Isolation and characterization of Clostridium difficile from a small survey of wastewater, food and animals in New Zealand.

The objective of this study was to undertake a microbiological survey of foods, animal faeces and wastewater samples for Clostridium difficile, and determine the genotypes and antimicrobial susceptibilities of isolates. A total of 211 samples were tested for C. difficile using culture methods. Thirteen toxigenic C. difficile isolates were obtained; ten from wastewater samples, one each from pig and duck faeces and another from a raw meat product. Eight PCR-ribotypes (RTs) were identified, including two novel RTs (878 and 879). Single-nucleotide polymorphism analysis using WGS data for all isolates provided greater discrimination between C. difficile isolates within the same RT and multilocus sequence typing (MLST) profiles. All C. difficile isolates were found to be susceptible to the first-line human antimicrobials used to treat C. difficile infection. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study to report the isolation of Clostridium difficile from animals, food and wastewater in New Zealand (NZ) and provides important data with respect to ribotypes and multilocus sequence typing profiles, whole genome sequence and antimicrobial susceptibilities. The results highlight the need for further investigations into the epidemiology of C. difficile in NZ and to elucidate the role of the environmental and food sources as transmission routes of human infection.

A Qualitative Approach in Understanding Illness Perception and Treatment Needs in Patients with Gamma Hydroxybutyrate Use Disorder.

BACKGROUND: The party drug gamma hydroxybutyrate (GHB) is highly addictive. GHB use disorder (GUD) has poor treatment outcome, with relapse rates over 60% within 3 months after detoxification. In order to get a better understanding of the limited treatment success, we explored GUD patients' illness perceptions and treatment needs. METHODS: In a qualitative cross-sectional observational study, using a semi-structured interview based on the works of Kleinmann, illness perceptions were explored among treatment seeking GUD patients (n = 20). The analysis was based on the principles of Grounded Theory by the 2 interviewers and an independent researcher. RESULTS: GUD patients had mainly positive views toward GHB. GHB was perceived as strongly rewarding and perceived as the solution to psychosocial problems, rather than the cause. After repeated readmissions, GUD patients perceived themselves as addicted to GHB and GHB use as more problematic. They reported a need for personalized treatment goals, which were mainly aimed toward dealing with psychiatric symptoms and social reintegration. CONCLUSION: GUD shares many characteristics with other substance use disorders, in line with gradual development from positive reinforcement in early-stage GUD to negative reinforcement in later stages of more compulsive GHB use. Future studies should investigate whether personalization of treatment goals, such as social reintegration, lead to better treatment outcomes.

Isolation and genotyping of Campylobacter species from kiwi (Apteryx spp.) in captivity: implications for transmission to and from humans.

AIMS: To investigate the presence of Campylobacter spp. in captive kiwi (Apteryx spp.) and compare their genotypic profiles with those of human and animal origin, in order to assess their potential for zoonotic or zooanthroponotic transmission. METHODS: Conventional selective enrichment and filter-based isolation methods were applied to isolate Campylobacter spp. from fresh faecal samples from 12 North Island brown kiwi (Apteryx mantelli) and one great spotted kiwi (A. haastii), housed in one of five different areas in a kiwi sanctuary in Christchurch, New Zealand. Isolates were identified using multiplex PCR and 16S rRNA gene sequencing. High-resolution rapid genotyping using multiplex ligation-dependant probe amplification-based binary typing (MBiT) was applied and profiles compared with similar results from 2,165 Campylobacter spp. isolates contained in a database derived from human clinical, veterinary and environmental samples. RESULTS: One isolate of C. jejuni, and one belonging to the C. lari phylogenetic group were recovered from faeces from two kiwi. High-resolution rapid genotyping by MBiT demonstrated these to be indistinguishable from isolates obtained previously from human cases of diarrhoea, and others from chicken, cattle, sheep and water. CONCLUSIONS: These data provide evidence for potential zoonotic or zooanthroponotic transmission of Campylobacter spp. in kiwi with implications for management of birds kept in captivity. We believe this is the first formal report of C. jejuni and a C. lari-like organism in kiwi. ABBREVIATIONS: MBiT: Multiplex ligation-dependant probe amplification-based binary typing.

The Neurobiological Mechanisms of Gamma-Hydroxybutyrate Dependence and Withdrawal and Their Clinical Relevance: A Review.

OBJECTIVE: x03B3;-Hydroxybutyrate (GHB) has gained popularity as a drug of abuse. In the Netherlands the number of patients in treatment for GHB dependence has increased sharply. Clinical presentation of GHB withdrawal can be life threatening. We aim, through this overview, to explore the neurobiological pathways causing GHB dependency and withdrawal, and their implications for treatment choices. METHODS: In this work we review the literature discussing the findings from animal models to clinical studies focused on the neurobiological pathways of endogenous but mainly exogenous GHB. RESULTS: Chronic abuse of GHB exerts multifarious neurotransmitter and neuromodulator effects on x03B3;-aminobutyric acid (GABA), glutamate, dopamine, serotonin, norepinephrine and cholinergic systems. Moreover, important effects on neurosteroidogenesis and oxytocin release are wielded. GHB acts mainly via a bidirectional effect on GABAB receptors (GABABR; subunits GABAB1 and GABAB2), depending on the subunit of the GIRK (G-protein-dependent ion inwardly rectifying potassium) channel involved, and an indirect effect of the cortical and limbic inputs outside the nucleus accumbens. GHB also activates a specific GHB receptor and ß1-subunits of α4-GABAAR. Reversing this complex interaction of neurobiological mechanisms by the abrupt cessation of GHB use results in a withdrawal syndrome with a diversity of symptoms of different intensity, depending on the pattern of GHB abuse. CONCLUSION: The GHB withdrawal symptoms cannot be related to a single mechanism or neurological pathway, which implies that different medication combinations are needed for treatment. A single drug class, such as benzodiazepines, gabapentin or antipsychotics, is unlikely to be sufficient to avoid life-threatening complications. Detoxification by means of titration and tapering of pharmaceutical GHB can be considered as a promising treatment that could make polypharmacy redundant.

