Possible reservoirs of thermotolerant Campylobacter at the farm between rearing periods and after the use of enrofloxacin as a therapeutic treatment.

Campylobacteriosis is a zoonosis and the most frequent cause of food-borne bacterial enteritis in humans. C. jejuni and C. coli are the most common species implicated in campylobacteriosis. Broilers and their products are considered the most important food sources of human infections. The aim of the present study was to evaluate the presence of thermotolerant Campylobacter in different reservoirs at the farm, and the permanence of this pathogen during four consecutive rearing periods. The samples were taken from the same house farm in the downtime period and during the last week of broiler rearing, prior to their slaughter during four consecutive cycles. Different reservoirs as potential sources of Campylobacter were analysed. The prevalence of Campylobacter in vectors was 23% in A. diaperinus larvae, 20% in wild birds, 13% in A. diaperinus adults, and 9% in flies; as regards fomites, the prevalence was 50% in workers' boots, 27% in litter, and 21% in feed, while in broilers it was 80%. Campylobacter jejuni was the most detected species (51%) in the samples analysed. In addition, some Campylobacter genotypes persisted in the house farm throughout consecutive rearing periods, indicating that those strains remain during downtime periods. However, our study could not identify the Campylobacter sources in the downtime periods because all the samples were negative for Campylobacter isolation. In addition, a remarkable finding was the effect of the use of enrofloxacin (as a necessary clinical intervention for flock health) in cycle 3 on the Campylobacter population. No Campylobacter could be isolated after that clinic treatment. Afterwards, we found a greater proportion of C. coli isolates, and the genotypes of those isolates were different from the genotypes found in the previous rearing periods. In conclusion, the effect of the use of enrofloxacin during the rearing period changed the Campylobacter species proportion, and this finding is particularly interesting for further evaluation. Furthermore, more studies should be conducted with the aim of detecting the Campylobacter sources between rearing periods.

Campylobacter sp.: Pathogenicity factors and prevention methods-new molecular targets for innovative antivirulence drugs?

Infections caused by bacterial species from the genus Campylobacter are one of the four main causes of strong diarrheal enteritis worldwide. Campylobacteriosis, a typical food-borne disease, can range from mild symptoms to fatal illness. About 550 million people worldwide suffer from campylobacteriosis and lethality is about 33 million p.a. This review summarizes the state of the current knowledge on Campylobacter with focus on its specific virulence factors. Using this knowledge, multifactorial prevention strategies can be implemented to reduce the prevalence of Campylobacter in the food chain. In particular, antiadhesive strategies with specific adhesion inhibitors seem to be a promising concept for reducing Campylobacter bacterial load in poultry production. Antivirulence compounds against bacterial adhesion to and/or invasion into the host cells can open new fields for innovative antibacterial agents. Influencing chemotaxis, biofilm formation, quorum sensing, secretion systems, or toxins by specific inhibitors can help to reduce virulence of the bacterium. In addition, the unusual glycosylation of the bacterium, being a prerequisite for effective phase variation and adaption to different hosts, is yet an unexplored target for combating Campylobacter sp. Plant extracts are widely used remedies in developing countries to combat infections with Campylobacter. Therefore, the present review summarizes the use of natural products against the bacterium in an attempt to stimulate innovative research concepts on the manifold still open questions behind Campylobacter towards improved treatment and sanitation of animal vectors, treatment of infected patients, and new strategies for prevention. KEY POINTS: • Campylobacter sp. is a main cause of strong enteritis worldwide. • Main virulence factors: cytolethal distending toxin, adhesion proteins, invasion machinery. • Strong need for development of antivirulence compounds.

Validation of the T86I mutation in the gyrA gene as a highly reliable real time PCR target to detect Fluoroquinolone-resistant Campylobacter jejuni.

BACKGROUND: Campylobacter jejuni is a leading cause of bacterial diarrhea worldwide, and increasing rates of fluoroquinolone (FQ) resistance in C. jejuni are a major public health concern. The rapid detection and tracking of FQ resistance are critical needs in developing countries, as these antimicrobials are widely used against C. jejuni infections. Detection of point mutations at T86I in the gyrA gene by real-time polymerase chain reaction (RT-PCR) is a rapid detection tool that may improve FQ resistance tracking. METHODS: C. jejuni isolates obtained from children with diarrhea in Peru were tested by RT-PCR to detect point mutations at T86I in gyrA. Further confirmation was performed by sequencing of the gyrA gene. RESULTS: We detected point mutations at T86I in the gyrA gene in 100% (141/141) of C. jejuni clinical isolates that were previously confirmed as ciprofloxacin-resistant by E-test. No mutations were detected at T86I in gyrA in any ciprofloxacin-sensitive isolates. CONCLUSIONS: Detection of T86I mutations in C. jejuni is a rapid, sensitive, and specific method to identify fluoroquinolone resistance in Peru. This detection approach could be broadly employed in epidemiologic surveillance, therefore reducing time and cost in regions with limited resources.

