Outbreak of Arcanobacterium haemolyticum in chronic wounds in The Netherlands.

INTRODUCTION: Aging and comorbidities such as diabetes and vascular problems contribute to the increasing occurrence of chronic wounds. From the beginning of 2016, a marked increase in Arcanobacterium haemolyticum (ARH) in chronic wound cultures was noted among patients visiting a wound expertise centre in The Netherlands. AIM: To report the outbreak investigation of ARH cultured from chronic wounds and describe the implemented infection prevention measures. METHODS: In total, 50 ARH isolates were sent to a reference laboratory for molecular typing. Samples for bacterial culture and ARH polymerase chain reaction were taken from care workers, the environment and items used for wound care. Infection prevention measures were implemented in a bundled approach, involving education, better aseptic wound care conditions and hygienic precautions. Before and after the implementation of infection prevention measures, two screening rounds of ARH testing were performed among all patients receiving home care. RESULTS: ARH isolates from wound care patients were found to be identical by core genome multi-locus sequence typing. No definite outbreak source could be determined by culture. However, three pairs of forceps, used by two nurses on multiple patients, were found to be ARH positive by polymerase chain reaction. In the two screening rounds before and after the implementation of infection prevention measures, the proportion of ARH-positive patients decreased significantly from 20% (20/99) to 3% (3/104). Subsequently, no new cases occurred. CONCLUSION: This first ARH outbreak was likely caused by re-using contaminated instruments. Through the implementation of improved infection prevention measures and re-education of all employees involved, the outbreak was controlled. With the current trend of care transition, infection control must be a major concern.

VIM-carbapenemase-producing Escherichia coli in a residential care home in The Netherlands.

BACKGROUND: Carbapenemase-producing Enterobacteriaceae (CPE) are an important and increasing threat to public health. In hospitals and long-term care facilities, carriers should be identified to prevent transmission; however, guidelines for infection control are not applicable to all types of care homes. AIM: To report the outbreak investigation of a VIM-metallo-ß-lactamase-producing Escherichia coli in a Dutch residential care home, where residents lived in private apartments but also used shared facilities. METHODS: Contact and environmental screening rounds were performed to assess carriage and colonization rates. Due to the domestic characteristics of the home, customized infection control measures were needed. A bundle of interventions was implemented, including contact precautions, improved hygiene and education. FINDINGS: In total, eight CPE carriers, including the index case, were identified among 110 residents. VIM-CPE spread was associated with the use of shared toilets in communal areas. Seven months after the first finding, all carriers were found to be VIM-negative, and after 1 year, VIM CPE was no longer detectable in the environment. CONCLUSION: A customized bundled approach was needed to control the outbreak successfully. Current guidelines should be adapted to be suitable for all types of residential care homes in order to combat the spread of multi-resistant pathogens effectively.

Reduced length of hospital stay through a point of care placed automated blood culture instrument.

Early appropriate antimicrobial treatment of patients with sepsis has a large impact on clinical outcome. To enable prompt and efficient processing of blood cultures, the inoculated vials should be placed into an automated continuously monitoring blood culture system immediately after sampling. We placed an extra BACTEC FX instrument at the emergency department of our hospital and validated the twice-daily re-entering of ongoing vials from this instrument into the BACTEC FX at the laboratory. We subsequently assessed the benefits of shortening the transport time between sampling and monitored incubation of blood culture vials by comparing the turnaround times of positive blood cultures from emergency department patients with a historical control group. Re-entering ongoing vials within 2 h raised no technical problems with the BACTEC FX and did not increase the risk of false-negative culture results. The decreased transport time resulted in significantly earlier available Gram stain results for a large proportion of patients in the intervention group and a significant shortening of the median total turnaround time to less than 48 h. The median length of hospital stay shortened by 1 day. Immediate entering of blood culture vials into a point of care placed BACTEC FX instrument and subsequent efficient processing enables earlier decision-making regarding antimicrobial treatment, preventing the development of antimicrobial resistance and reducing healthcare costs.

Randomised clinical study: Aspergillus niger-derived enzyme digests gluten in the stomach of healthy volunteers.