Use of a bacteriophage to inactivate Escherichia coli O157:H7 on beef.

A number of outbreaks of Escherichia coli O157:H7 infections involving beef have been reported. Options for controlling bacterial pathogens in raw foods are limited, but one is to use bacteriophages (phages). We describe the isolation and characterisation of phage FAHEc1, which infects E. coli O157, and its ability to kill its host in vitro and on beef. The phage belonged to the family Myoviridae and lysed 28 of 30 E. coli O157 (:H7, :HNM and :H not specified) isolates, only one other non-O157 E. coli serotype (O162:H7), and none of the other 13 bacterial species tested. The phage did not contain stx1, stx2, eae or ehxA virulence genes as assessed by PCR. An approximate 4 log10 inactivation of E. coli O157:H7 occurred at 5 °C in the presence of phage FAHEc1 at >107 PFU/ml in broth in vitro. On thinly sliced beef pieces incubated at 37 °C, a > 2.7 log10 reduction occurred with 3.2 × 107 PFU/4 cm² meat piece. At lower phage concentrations (10³-104 PFU/4 cm² piece) phage replication occurred on beef at 37 °C. When the phage was applied to beef pieces under conditions simulating hot boning and conventional carcass cooling, inactivation of E. coli O157:H7 of approximately 2 log10 was measured under optimal conditions with phages applied at 3.2 × 107 PFU/4 cm² meat piece.

Detection, isolation and enumeration of Yersinia enterocolitica from raw pork.

The methods available for the isolation of Yersinia enterocolitica from foods are generally considered to be less than optimal, and methods for estimation of numbers are lacking. Such methods are needed to understand better the significance of foodborne yersiniosis and to provide data for exposure assessment. We describe a method for the detection and enumeration of Y. enterocolitica containing the pYV virulence plasmid (YeP+) in samples from pork surfaces. The method uses a multiplex PCR targeting the ail and virF genes to detect Y. enterocolitica after incubation of surface swabs in Yersinia enrichment broth according to Ossmer. Enumeration was achieved by adapting the enrichment to a most probable number (MPN) method format. A presumptive result was available within 24 h of sample receipt, and YeP+ isolates were confirmed within four days. The presence/absence and MPN methods were evaluated in a pilot survey of 34 packs of raw pork meat purchased from retail outlets in Christchurch, New Zealand. YeP+ was detected by PCR on meat from 32% of the packs, and YeP+ isolates were obtained from 18% of the samples. YeP+ were present at numbers ranging from 0.30 to 5.42 MPN/cm(2). This improved method for the detection and enumeration of YeP+ from meat samples can be used for microbiological surveys to obtain data for assessments of consumer exposure to virulent Y. enterocolitica, and in outbreak investigations.

Enumeration and growth of naturally occurring Listeria spp. in unpackaged ham.

A total of 301 unpackaged retail ham samples were tested for the presence and number of Listeria spp. after 7 days at 5 degrees C to simulate domestic storage. Thirteen samples (4.3%) contained Listeria monocytogenes, with the highest count being 1.6 x 10(3)cfu g(-1). Thirteen samples contained other Listeria spp. Genotyping showed that only one L. monocytogenes isolate from the 14 tested was of a type previously identified in New Zealand human cases. Listeria-contaminated batches were incubated at 5 degrees C over approximately 3 weeks to assess the growth rate of natural contaminants. None contained L. monocytogenes, but growth occurred in one sample containing Listeria welshimeri and four containing Listeria innocua. Growth was usually slow at 0.002-0.004 log h(-1). In one sample, L. innocua grew at 0.02 log h(-1) although the maximum number reached was only 4.0-5.0 x 10(3)cfu g(-1). In five other samples little growth, if any, occurred. Growth of naturally occurring Listeria spp. at 5 degrees C was therefore generally slower than predicted by the Pathogen Modelling Programme (PMP) or did not occur.

Campylobacter spp. in New Zealand raw sheep liver and human campylobacteriosis cases.

Sheep liver samples were tested for the presence and numbers of Campylobacter jejuni and C. coli during both spring and autumn. Over the same period, isolates were obtained from human clinical cases from the same geographical area as where the food samples were purchased. A subset of the C. jejuni isolates was typed by both Penner serotyping and pulsed field gel electrophoresis using the restriction enzyme SmaI, to estimate the proportion of liver isolate types that were also isolated from human cases of campylobacteriosis. Of the 272 liver samples tested, 180 (66.2%) contained Campylobacter. Most of the positive samples contained <3 MPN/g of the organism, and only 12 (6.7%) were contaminated at a level exceeding 100 MPN/g. A total of 180 C. jejuni isolates were obtained from sheep liver and another 200 from human faeces. Of these, 212 isolates were randomly selected for typing, half from raw liver and half from human faeces. More than half (61.1%) of the 106 C. jejuni isolates from liver were of subtypes that were also isolated from human cases. While the C. jejuni present in sheep liver were mostly of subtypes also isolated from human cases, the significance of this food as a vehicle of human campylobacteriosis needs to be examined further in respect to other factors such as dose-response information, consumption data, frequency of undercooking and cross contamination.