New and alternative strategies for the prevention, control, and treatment of antibiotic-resistant Campylobacter.

Campylobacter is an enteric pathogen and a leading bacterial cause of diarrhea worldwide. It is widely distributed in food animal species and is transmitted to humans primarily through the foodborne route. While generally causing self-limited diarrhea in humans, Campylobacter may induce severe or systemic infections in immunocompromised or young/elderly patients, which often requires antibiotic therapy with the first-line antibiotics including fluoroquinolones and macrolides. Over the past decades, Campylobacter has acquired resistance to these clinically significant antibiotics, compromising the effectiveness of antibiotic treatments. To address this concern, many studies have been conducted to advance novel and alternative measures to control antibiotic-resistant Campylobacter in animal reservoirs and in the human host. Although some of these undertakings have yielded promising results, efficacious and reliable alternative approaches are yet to be developed. In this review article, we will describe Campylobacter-associated disease spectrums and current treatment options, discuss the state of antibiotic resistance and alternative therapies, and provide an evaluation of various approaches that are being developed to control Campylobacter infections in animal reservoirs and the human host.

Campylobacter: A Review of New Promising Remedies with Medicinal Plants and Natural Antioxidants.

Campylobacter (curved bacteria) is considered one of the most important and common zoonotic bacteria and the three leading bacterial causes of gastroenteritis and diarrhea. Antibacterial resistance is growing and expanding. The aim of this review article is to report anti-Campylobacter medicinal plants. For this purpose, the search terms consisting of Campylobacter, medicinal plants, essential oil, extract, and traditional medicine were used to retrieve the relevant articles published in the journals indexed in Information Sciences Institute, Web of Science, PubMed, Scopus, Google Scholar, and Scientific Information Databases. Then, the findings of eligible articles were analyzed. According to the analysis, 71 medicinal plants were found to exert anti-Campylobacter effect. The active compounds of these plants are possibly nature-based antibiotic agents that are effective on Campylobacter. If these compounds are isolated, purified, and studied in pharmaceutical investigations, they can be used to produce nature-based, anti-Campylobacter antibiotics.

Quinolone and Macrolide-Resistant Campylobacter jejuni in Pediatric Gastroenteritis Patients from Central Iran.

Aims: To determine the prevalence and the antibiotic resistance patterns of Campylobacter jejuni isolated from pediatric diarrhea patients in central Iran. Materials and Methods: Stool specimens (n = 230) were investigated using a modified Gram stain, two specific culture media, and C. jejuni-specific PCR. Antibiotic resistance profiles and relevant resistance genes were determined. Genetic relationships among a selection of the isolates were studied by Fla typing. Results: Out of the 230 diarrhea samples, 48 (20.8%) cases of C. jejuni were identified using modified Gram stain, 45 (19.5%) using the culture media, and 76 (33%) cases were identified using PCR. The highest antibiotic resistance rates were observed in 37 (82.2%) strains against tetracycline, in 32 (71.1%) against ciprofloxacin, and in 31 (68.8%) against erythromycin. Twenty (44.4%) isolates were resistant to ciprofloxacin and erythromycin simultaneously. Genotypic investigations found 36 (97.3%) strains carrying the tet (o) gene, 31 (96.8%) harboring the cmeB gene, 22 (68.7%) strains with the gyrA6 gene, 20 (64.5%) strains containing a 23S rRNA mutation, and 21 (65.6%) strains with the qnrS gene. Fla typing of a random subset of 14 strains revealed 11 different types showing the genomic diversity of the isolates. Strains sharing the same Fla type could be easily distinguished by their resistance gene profile. Conclusions: This is the first study to demonstrate that genetically diverse quinolone-macrolide-resistant C. jejuni is an important cause of gastroenteritis in children from central Iran. Pediatricians should consider these resistance features once the antibiotic prescription is necessary for prevention of possible complications, especially in those under 5 years of age. Of note, most cases of Campylobacter diarrhea are self-limiting and antibiotics should only be prescribed in those cases where severe complications evolve.