BACKGROUND: Aspergillus niger prolyl endoprotease (AN-PEP) efficiently degrades gluten molecules into non-immunogenic peptides in vitro. AIM: To assess the efficacy of AN-PEP on gluten degradation in a low and high calorie meal in healthy subjects. METHODS: In this randomised, double-blind, placebo-controlled, cross-over study 12 healthy volunteers attended to four test days. A liquid low or high calorie meal (4 g gluten) with AN-PEP or placebo was administered into the stomach. Via a triple-lumen catheter gastric and duodenal aspirates were sampled, and polyethylene glycol (PEG)-3350 was continuously infused. Acetaminophen in the meals tracked gastric emptying time. Gastric and duodenal samples were used to calculate 240-min area under the curve (AUC0-240 min ) of ?-gliadin concentrations. Absolute ?-gliadin AUC0-240 min was calculated using duodenal PEG-3350 concentrations. RESULTS: AN-PEP lowered α-gliadin concentration AUC0-240 min, compared to placebo, from low and high calorie meals in stomach (low: 35 vs. 389 µg × min/mL; high: 53 vs. 386 µg × min/mL; P < 0.001) and duodenum (low: 7 vs. 168 µg × min/mL; high: 4 vs. 32 µg × min/mL; P < 0.001) and absolute α-gliadin AUC0-240 min in the duodenum from low (2813 vs. 31 952 µg × min; P < 0.001) and high (2553 vs. 13 095 µg × min; P = 0.013) calorie meals. In the placebo group, the high compared to low calorie meal slowed gastric emptying and lowered the duodenal α-gliadin concentration AUC0-240 min (32 vs. 168 µg × min/mL; P = 0.001). CONCLUSIONS: AN-PEP significantly enhanced gluten digestion in the stomach of healthy volunteers. Increasing caloric density prolonged gastric residence time of the meal. Since AN-PEP already degraded most gluten from low calorie meals, no incremental effect was observed by increasing meal caloric density. ClinicalTrials.gov, Number: NCT01335503; www.trialregister.nl, Number: NTR2780.

Diagnosis of acute puerperal metritis by electronic nose device analysis of vaginal discharge in dairy cows.

The objective of this study was to estimate the diagnostic accuracy of an electronic nose device using vaginal discharge samples to diagnose acute puerperal metritis (APM) in dairy cows. Uterine fluid was sampled manually with a gloved hand and under sterile conditions for electronic nose device analysis (day in milk (DIM) 2, 5, and 10) and bacteriologic examination (DIM 5), respectively, and on additional days, if APM was diagnosed during the daily clinical examinations. A dataset containing samples from 70 cows was used to create a model and to validate the APM status predicted by this model, respectively. Half of the dataset (n = 35; 14 healthy and 21 metritic cows) was provided with information regarding the APM diagnosis and contained all three measurements (DIM 2, 5, and 10) for each cow and was used as a training set whereas the second half was blinded (n = 35; 14 healthy and 21 metritic cows) and contained only the samples collected on DIM 5 of each cow and was used to validate the created prediction model. A receiver operating characteristic curve was calculated using the prediction results of the validation test. The best observed sensitivity was 100% with specificity of 91.6% when using a threshold value of 0.3. The calculated P-value for the receiver operating characteristic curve was less than 0.01. Overall, Escherichia coli was isolated in eight of 28 (28.6%) and 22 of 42 (52.4%) samples collected from healthy and metritic cows, respectively. Trueperella pyogenes and Fusobacterium necrophorum were isolated in 14 and six of 28 (50.0% and 21.4%) and 17 and 16 of 42 (40.5% and 38.1%) samples collected from healthy and metritic cows, respectively. The prevalence of Escherichia coli and Trueperella pyogenes was similar in the samples obtained from metritic cows used for the training set and the validation test. The results are promising especially because of the objective nature of the measurements obtained by the electronic nose device.

Technical note: Evaluation of odor from vaginal discharge of cows in the first 10 days after calving by olfactory cognition and an electronic device.

The objective of this study was to determine test characteristics (i.e., intra- and interobserver variability, intraassay variability, sensitivity, and specificity) of an evaluation of odor from vaginal discharge (VD) of cows in the first 10 d postpartum conducted by olfactory cognition and an electronic device, respectively. In experiment 1, 16 investigators (9 veterinary students and 7 licensed veterinarians) evaluated 5 VD samples each on 10 different days. The kappa test revealed an agreement between investigators (interobserver) of κ=0.43 with a Fleiss adjusted standard error of 0.0061. The overall agreement was the same for students (κ=0.28) and veterinarians (κ=0.28). Mean agreement within observers (intraobserver) was κ=0.52 for all observers, and 0.49 and 0.62 for students and veterinarians, respectively. In experiment 2, the repeatability of an electronic device (DiagNose; C-it, Zutphen, the Netherlands) was tested. Therefore, 5 samples of VD from 5 cows were evaluated 10 times each. The repeatability was 0.97, determined by Cronbach's α. In experiment 3, 20 samples collected from healthy cows and 20 of cows with acute puerperal metritis were evaluated by the 16 investigators and the DiagNose using a dichotomous scale (1=cow with acute puerperal metritis; 0=healthy cow). Sensitivity and specificity of olfactory evaluation was 75.0 and 60.1% compared with 92.0 and 100%, respectively, for the electronic nose device. The study revealed a considerable subjectivity of the human nose concerning the classification into healthy and sick animals based on the assessment of vaginal discharge. The repeatability of the electronic nose was higher. In conclusion, the DiagNose system, although imperfect, is a reasonable tool to improve odor assessment of VD. The current system, however, is not suitable as a screening tool in the field. Further research is warranted to adapt such electronic devices to practical on-farm screening tools.

