Epidemiological typing of Neisseria gonorrhoeae and detection of markers associated with antimicrobial resistance directly from urine samples using next generation sequencing.

OBJECTIVES: To investigate the potential for next generation sequencing (NGS) to be used directly on clinical specimens that have tested positive for Neisseria gonorrhoeae by nucleic acid amplification testing (NAAT), to generate information on epidemiological genotyping and antimicrobial resistance (AMR) markers. METHODS: DNA was extracted from 13 N. gonorrhoeae NAAT-positive urine specimens, enriched for microbial DNA and sequenced using the Ion Torrent PGM workflow. Sequences that aligned to the human genome were filtered out and the remaining sequences were de novo assembled. The resulting contigs were searched for regions of interest using Ridom SeqSphere. MLST and NG-MAST alleles were assigned according to the schemes at PubMLST.org and NG-MAST.net, respectively. RESULTS: In total, 11 of the 13 samples tested generated a sufficient number of N. gonorrhoeae sequence reads to provide full coverage of the genome at a depth of 6-130×. Complete MLST and NG-MAST sequence types could be generated for each of these samples. The presence of 10 different AMR markers was investigated, and both previously reported and novel mutations were identified in genes associated with reduced susceptibility to several antimicrobials. CONCLUSIONS: We found that sequencing the entire genome of N. gonorrhoeae directly from clinical samples is possible using NGS, and that multiple levels of N. gonorrhoeae typing information can be generated. As NAAT only testing becomes more common, this method could be used to detect both known and novel mutations associated with AMR and to generate genotyping information, supporting AMR and epidemiological surveillance in the absence of culturing.

Azithromycin suppresses P. gingivalis LPS-induced pro-inflammatory cytokine and chemokine production by human gingival fibroblasts in vitro.

OBJECTIVE: Azithromycin is a macrolide antibiotic that appears to have both antibacterial and anti-inflammatory properties. This study aimed to investigate the effect of azithromycin on the production of pro-inflammatory cytokines and chemokines by human gingival fibroblasts (HGF) in vitro. MATERIALS AND METHODS: The effects of azithromycin (0.1 to 10 µg/mL) on the production of interleukin-6 (IL-6), interleukin-8 (IL-8), monocyte chemoattractant protein 1 (MCP-1), and growth-regulated oncogene (GRO) by human gingival fibroblasts cultured in the presence or absence of Porphyromonas gingivalis lipopolysaccharide (LPS) was studied. Cytokine and chemokine protein levels in the culture supernatant were assessed using a Luminex® multiplex immunoassay. RESULTS: P. gingivalis LPS induced cytokine/chemokine (IL-6, IL-8, MCP-1, and GRO) protein production in HGFs, and this effect was suppressed by azithromycin at all concentrations tested. CONCLUSIONS: This study demonstrates that azithromycin suppresses P. gingivalis LPS-induced cytokine/chemokine protein production in HGF, which may explain some of the clinical benefits observed with the adjunctive use of azithromycin in the treatment of periodontitis. CLINICAL RELEVANCE: The current study examines the anti-inflammatory properties of azithromycin which may make it useful as an adjunct treatment to periodontitis. Specifically, we used azithromycin to modulate the production of pro-inflammatory cytokines by gingival fibroblasts known to be important in periodontal inflammation.

Real-time investigation of a Legionella pneumophila outbreak using whole genome sequencing.

Legionella pneumophila is the main pathogen responsible for outbreaks of Legionnaires' disease, which can be related to contaminated water supplies such as cooling towers or water pipes. We combined conventional molecular methods and whole genome sequence (WGS) analysis to investigate an outbreak of L. pneumophila in a large Australian hospital. Typing of these isolates using sequence-based typing and virulence gene profiling, was unable to discriminate between outbreak and non-outbreak isolates. WGS analysis was performed on isolates during the outbreak, as well as on unlinked isolates from the Public Health Microbiology reference collection. The more powerful resolution provided by analysis of whole genome sequences allowed outbreak isolates to be distinguished from isolates that were temporally and spatially unassociated with the outbreak, demonstrating that this technology can be used in real-time to investigate L. pneumophila outbreaks.

Prolonged and mixed non-O157 Escherichia coli infection in an Australian household.

An Australian family was identified through a Public Health follow up on a Shiga-toxigenic Escherichia coli (STEC) positive bloody diarrhoea case, with three of the four family members experiencing either symptomatic or asymptomatic STEC shedding. Bacterial isolates were submitted to stx sequence sub-typing, multi-locus variable number tandem repeat analysis (MLVA), multi-locus sequence typing (MLST) and binary typing. The analysis revealed that there were multiple strains of STEC being shed by the family members, with similar virulence gene profiles and the same serogroup but differing in their MLVA and MLST profiles. This study illustrates the potentially complicated nature of non-O157 STEC infections and the importance of molecular epidemiology in understanding disease clusters.

Assessment of metal toxicity in sediment pore water from Lake Macquarie, Australia.