Antimicrobial Resistance in Campylobacter Species: Mechanisms and Genomic Epidemiology.

The Campylobacter genus is a large and diverse group of Gram-negative bacteria that are known to colonize humans and other mammals, birds, reptiles, and shellfish. While it is now recognized that several emerging Campylobacter species can be associated with human disease, two species, C. jejuni and C. coli, are responsible for the vast majority of bacterial gastroenteritis in humans worldwide. Infection with C. jejuni, in particular, has also been associated with a number of extragastrointestinal manifestations and autoimmune conditions, most notably Guillain-Barré syndrome. The antimicrobial drugs of choice for the treatment of severe Campylobacter infection include macrolides, such as erythromycin, clarithromycin, or azithromycin. Fluoroquinolones, such as ciprofloxacin, are also commonly used for empirical treatment of undiagnosed diarrheal disease. However, resistance to these and other classes of antimicrobial drugs is increasing and is a major public health problem. The US Centers for Disease Control and Prevention estimates that over 300,000 infections per year are caused by drug-resistant Campylobacter. In this chapter, we discuss the taxonomy of the Campylobacter genus, the clinical and global epidemiological aspects of Campylobacter infection, with an emphasis on C. jejuni and C. coli, and issues related to the treatment of infection and antimicrobial resistance mechanisms. We further discuss the use of next-generation sequencing for the detection and surveillance of antimicrobial resistance genes.

Therapeutic administration of enrofloxacin in mice does not select for fluoroquinolone resistance in Campylobacter jejuni.

Enrofloxacin is registered for therapeutic use in beef cattle to treat bovine respiratory disease in Canada. A murine model was used to experimentally examine the impact of therapeutic administration of enrofloxacin on fluoroquinolone resistance development in Campylobacter jejuni. Administration of enrofloxacin to mice via subcutaneous injection or per os routes resulted in equivalent levels of bioactive enrofloxacin within the intestine, but bioactivity was short-lived (<48 h after cessation). Enrofloxacin administration did not affect densities of total bacteria, Firmicutes, or Bacteroidetes in digesta and had modest impacts on densities of Enterobacteriaceae. All mice inoculated with C. jejuni NCTC 11168 became persistently colonized by the bacterium. Enrofloxacin reduced C. jejuni cell densities within the cecal and colonic digesta for all treatments, and densities shed in feces as a function of antibiotic duration. None of the C. jejuni isolates recovered from mice after administration of enrofloxacin (n = 260) developed resistance to ciprofloxacin regardless of method or duration of administration. Furthermore, only modest shifts in the minimum inhibitory concentration of the isolates by treatment were noted. The study findings indicate that the risk posed by short-term subcutaneous administration of enrofloxacin for the development of fluoroquinolone resistance in mammals is low.

Pharmacokinetic and antimicrobial activity of a new carvacrol-based product against a human pathogen, Campylobacter jejuni.

AIM: In vitro and in vivo studies were conducted to test a new carvacrol-based product designed to delay the carvacrol release so that it could reach the caeca of broiler chickens in order to control Campylobacter jejuni. METHODS AND RESULTS: Antimicrobial activity of carvacrol, a constituent of oregano and thyme essential oil, has been demonstrated against C. jejuni in vitro, and this compound was found beneficial for broiler growth. Here, we tested a new liquid formulation that did not change the antibacterial efficacy of carvacrol against C. jejuni in vitro, as assessed by broth microdilution. The mode of action of carvacrol also remained unchanged as illustrated by electronic microscopy. A pharmacokinetic assay monitored carvacrol of the solid galenic formulation in the avian digestive tract and this showed that this compound was mainly found in the last part (caeca, large intestine) and in the droppings. Extremely low concentrations of free carvacrol were present in blood plasma, with larger amounts of carvacrol metabolites: carvacrol glucuronide and sulphate. A qPCR analysis showed that the solid galenic form of carvacrol added at 5 kg per tonne of food (i.e. 9·5 mg of carvacrol per kg of bodyweight per day) significantly decreased the C. jejuni caecal load by 1·5 log. CONCLUSIONS: The new liquid formulation was as effective as unformulated carvacrol in vitro. In vivo the solid galenic form seems to delay the carvacrol release into the caeca and presented interesting results on C. jejuni load after 35 days. SIGNIFICANCE AND IMPACT OF THE STUDY: Results suggested that this product could be promising to control Campylobacter contamination of broilers.