Otitis externa following aural irrigation linked to instruments contaminated with Pseudomonas aeruginosa.

BACKGROUND: The incidence of acute otitis externa, an infection of the external auditory canal, in general practitioners' (GP) practices in The Netherlands is about 14 per 1000 patients per year. In early 2010, one of the authors noted that some of the otitis externa patients in his GP practice had undergone cerumen removal by ear syringing a few weeks earlier. Bacterial cultures of samples taken from the instruments used showed contamination of an ear syringe by Pseudomonas aeruginosa. From then on, P. aeruginosa isolates from patients' ears were stored in the laboratory. AIM: It was assessed whether cross-contamination with P. aeruginosa between patients in the same GP practice could occur through the use of contaminated ear lavage instruments. METHODS: From 17 GP practices, the otolaryngology Outpatient Department and the Out-of-Hours GP Service, instruments used for examining and cleaning the outer ear were swabbed. Strains of P. aeruginosa cultured from the instruments were genotyped together with isolates of patients registered in the same practice. FINDINGS: In four practices where contaminated instruments were found, genotyping showed similarity between P. aeruginosa strains isolated from a patient and the ear syringe, and/or between strains of different patients in the same practice. CONCLUSIONS: Transmission of P. aeruginosa from ear lavage instruments to patients appears to occur with otitis externa as a result. Together with the Infection Control Unit of our hospital we have formulated recommendations for the appropriate cleaning, disinfection and storage of re-usable ear lavage instruments for the GP practices to implement.

Evaluation of three enzyme immunoassays and a loop-mediated isothermal amplification test for the laboratory diagnosis of Clostridium difficile infection.

The laboratory diagnosis of Clostridium difficile infection (CDI) consists of the detection of toxigenic Clostridium difficile, and/or its toxins A or B in stool preferably in a two-step algorithm. In a prospective study, we compared the performance of three toxin enzyme immunoassays (EIAs)-ImmunoCard Toxins A & B, Premier Toxins A & B and C. diff Quik Chek Complete, which combines a toxins test and a glutamate dehydrogenase (GDH) antigen EIA in one device -and the loop-mediated isothermal amplification assay Illumigene C. difficile. In total 986 stool samples were analyzed. Compared with toxigenic culture as the gold standard, sensitivities, specificities, PPV and NPV values of the toxin EIAs were 41.1-54.8 %, 98.9-100 %, 75.0-100 % and 95.5-96.5 % respectively, of the Illumigene assay 93.3 %, 99.7 %, 95.8 % and 99.5 %. Illumigene assays performed significantly better for non-014/020 PCR-ribotypes than for C. difficile isolates belonging to 014/020. Discrepant analysis of three culture-negative, but Illumigene-positive samples, revealed the presence of toxin genes using real-time PCRs. In addition to the GDH EIA (NPV of 99.8 %), the performance of Illumigene allows this test to be introduced as a first screening test for CDI- or as a confirmation test for GDH -positive samples, although the initial invalid Illumigene result of 4.4 % is a point of concern.

In vitro study of the antimicrobial effects of radiological contrast agents used in arthrography.

Aspiration arthrography using an iodinated contrast medium is a useful tool for the investigation of septic or aseptic loosening of arthroplasties and of septic arthritis. Previously, the contrast media have been thought to cause false negative results in cultures when present in aspirated samples of synovial fluid, probably because free iodine is bactericidal, but reports have been inconclusive. We examined the influence of the older, high osmolar contrast agents and the low osmolar media used currently on the growth of ten different micro-organisms capable of causing deep infection around a prosthesis. Five media were tested, using a disc diffusion technique and a time-killing curve method in which high and low inocula of micro-organisms were incubated in undiluted media. The only bactericidal effects were found with low inocula of Escherichia coli and Pseudomonas aeruginosa in ioxithalamate, one of the older ionic media. The low and iso-osmolar iodinated contrast media used currently do not impede culture. Future study must assess other causes of false negative cultures of synovial fluid and new developments in enhancing microbial recovery from aspirated samples.