Recent investigations into the level of heavy metal enrichment in the sediments of Lake Macquarie have indicated that significant contamination has occurred over the past 100 years, with elevated levels of lead, zinc, cadmium, copper, and selenium being observed in most parts of the lake. Pore water extracted from sediments showing the greatest contamination by these metals exhibited toxicity to the larval development of the sea urchin Heliocidaris tuberculata. However, an analysis of pore water metal concentrations revealed that the concentrations of these metals were too low to cause toxicity. Rather, pore water toxicity was highly correlated with manganese for the majority of sites sampled; subsequent spiking experiments confirmed manganese as a cause of toxicity. Current levels of manganese in the sediments of Lake Macquarie have arisen from natural sources and are not the result of anthropogenic activities. These results reiterate the importance of identifying the causes of toxicity in assessments of sediment contamination, particularly when testing sediment pore waters using sensitive early life stages.

Critical incident stress and challenges for the emergency workplace.

A formal response to all levels of critical incidents that are potentially psychologically disturbing is needed. This response is needed for small-scale and large-scale incidents, such as natural or manmade disasters. Health care providers at all levels should be encouraged to participate in wellness programs and to understand workplace stressors. Critical incident stress debriefing teams have a valuable role in preventing posttraumatic stress disorder. Procedures for establishing response teams are presented, and future directions are discussed.

Comparison of the kinetic fibrinogen assay with the von Clauss method and the clot recovery method in plasma of patients with conditions affecting fibrinogen coagulability.

A Kinetic Fibrinogen Assay (KFA), a method based on the kinetic reaction of the developing fibrin clot, was used to determine fibrinogen concentration in plasma. Two other methods employing different quantification principles were used for comparison: the von Clauss method and the procedure measuring protein concentration in an isolated and washed plasma clot (World Health Organization [WHO] method). All three methods quantified functional thrombin-coagulable fibrinogen. Plasma specimens were obtained from three groups of patients: those with liver disease and those taking either coumarin derivative or heparin. In all of these conditions, there are deviations from the normal process of fibrin clot formation. The KFA method yielded results that were consistent and provided excellent precision and accuracy allowing quantification of plasma fibrinogen in the range of 70-800 mg/dL (2-23.5 microM). The determination of fibrinogen by the KFA method was not adversely affected using plasma from patients treated with heparin and those undergoing coumarin therapy. Statistical analysis of the results indicated that the KFA method compared very favorably with the von Clauss and WHO methods. In assessing the clinical utility of each method, the WHO method was found to be labor intensive and time consuming; therefore, not suitable for routine use in a clinical laboratory. The von Clauss method required a trained laboratory technician and some laboratory manipulations. The KFA method was not only reliable and accurate, but also fully automated, making it the easiest and the fastest to perform routinely.

Thrombophlebitic retroperitoneal collateral veins mimicking lymphadenopathy: MR and CT appearance.

The authors report the magnetic resonance (MR) and computed tomography (CT) features of what is believed to be thrombophlebitis of the left cava of a duplicated IVC, which mimicked lymphadenopathy, and prompted both a CT-guided needle aspiration biopsy and surgical exploration. Knowledge that retroperitoneal vascular anomalies can mimick lymphadenopathy, both clinically and radiographically, may help obviate further testing and intervention.

Effect of staff turnover and the social environment on depressive symptoms in nursing home residents.

Although one would expect a relationship on the basis of anecdotal evidence, there is little objective evidence for a hypothesized relationship between the social environment provided in institutional settings and the psychological well-being of the residents. In an attempt to operationalize this hypothesis, the present study examined the effect of nursing staff turnover, frequency of visitors, and the presence or absence of close friends on depressive symptoms among nursing home residents. Once physical dependency and past history of depression were taken into account, there was no demonstrable relationship between staff turnover and depressive symptoms among the residents of this sample of nursing homes. There was, however, a significant relationship between absence of close friends among other residents and depressive symptoms, although only 13% of the total variance was explained.

The Canberra Interview for the Elderly: assessment of its validity in the diagnosis of dementia and depression.

The Canberra Interview for the Elderly (CIE) has been developed as a field instrument for identifying cases of dementia and depression, according to the diagnostic criteria in both draft ICD-10 and DSM-III-R. It has been designed to be administered by lay interviewers and responses are assembled algorithmically to derive diagnoses. The validity of the CIE was assessed using a sample of 75 elderly patients attending a hospital clinic. The CIE diagnoses were compared with clinical judgements made at the time of recruitment into the study and later by 3 clinicians using the information collected by the lay interviewers. Agreement between the CIE and the clinicians' diagnoses was as great as agreement between the clinicians themselves, meeting or exceeding agreement observed for comparable instruments designed for lay administration.

Cutaneous ureterostomy in adults.