Campylobacter Prevalence and Quinolone Susceptibility in Feces of Preharvest Feedlot Cattle Exposed to Enrofloxacin for the Treatment of Bovine Respiratory Disease.

Campylobacter spp. can be pathogenic to humans and often harbor antimicrobial resistance genes. Data on resistance in relation to fluoroquinolone use in beef cattle are scarce. This cross-sectional study of preharvest cattle evaluated Campylobacter prevalence and susceptibility to nalidixic acid and ciprofloxacin in feedlots that previously administered a fluoroquinolone as primary treatment for bovine respiratory disease. Twenty fresh fecal samples were collected from each of 10 pens, in each of five feedlots, 1-2 weeks before harvest. Feces were cultured for Campylobacter using selective enrichment and isolation methods. Genus and species were confirmed via PCR. Minimum inhibitory concentrations (MICs) of ciprofloxacin and nalidixic acid were determined using a micro-broth dilution method and human breakpoints. Antimicrobial use within each pen was recorded. Data were analyzed using generalized linear mixed-models (prevalence) and survival analysis (MICs). Overall, sample-level prevalence of Campylobacter was 27.2% (272/1000) and differed significantly among feedlots (p < 0.01). Campylobacter coli was the most common species (55.1%; 150/272), followed by Campylobacter hyointestinalis (42.6%; 116/272). Within-pen prevalence was not significantly associated with the number of fluoroquinolone treatments, sex, body weight, or metaphylaxis use, but was associated with the number of days cattle were in the feedlot (p = 0.03). The MICs for the majority of Campylobacter isolates were above the breakpoints for nalidixic acid (68.4%; 175/256) and for ciprofloxacin (65.6%; 168/256). Distributions of MICs for nalidixic acid (p ≤ 0.01) and ciprofloxacin (p ≤ 0.05) were significantly different among feedlots, and by Campylobacter species. However, fluoroquinolone treatments, sex, body weight, days on feed, and metaphylaxis were not significantly associated with MIC distributions within pens. We found no evidence that the number of fluoroquinolone treatments within feedlot pens significantly affected the within-pen fecal prevalence or quinolone susceptibilies of Campylobacter in feedlots that used a fluoroquinolone as primary treatment for bovine respiratory disease.

TYPLEX® Chelate, a novel feed additive, inhibits Campylobacter jejuni biofilm formation and cecal colonization in broiler chickens.

Reducing Campylobacter spp. carriage in poultry is challenging, but essential to control this major cause of human bacterial gastroenteritis worldwide. Although much is known about the mechanisms and route of Campylobacter spp. colonization in poultry, the literature is scarce on antibiotic-free solutions to combat Campylobacter spp. colonization in poultry. In vitro and in vivo studies were conducted to investigate the role of TYPLEX® Chelate (ferric tyrosine), a novel feed additive, in inhibiting Campylobacter jejuni (C. jejuni) biofilm formation and reducing C. jejuni and Escherichia coli (E. coli) colonization in broiler chickens at market age. In an in vitro study, the inhibitory effect on C. jejuni biofilm formation using a plastic bead assay was investigated. The results demonstrated that TYPLEX® Chelate significantly reduces biofilm formation. In an in vivo study, 800 broilers (one d old) were randomly allocated to 4 dietary treatments in a randomized block design, each having 10 replicate pens with 20 birds per pen. At d 21, all birds were challenged with C. jejuni via seeded litter. At d 42, cecal samples were collected and tested for volatile fatty acid (VFA) concentrations and C. jejuni and E. coli counts. The results showed that TYPLEX® Chelate reduced the carriage of C. jejuni and E. coli in poultry by 2 and 1 log10 per gram cecal sample, respectively, and increased cecal VFA concentrations. These findings support TYPLEX® Chelate as a novel non-antibiotic feed additive that may help produce poultry with a lower public health risk of Campylobacteriosis.

[COMPARISON OF CLINICAL AND DEMOGRAPHIC CHARACTERISTICS OF ARAB AND JEWISH CHILDREN LIVING IN THE GALILEE WHO HAVE SUFFERED FROM AN INTESTINAL INFECTION CAUSED BY CAMPYLOBACTER].