Black tea reduces diarrhoea prevalence but decreases growth performance in enterotoxigenic Escherichia coli-infected post-weaning piglets.

Enterotoxigenic Escherichia coli (ETEC) is a main cause of diarrhoea in humans and piglets. In vitro, black tea extract (BTE) has anti-pathogenic properties. Anti-diarrhoeal properties of BTE were assessed in a pig model of gastrointestinal infection. At weaning (day 0), piglets (n = 96) were randomly assigned to a diet containing 0% (control), 0.4% or 0.8% (wt/wt) BTE during 27 days. Piglets were orally infected with 6.4 × 10(6) cfu of ETEC on day 6. Faecal consistency, feed intake and body weight were measured. In a sub-study (n = 30 piglets), the effect of BTE palatability on feed intake was assessed. Additionally, the effect of BTE on ETEC growth in the presence or absence of iron was studied in vitro. The 0.8% BTE diet reduced diarrhoea prevalence by 20% but also decreased feed intake by 16% and feed efficiency by 12% over the total period. The 0.4% BTE diet decreased feed efficiency and weight gain from day 13 onwards. The palatability study demonstrated that piglets preferred the control to the BTE diets. In vitro, BTE delayed ETEC exponential growth, which was reversed by iron addition. Although BTE had anti-diarrhoeal properties, this effect was accompanied by impaired performance. The absence of a correlation between diarrhoea prevalence and feed intake suggests that reduced diarrhoea directly results from BTE rather than from reduced feed intake caused by BTE astringency.

Evaluation of a rapid immunochromatographic test for the detection of norovirus in stool samples.

Worldwide noroviruses are an important cause of gastroenteritis and are major agents of both sporadic as well as epidemic infection. Because of the rapid transmission of the virus, early detection is essential. Until recently, the available test methods for the detection in stool were enzyme immunoassays and real-time reverse transcription polymerase chain reaction (RT-PCR), both of which take several hours to perform. We evaluated the rapid immunochromatographic test RIDA(R)QUICK Norovirus for the detection of norovirus in the stool of patients with acute gastroenteritis. This test is easy to perform and read and only takes 20 min. The sensitivity and specificity compared to RT-PCR results and the positive and negative predictive values were 57.1%, 99.1%, 93.3% and 91.2%, respectively. The rapid test is useful for quick screening, but a negative result should be followed up by RT-PCR.

Parasitological diagnosis combining an internally controlled real-time PCR assay for the detection of four protozoa in stool samples with a testing algorithm for microscopy.

Molecular detection of gastrointestinal protozoa is more sensitive and more specific than microscopy but, to date, has not routinely replaced time-consuming microscopic analysis. Two internally controlled real-time PCR assays for the combined detection of Entamoeba histolytica, Giardia lamblia, Cryptosporidium spp. and Dientamoeba fragilis in single faecal samples were compared with Triple Faeces Test (TFT) microscopy results from 397 patient samples. Additionally, an algorithm for complete parasitological diagnosis was created. Real-time PCR revealed 152 (38.3%) positive cases, 18 of which were double infections: one (0.3%) sample was positive for E. histolytica, 44 (11.1%) samples were positive for G. lamblia, 122 (30.7%) samples were positive for D. fragilis, and three (0.8%) samples were positive for Cryptosporidium. TFT microscopy yielded 96 (24.2%) positive cases, including five double infections: one sample was positive for E. histolytica/Entamoeba dispar, 29 (7.3%) samples were positive for G. lamblia, 69 (17.4%) samples were positive for D. fragilis, and two (0.5%) samples were positive for Cryptosporidium hominis/Cryptosporidium parvum. Retrospective analysis of the clinical patient information of 2887 TFT sets showed that eosinophilia, elevated IgE levels, adoption and travelling to (sub)tropical areas are predisposing factors for infection with non-protozoal gastrointestinal parasites. The proposed diagnostic algorithm includes application of real-time PCR to all samples, with the addition of microscopy on an unpreserved faecal sample in cases of a predisposing factor, or a repeat request for parasitological examination. Application of real-time PCR improved the diagnostic yield by 18%. A single stool sample is sufficient for complete parasitological diagnosis when an algorithm based on clinical information is applied.

Association between group A beta-haemolytic streptococci and vulvovaginitis in adult women: a case-control study.