In the patient with metastatic carcinoma, urinary diversion is usually achieved with indwelling ureteral stents or placement of a percutaneous nephrostomy tube. Most forms of surgical diversion carry an unacceptable morbidity rate, especially in the debilitated patient. Over a fifteen-year period (1974-1989), 29 adult patients with pelvic malignancy (32 ureters) underwent palliative cutaneous ureterostomy. This previously reported technique involves transverse nephropexy and construction of a stoma using a small skin flap. Indications included ureteral obstruction or severe urinary tract symptoms. Hydroureter, often considered a precondition for this procedure, was not present in several patients and was not a prerequisite to success. Complications related to the procedure included one postoperative death due to stroke, one death due to uremia and sepsis, and one instance of severe renal arterial stenosis resulting in renal failure. Preservation of renal function was possible in the 10 patients known to have survived from one to thirteen years postoperatively; only 3 patients eventually required stomal revision. By adherence to the surgical techniques described, the usually high incidence of stomal stenosis was avoided. Our experience reveals that although the indications for cutaneous ureterostomy are limited, this procedure can provide an alternative to permanent nephrostomy drainage or to a higher risk intestinal urinary diversion in carefully selected patients with a reasonable life expectancy.

Assessing the quality of residential care for the aged.

The standard of quality of care in Australian nursing homes is assessed by standards monitoring teams who use their expert judgement and a set of guidelines based on seven objectives and thirty-one standards. In this article, the possibility of supplementing the teams' expert judgement with information gained from standardised questionnaires is explored. Following a review of environment-focussed and resident-focussed instruments, an overview of the scales is provided and the idea of using standardised questionnaires in the standards monitoring process is discussed in terms of possible future developments.

Risk Assessment Studies: Detailed Host Range Testing of Wild-Type Cabbage Moth, Mamestra brassicae (Lepidoptera: Noctuidae), Nuclear Polyhedrosis Virus.

The host range of a multiply enveloped nuclear polyhedrosis virus (NPV) (Baculoviridae) isolated from the cabbage moth, Mamestra brassicae (Lepidoptera: Noctuidae), was determined by challenging a wide range of insect species with high (10 polyhedral inclusion bodies) and low (10 polyhedral inclusion bodies) doses of the virus. The identity of the progeny virus was confirmed by dot blotting. Analysis of 50% lethal dose was carried out on selected species, and the progeny virus was identified by using restriction enzyme analysis and Southern blotting. Other than the Lepidoptera, none of the species tested was susceptible to M. brassicae NPV. Within the Lepidoptera, M. brassicae NPV was infective to members of four families (Noctuidae, Geometridae, Yponomeutidae, and Nymphalidae). Of 66 lepidopterous species tested, M. brassicae NPV was cross-infective to 32 of them; however, 91% of the susceptible species were in the Noctuidae. The relevance of host range data in risk assessment studies is discussed.

Mass casualty incident. Integration with prehospital care.

Mass casualty incident involves the use of limited resources for multiple casualties. The emergency physician must be familiar with both prehospital and hospital plans for mass casualty care in order to facilitate optimal care and to maintain the continuum from field care to definitive treatment. It is essential that the emergency physician become involved in the disaster planning processes to ensure that the victims receive the best care available under the circumstances and that the safety and emotional well-being of both prehospital and hospital personnel are assured. Emergency physicians involved in prehospital care should be certain that the local EMS system has adequate training and chances to update their skills and knowledge. Disaster drills of the EMS system are excellent ways to practice, to identify weaknesses, and for preplanning to enhance disaster medical care.

Retroperitoneal actinomycosis: a case report and review of the literature.

A 25-year-old Haitian man presented with a post-traumatic left retroperitoneal abscess. Culture of material obtained during surgical drainage yielded Actinomyces israelii. Retroperitoneal actinomycosis is a particularly unusual manifestation of infection with this organism. Because it may mimic subacute infections or malignant masses in terms of clinical findings, diagnosis of this entity may be difficult. Our diagnosis was based on the results of culture, and the patient responded to long-term penicillin therapy.

Family participation during resuscitation: an option.

We began to question the fairness of a policy to exclude close family members from the treatment room during attempted resuscitation of cardiac arrest victims in 1982 after 13 of 18 surviving relatives (72%) who were surveyed about their experiences during the attempted resuscitation of a family member responded that they would have liked to have been present during the resuscitation. We report the results of a program instituted at that time that allowed selected family members to be present during resuscitation efforts. Family members were asked by a chaplain or nurse if they wished to be present in the resuscitation room, and those accepting were accompanied by a supporting emergency staff member who explained the milieu of the code room. None of the participants interfered with resuscitation efforts. Seventy persons who participated were later contacted by one of the chaplains and asked to complete a survey form. Forty-four of 47 respondents (94%) who had been present during resuscitation believed that they would participate again. Thirty-six (76%) thought that adjustment to the death or grieving was facilitated by their witnessing the resuscitation; 30 (64%) felt that their presence was beneficial to the dying family member. We conclude that lay person may wish to be with family members who may be dying even though resuscitation efforts are being made, and that it is reasonable to inquire about this wish. This experience has assisted the grieving process for many and has not interrupted or adversely affected medical efforts at resuscitation.

Hazardous chemicals.

Resuscitation of the patient who has been exposed to a toxic substance requires attention to good supportive care, attention to the safety of the rescuers and emergency department personnel, prompt and thorough decontamination, and use of specific antidotes when available.