INTRODUCTION: Among all infectious agents that cause gastrointestinal infection in children, the most common is the Campylobacter bacterium. The bacterium has multiple virulence factors such as motility, adhesion and invasion of the human intestinal lining, and enzyme secretion. In recent years, there has been a worldwide increase in Campylobacter resistance to antibiotics. AIMS: To examine the frequency of Campylobacter among children who were hospitalized at the Poriya Medical Center during 2012-2014 and suffered from an intestinal infection caused by Campylobacter; to compare the demographic, clinical, and laboratory data of Jewish and Arab children; to examine the resistance rate of the bacterium to antibiotics. METHODS: The data on Campylobacter frequency in children who suffered from an intestinal infection was extracted from the medical records: age, sex, hospitalization duration, hemoglobin and leukocyte values in blood chemistry, the residential environment, and antibiotic treatment during hospitalization. In addition, antibiotic susceptibility tests were performed for Erythromycin and Ciprofloxacin for all Campylobacter cultures that were isolated from patients' stool samples and kept frozen. RESULTS: Campylobacter is the most prevalent bacterial factor among children who were hospitalized following enteritis. There are differences in the bacterium frequency among Jewish children in comparison to frequency in Arab children in the following aspects: Campylobacter is more frequent in Arab children, more common among children living in rural areas, and especially those of Arab origin. Arab children were hospitalized for longer durations than Jewish children. The mean age of Jewish children who suffered from infection caused by Campylobacter was higher compared to the mean age of Arab children. No difference was found in leukocyte values in the cell count. Hemoglobin values were lower among Jewish children compared to Arab children. There was a high percentage of children treated with antibiotics due to intestinal infection caused by Campylobacter, especially among Arab children. Resistance to Erythromycin was not found; however the rate of resistance to Ciprofloxacin was 10.7%. CONCLUSIONS: There are significant differences in intestinal infection caused by Campylobacter among Jewish and Arab children in parameters such as: mean age, hospitalization duration, and residential area. The antibiotic resistance rate that was found is low; however, presently, it still exists.

Antimicrobial Use for and Resistance of Zoonotic Bacteria Recovered from Nonhuman Primates.

As a growing threat to human and animal health, antimicrobial resistance (AMR) has become a central public-health topic. Largescale surveillance systems, such as the National Antimicrobial Resistance Monitoring System (NARMS), are now established to monitor and provide guidance regarding AMR, but comprehensive literature on AMR among NHP is sparse. This study provides data regarding current antimicrobial use strategies and the prevalence of AMR in zoonotic bacteria recovered from NHP within biomedical research institutions. We focused on 4 enteric bacteria: Shigella flexneri, Yersinia enterocolitica, Y. pseudotuberculosis, and Campylobacter jejuni. Fifteen veterinarians, 7 biomedical research institutions, and 4 diagnostic laboratories participated, providing susceptibility test results from January 2012 through April 2015. Veterinarians primarily treated cases caused by S. flexneri, Y. enterocolitica, and Y. pseudotuberculosis with enrofloxacin but treated C. jejuni cases with azithromycin and tylosin. All isolates were susceptible to the associated primary antimicrobial but often showed resistance to others. Specifically, S. flexneri isolates frequently were resistant to erythromycin (87.5%), doxycycline (73.7%), and tetracycline (38.3%); Y. enterocolitica isolates to ampicillin (100%) and cefazolin (93.6%); and C. jejuni isolates to methicillin (99.5%) and cephalothin (97.5%). None of the 58 Y. pseudotuber-culosis isolates was resistant to any tested antimicrobial. Notably, resistance patterns were not shared between this study's NHP isolates and human isolates presented by NARMS. Our findings indicate that zoonotic bacteria from NHP diagnostic samples are broadly susceptible to the antimicrobials used to treat the clinical infections. These results can help veterinarians ensure effective antimicrobial therapy and protect staff by minimizing occupational risk.

Identification and antimicrobial resistance patterns of bacterial enteropathogens from children aged 0-59 months at the University Teaching Hospital, Lusaka, Zambia: a prospective cross sectional study.