Guidelines for the management of vaginal discharge mention Candida albicans, Trichomonas vaginalis, bacterial vaginosis, Chlamydia trachomatis and Neisseria gonorrhoeae as causes and do not recommend full microbiological culture. The role of non-group B beta-haemolytic streptococci in vaginal cultures is unclear, except for group A streptococci that are known to cause vulvovaginitis in children. In a case-control study, we investigated the association between non-group B beta-haemolytic streptococci and vulvovaginitis in adult women. Cases were women with recurrent vaginal discharge from whom a sample was cultured. Controls were asymptomatic women who consented to submitting a vaginal swab. Group A streptococci were isolated from 49 (4.9%) of 1,010 cases and not from the 206 controls (P < 0.01). Isolation rates of group C, F and G streptococci were low and did not differ statistically between cases and controls. Group A beta-haemolytic streptococci are associated with vaginal discharge in adult women. The other non-group B streptococci require more study. For the adequate management of vaginal discharge, culturing is necessary if initial treatment fails. Guidelines should be amended according to these results.

Device-independent, real-time identification of bacterial pathogens with a metal oxide-based olfactory sensor.

A novel olfactory method for bacterial species identification using an electronic nose device called the MonoNose was developed. Differential speciation of micro-organisms present in primary cultures of clinical samples could be performed by real-time identification of volatile organic compounds (VOCs) produced during microbial replication. Kinetic measurements show that the dynamic changes in headspace gas composition are orders of magnitude larger than the static differences at the end of fermentation. Eleven different, clinically relevant bacterial species were included in this study. For each of the species, two to eight different strains were used to take intra-species biodiversity into account. A total of 52 different strains were measured in an incubator at 37 degrees C. The results show that the diagnostic specificities varied from 100% for Clostridium difficile to 67% for Enterobacter cloacae with an overall average of 87%. Pathogen identification with a MonoNose can be achieved within 6-8 h of inoculation of the culture broths. The diagnostic specificity can be improved by broth modification to improve the VOC production of the pathogens involved.

[Gastroenteritis caused by Salmonella from pet snakes]. / Gastro-enteritis door salmonella aflkomstig van als huisdier gehouden slangen.

A Salmonella subspecies associated with reptiles (Salmonella enterica subspecies diarizonae) was isolated from the stool of a 19-year-old man with gastroenteritis. The same species was isolated from stool and urine samples taken from terraria found in the home of the patient's parents where snakes were kept. A high percentage of reptiles in the wild and in captivity are asymptomatic carriers of Salmonella species that can be transmitted to humans who come in contact with these animals. Unlike in the United States of America, for example, cases of reptile-associated infections have scarcely been published in the Netherlands and targeted information on the risk of infection is lacking. Because the popularity of exotic pets--and thereby the risk of infection--is increasing in The Netherlands, targeted information for veterinarians, traders and owners of exotic pets is warranted to prevent reptile-associated salmonellosis.

In vivo and in vitro effects of tea extracts on enterotoxigenic Escherichia coli-induced intestinal fluid loss in animal models.

OBJECTIVES: Enterotoxigenic Escherichia coli (ETEC) infection is a major cause of dehydrating diarrhoea in infants and early-weaned piglets living under subhygienic conditions. We studied the effect of different tea types and subfractions on the intestinal fluid and electrolyte losses involved in ETEC diarrhoea. MATERIALS AND METHODS: Jejunal segments of anaesthetised piglets were infected with ETEC or ETEC heat-labile toxin (LT) and subsequently perfused for 8 hours with control or tea solutions containing green or black tea extract (BTE) or 3 different BTE subfractions containing small-size, large-size or no phenolics. Changes in intestinal fluid and electrolyte net absorption were measured. To assess the antisecretory effects of tea, BTE was incubated before or after administration of the secretagogue forskolin in rat jejunal tissue placed in Ussing chambers and Cl- secretion measured as changes in short-circuit current (I(SC)). RESULTS: Enterotoxigenic E. coli infection of piglet jejunal segments significantly reduced net absorption of fluid, Na+ and Cl- and increased net secretion of K+ compared with controls. Perfusion of the ETEC-infected segments with both 3 g/L green tea extract and BTE significantly inhibited these disturbances in fluid and electrolyte balance. The BTE subfraction rich in polymeric phenolics but not the other subfractions improved the fluid and electrolyte balance. Addition of forskolin to rat jejunal tissue induced a significant increase in I(SC). Pretreating but not posttreating the jejunal tissue with BTE inhibited the forskolin-induced increase in I(SC). CONCLUSIONS: Tea may inhibit net fluid and electrolyte losses involved in secretory diarrhoea from ETEC.