm that genotypic patterns were specific for clonal lineages rather than for the origin of isolates from individual farms

Evaluation of a Method for Standardized Antimicrobial Susceptibility Testing with Mycoplasma hyorhinis Field Isolates

Organizations like the Clinical and Laboratory Standards Institute (CLSI) or the European Committee of Antimicrobial Susceptibility Testing (EUCAST) provide standardized methodologies for antimicrobial susceptibility testing of a wide range of nonfastidious and fastidious bacteria, but so far not for Mycoplasma spp. of animal origin. Recently, a proposed method for the standardized broth microdilution testing of Mycoplasma hyorhinis using commercial Sensititre microtiter plates was presented. In this study, we evaluated this broth microdilution method with 37 field isolates and tested their susceptibility toward the following antimicrobial agents: doxycycline, enrofloxacin, erythromycin, florfenicol, gentamicin, marbofloxacin, tetracycline, tiamulin, tilmicosin, tulathromycin, and tylosin. The isolates originated from different countries, isolation sites, and years. The broth microdilution method was carried out using a modified Friis broth as the culture and test medium. For macrolides and lincosamides, a bimodal distribution with elevated MIC values could be observed for almost half of the tested field isolates, deducing reduced susceptibility toward these substances. With a recently published protocol, we were able to test a variety of field isolates, and consistent data could be obtained. Using this method, monitoring studies of Mycoplasma hyorhinis isolates can be carried out in a comparable manner, and the observed susceptibility profiles can be screened for possible changes in MIC values in the future

Influence of systemic fluoroquinolone administration on the presence of Pasteurella multocida in the upper respiratory tract of clinically healthy calves

The influence of enrofloxacin administration (5 mg/kg) for five consecutive days on the occurrence of Pasteurella multocida in the upper respiratory tract of two healthy calves was monitored over a 10-day period. From nasal swabs of two additional healthy control calves, which received a placebo saline administration, P. multocida was isolated throughout the study period. In the enrofloxacin treated calves, P. multocida was not demonstrated in the nasopharynx from 48 h after the first injection until two days after the last administration, when P. multocida reappeared and proved to be clonal in nature to the original isolates. During the experiment, no change in minimal inhibitory concentration for enrofloxacin of the P. multocida isolates was detected (MIC ≤ 0.015 μg/mL). Enrofloxacin concentrations were determined in the plasma by a high-performance liquid chromatography method with fluorescence detection. The PK/PD indices AUC/MIC and Cmax/MIC ratio were calculated and found to be 1157.7 and 129.8, respectively. Remarkably, the respiratory pathogen Arcanobacterium pyogenes became the predominant recovered organism in the nasopharynx of one animal following enrofloxacin therapy throughout the remaining of the experiment

Curcumin Mitigates Immune-Induced Epithelial Barrier Dysfunction by Campylobacter jejuni

Campylobacter jejuni (C. jejuni) is the most common cause of foodborne gastroenteritis worldwide. The bacteria induce diarrhea and inflammation by invading the intestinal epithelium. Curcumin is a natural polyphenol from turmeric rhizome of Curcuma longa, a medical plant, and is commonly used in curry powder. The aim of this study was the investigation of the protective effects of curcumin against immune-induced epithelial barrier dysfunction in C. jejuni infection. The indirect C. jejuni-induced barrier defects and its protection by curcumin were analyzed in co-cultures with HT-29/B6-GR/MR epithelial cells together with differentiated THP-1 immune cells. Electrophysiological measurements revealed a reduction in transepithelial electrical resistance (TER) in infected co-cultures. An increase in fluorescein (332 Da) permeability in co-cultures as well as in the germ-free IL-10-/- mouse model after C. jejuni infection was shown. Curcumin treatment attenuated the C. jejuni-induced increase in fluorescein permeability in both models. Moreover, apoptosis induction, tight junction redistribution, and an increased inflammatory response-represented by TNF-α, IL-1β, and IL-6 secretion-was observed in co-cultures after infection and reversed by curcumin. In conclusion, curcumin protects against indirect C. jejuni-triggered immune-induced barrier defects and might be a therapeutic and protective agent in patients