BACKGROUND: Bacterial diarrhoeal disease is among the most common causes of mortality and morbidity in children 0-59 months at the University Teaching Hospital in Lusaka, Zambia. However, most cases are treated empirically without the knowledge of aetiological agents or antimicrobial susceptibility patterns. The aim of this study was, therefore, to identify bacterial causes of diarrhoea and determine their antimicrobial susceptibility patterns in stool specimens obtained from the children at the hospital. METHODS: This hospital-based cross-sectional study involved children aged 0-59 months presenting with diarrhoea at paediatrics wards at the University Teaching Hospital in Lusaka, Zambia, from January to May 2016. Stool samples were cultured on standard media for enteropathogenic bacteria, and identified further by biochemical tests. Multiplex polymerase chain reaction was used for characterization of diarrhoeagenic Escherichia coli strains. Antimicrobial susceptibility testing was performed on antibiotics that are commonly prescribed at the hospital using the Kirby-Bauer disc diffusion method, which was performed using the Clinical Laboratory Standards International guidelines. RESULTS: Of the 271 stool samples analysed Vibrio cholerae 01 subtype and Ogawa serotype was the most commonly detected pathogen (40.8%), followed by Salmonella species (25.5%), diarrhoeagenic Escherichia coli (18%), Shigella species (14.4%) and Campylobacter species (3.5%). The majority of the bacterial pathogens were resistant to two or more drugs tested, with ampicillin and co-trimoxazole being the most ineffective drugs. All diarrhoeagenic Escherichia coli isolates were extended spectrum ß-lactamase producers. CONCLUSION: Five different groups of bacterial pathogens were isolated from the stool specimens, and the majority of these organisms were multidrug resistant. These data calls for urgent revision of the current empiric treatment of diarrhoea in children using ampicillin and co-trimoxazole, and emphasizes the need for continuous antimicrobial surveillance as well as the implementation of prevention programmes for childhood diarrhoea.

Antimicrobial resistance in Campylobacter coli and Campylobacter jejuni in cynomolgus monkeys (Macaca fascicularis) and eradication regimens.

BACKGROUND: Campylobacter spp. are zoonotic pathogens, however, knowledge about their presence and antimicrobial resistance in nonhuman primates is limited. Our animal facility purchased cynomolgus monkeys (Macaca fascicularis) from various Asian countries: China, Cambodia, Indonesia, the Philippines, and Vietnam. METHODS: Colonization by Campylobacter spp. was investigated in 238 of the monkeys from 2009 to 2012 and antimicrobial susceptibility testing was carried out for these isolates. Furthermore, we eradicated these pathogens from these monkeys. RESULTS: Campylobacter spp. were isolated from 47 monkeys from three specific countries: China, Cambodia, and Indonesia, with respective isolation rates of 15%, 36%, and 67%. Two monkeys, which were each infected with Campylobacter jejuni and Campylobacter coli, showed clinical symptoms of diarrhea and bloody feces. In total, 41 isolates of C. coli and 17 isolates of C. jejuni were detected. Antimicrobial susceptibility varied: in the monkeys from China, erythromycin (ERY)-, tetracycline (TET)-, and ciprofloxacin-resistant C. coli, in the monkeys from Cambodia, amoxicillin-intermediate, TET- and ciprofloxacin-resistant C. coli and amoxicillin- and ciprofloxacin-resistant C. jejuni, and in the monkeys from Indonesia, ciprofloxacin-resistant C. coli and TET- and ciprofloxacin-resistant C. jejuni were common (>75%). Multiresistant isolates of C. coli were found in monkeys from all countries and multiresistant isolates of C. jejuni were found in monkeys from Indonesia. The eradication rate with azithromycin was comparable to that with gentamicin (GEN) by oral administration, and was higher than those with amoxicillin-clavulanic acid (AMC) and chloramphenicol (CHL). CONCLUSION: From the perspective of zoonosis, we should acknowledge multiresistant Campylobacter spp. isolated from the monkeys as a serious warning.

In vitro investigations into the use of antimicrobials in combination to maintain efficacy of fluoroquinolones in poultry.

The aim of this study was to determine if apramycin, colistin or lincomycin-spectinomycin, in combination with enrofloxacin, was able prevent the emergence of mutants with reduced susceptibility to fluoroquinolone antibiotics in vitro. MICs were determined for enrofloxacin alone and in combination for panels of Campylobacter (n=37), Escherichia coli (n=52) and Salmonella (n=52) isolates. MIC results suggested that apramycin, colistin and lincomycin-spectinomycin worked in an additive/indifferent way when each was combined with enrofloxacin. Apramycin was considered the most promising antibiotic for combination-therapy in conjunction with enrofloxacin, and further evaluations (MBCs, MPCs and time-kill-curves) were performed for this combination for selected isolates. Results suggest combination-therapy of enrofloxacin with apramycin increases the efficacy, as well as decreasing the emergence and survival of bacteria with mutational resistance to fluoroquinolone antibiotics. Such combination-therapy, minimising the development of mutational resistance, may have relevance for Campylobacter, E. coli and Salmonella infections in poultry.

