Phenazine-1 carboxylic acid of Pseudomonas aeruginosa induces the expression of Staphylococcus aureus Tet38 MDR efflux pump and mediates resistance to phenazines and antibiotics.

In this study, we showed that phenazine-1 carboxylic acid (PCA) of Pseudomonas aeruginosa induced the expression of Tet38 efflux pump triggering Staphylococcus aureus resistance to tetracycline and phenazines. Exposure of S. aureus RN6390 to supernatants of P. aeruginosa PA14 and its pyocyanin (PYO)-deficient mutants showed that P. aeruginosa non-PYO phenazines could induce the expression of Tet38 efflux pump. Direct exposure of RN6390 to PCA compound at 0.25× MIC led to a five-fold increase in tet38 transcripts. Expression of Tet38 protein was identified through confocal microscopy using RN6390(pRN-tet38p-yfp) that expressed YFP under control of the tet38 promoter by PCA at 0.25× MIC. The MICs of PCA of a Tet38-overexpressor and a Δtet38 mutant showed a three-fold increase and a two-fold decrease, respectively, compared with that of wild-type. Pre-exposure of RN6390 to PCA (0.25× MIC) for 1 hour prior to addition of tetracycline (1× or 10× MIC) improved bacteria viability of 1.5-fold and 2.6-fold, respectively, but addition of NaCl 7% together with tetracycline at 10× MIC reduced the number of viable PCA-exposed RN6390 of a 2.0-log10 CFU/mL. The transcript levels of tetR21, a repressor of tet38, decreased and increased two-fold in the presence of PCA and NaCl, respectively, suggesting that the effects of PCA and NaCl on tet38 production occurred through TetR21 expression. These data suggest that PCA-induced Tet38 protects S. aureus against tetracycline during coinfection with P. aeruginosa; however, induced tet38-mediated S. aureus resistance to tetracycline is reversed by NaCl 7%, a nebulized treatment used to enhance sputum mobilization in CF patients.

NorA efflux pump mediates Staphylococcus aureus response to Pseudomonas aeruginosa pyocyanin toxicity.

Endogenous transporters protect Staphylococcus aureus against antibiotics and also contribute to bacterial defense from environmental toxins. We evaluated the effect of overexpression of four efflux pumps, NorA, NorB, NorC, and Tet38, on S. aureus survival following exposure to pyocyanin (PYO) of Pseudomonas aeruginosa, using a well diffusion assay. We measured the PYO-created inhibition zone and found that only an overexpression of NorA reduced S. aureus susceptibility to pyocyanin killing. The MICPYO of the NorA overexpressor increased threefold compared to that of wild-type RN6390 and was reduced 2.5-fold with reserpine, suggesting that increased NorA efflux caused PYO resistance. The PYO-created inhibition zone of a ΔnorA mutant was consistently larger than that of a plasmid-borne NorA overexpressor. PYO also produced a modest increase in norA expression (1.8-fold at 0.25 µg/mL PYO) that gradually decreased with increasing PYO concentrations. Well diffusion assays carried out using P. aeruginosa showed that ΔnorA mutant was less susceptible to killing by PYO-deficient mutants PA14phzM and PA14phzS than to killing by PA14. NorA overexpression led to reduced killing by all tested P. aeruginosa. We evaluated the NorA-PYO interaction using a collection of 22 clinical isolates from adult and pediatric cystic fibrosis (CF) patients, which included both S. aureus (CF-SA) and P. aeruginosa (CF-PA). We found that when isolated alone, CF-PA and CF-SA expressed varying levels of PYO and norA transcripts, but all four CF-PA/CF-SA pairs isolated concurrently from CF patients produced a low level of PYO and low norA transcript levels, respectively, suggesting a partial adaptation of the two bacteria in circumstances of persistent co-colonization.

Smoking in pregnancy is a key factor for sudden infant death among Maori.

AIM: To examine the sudden unexpected death in infancy (SUDI) disparity between Maori and non-Maori in New Zealand. METHODS: A nationwide prospective case-control study ran from March 2012 to February 2015. Exposure to established SUDI risk factors was analysed to investigate the disparity experienced by Maori. Infant ethnicity was based on mother's ethnicity. Maori ethnicity was prioritised. Non-Maori includes Pacific, Asian, NZ European and Other. RESULTS: There were 137 cases and 649 controls. The Maori SUDI rate was 1.41/1000 live births compared to 0.53/1000 for non-Maori. Parents/caregivers of 132 cases (96%) and 258 controls (40%) were interviewed. Smoking in pregnancy was associated with an equally increased SUDI risk for Maori (adjusted OR = 8.11, 95% CI = 2.64, 24.93) and non-Maori (aOR = 5.09, 95% CI = 1.79, 14.47), as was bed-sharing (aOR = 3.66, 95% CI = 1.49, 9.00 vs aOR = 11.20, 95% CI = 3.46, 36.29). Bed-sharing prevalence was similar; however, more Maori controls smoked during pregnancy (46.7%) than non-Maori (22.8%). The main contributor relating to increased SUDI risk for Maori/non-Maori infants is the combination of smoking in pregnancy and bed sharing. CONCLUSION: The association between known SUDI risk factors, including bed sharing and/or smoking in pregnancy and SUDI risk, is the same regardless of ethnicity. Maori infants are exposed more frequently to both behaviours because of the higher Maori smoking rate.