[Invasive Campylobacter jejuni/coli Infections: 9 Case Reports at a Single Center between 2000 and 2015, and a Review of Literature Describing Japanese Patients].

There have been few coherent reports on extraintestinal infection or bacteremia caused by Campylobacter jejuni (C. jejuni) or C. coli in Japan. To clarify the clinical and microbiological characteristics of invasive infections caused by these two species, we retrospectively analyzed the records of patients from whom these pathogens had been isolated from sterile sites between 2000 and 2015. During this study period, we identified 9 patients. The clinical syndrome of all of these patients was bacteremia. Three patients had underlying diseases with both liver cirrhosis and malignant neoplasm, and all of these patients were aged 60 years or older. The remaining 6 patients were immunocompetent and younger than 40 years of age. All 9 patients had a fever of 38.5 degrees C or higher. The proportion of patients with gastrointestinal symptoms was lower for the 3 patients with underlying diseases, compared with the 6 patients without underlying diseases (1/3 cases vs, 4/6 cases). Of the 8 strains evaluated for antimicrobial susceptibility, all were susceptible to imipenem/cilastatin, kanamycin and erythromycin, and 2 were resistant to levofloxacin. Antimicrobial treatment was administered to 8 patients, but one spontaneously recovered without any treatment. We were able to follow the outcomes of 8 patients, and all of these patients completely recovered without relapses. We also reviewed 14 Japanese patients reported in the Japanese and English literature and found similar clinical features consisting of a high-grade fever and an association with underlying diseases and gastrointestinal symptoms. Of note, 3 agammaglobulinemic patients presented with bacteremia and extraintestinal infections and had multiple relapses. Based on the findings of our 9 cases and previous reports, the affected patients were divided into two groups according to clinical syndrome and therapeutic intervention. One group consisted of previously healthy children or young adults showing bacteremia. Most of them had enterocolitis complications but had a good prognosis. The other group consisted of patients with underlying diseases or elderly patients who presented with bacteremia alone or bacteremia with extraintestinal infections. The latter group, especially among those with humoral immunodeficiency, should be parentally treated with antimicrobial agents and requires careful monitoring for relapse. This is the largest case series study to examine invasive C. jejuni/coli infections in Japan, and it provides important epidemiological information on this rare infection.

Efficacy of feed additives against Campylobacter in live broilers during the entire rearing period: Part B.

A total of 636 day-of-hatch Ross 308 broilers chicks were used in 4 independent trials carried out to screen the effect of 12 feed additives on reducing cecal colonization of Campylobacterin broilers. The tested additives were probiotics based on B. subtilis and S. cerevisae, a garlic extract, a blend of herbal substances and essential oils, two different combinations of essential oils and organic acids (OA), two mixtures of flavoring compounds, medium chain fatty acids (MCFA), monoglycerides (MG) of MCFA and MG-MCFA+OA. At 14 days of age, all the birds were orally infected with 0.1 mL of a bacterial suspension of C. jejuni ST-45 diluted at 10(5) cfu/mL in tryptone salt broth. In each trial, there was a positive control group and 2 (Trials 1 and 2) or 4 (Trials 3 and 4) additional treatment groups supplemented with additives, which were added to feed or water only to the finisher (21 to 42 d) diet (Trials 1 and 2) or to the starter (0 to 21 d) and finisher diets (Trials 3 and 4). Feed and water were available ad libitum. On days 35 and 42 of age in Trials 1 and 2, and on days 21, 35 and 42 of age in Trials 3 and 4, 10 (Trials 1 and 2) or 12 birds (Trials 3 and 4) per group were euthanized for cecal sampling. In Trial 1, birds fed with MCFA and MG-MCFA had a significant (P<0.05) reduction in cecal Campylobacter colonization compared to control at 35 d, but only the group treated with MG-MCFA maintained the reduction at 42 d. In Trials 2 to 4, no significant differences (P>0.05) in cecal Campylobacter counts were found between the treated and control animals. In conclusion, although none of the treatments were able to completely prevent the colonization of chickens with C. jejuni, MCFA and MG-MCFAs could reduce the pathogen counts when supplemented from 21 days onwards.