Role of L-type calcium channels in altered microvascular responses to propofol in hypertension.

BACKGROUND: Propofol acts as an L-type calcium channel (LTCC) antagonist to decrease peripheral resistance and initiate hypotension. This study investigated LTCC sensitivity/expression in hypertension and the role of LTCCs in exaggerated hypotension to propofol in this situation. METHODS: Age-matched 12- to 15-week-old normotensive rats [male Wistar Kyoto (WKY)] and spontaneously hypertensive rats (SHR) were used. Propofol (10 mg kg(-1), 10-50 mg kg(-1) h(-1) i.v.) was administered and the mesenteric microcirculation (<70 µm) observed with fluorescent in vivo microscopy using fluorescein isothiocyanate-conjugated bovine serum albumin (100 mg kg(-1) i.v.). Western blotting was used to measure tissue expression of the α(1C) LTCC subtype. Pressure myography was used to assess isolated mesenteric arterioles (<350 µm) in response to BAYK8644 (0.1 nM-1 µM), a specific LTCC channel agonist. RESULTS: Propofol dilated isolated arterioles {336.6 µM [mean (sd) change 16.2 (5.8)%]}. However, constriction to BAYK8644 was reduced at this concentration of propofol [EC(50)=8.3 (0.1) log mol(-1)] compared with controls [7.4 (0.1) log mol(-1), P<0.05], suggesting that propofol inhibited LTCCs. The sensitivity of LTCCs increased during hypertension, as in vivo there was a greater increase in mean arterial pressure (MAP) to BAYK8644 [10 µg kg(-1), WKY: 59.5 (9.3)%; SHR: 97.7 (6.3)%, P<0.05] with exaggerated constriction of arterioles [10 µg kg(-1), WKY: 9.1 (2.5)%; SHR: 19.1 (2.6)%, P<0.05]. Propofol also decreased MAP in SHR over time (P<0.05), but remained unchanged in WKY. Using western blotting, expression of α(1C) was greater in SHR compared with WKY (P<0.05). CONCLUSIONS: Propofol acts via LTCC channels, with increased channel expression and sensitivity in genetically hypertensive rats. We suggest that increased sensitivity and expression of LTCCs may be a mechanism for exaggerated hypertension during propofol anaesthesia.

Translation of flecainide- and mexiletine-induced cardiac sodium channel inhibition and ventricular conduction slowing from nonclinical models to clinical.

INTRODUCTION: Nonclinical in vivo models used for cardiovascular safety testing have not previously been studied for their sensitivity for detection of conduction slowing resulting from cardiac sodium channel block. The goal of this study was to examine the sensitivity of in vivo models to cardiac sodium channel block, and translation of the effect from in vitro to in vivo models using sodium channel inhibitors flecainide and mexiletine; flecainide, but not mexiletine is commonly associated with QRS complex prolongation in humans. METHODS: Inhibition of cloned cardiac sodium channels (hNav1.5) was studied using the IonWorks platform. Conduction slowing was measured in vitro in the rabbit isolated ventricular wedge (RVW) and in vivo in the conscious telemetered rat and dog, and anaesthetised dog. RESULTS: Flecainide and mexiletine inhibited hNav1.5 channels with IC50 values of 10.7 and 67.2 µM respectively. In the RVW, QRS was increased by flecainide at 60 bpm, and at 120bpm, there was an increased effect of both drugs. In conscious rats, flecainide significantly increased QRS complex duration; mexiletine had no significant effect, but there was an increase at the highest dose in 4/6 animals. QRS complex was increased by flecainide and mexiletine in anaesthetised dogs but this was not statistically significant; in conscious dog, only flecainide produced a significant increase in QRS complex. DISCUSSION: When compared to clinical data, effects of flecainide and mexiletine in RVW and conscious dog compared well with effects in patients and healthy volunteers in terms of sensitivity. The anaesthetised dog was least sensitive for detection of changes in QRS. All assays showed some differentiation between the expected conduction slowing activity of flecainide and mexiletine. Based on these data, RVW and conscious dog were most predictive for effects of compounds on QRS complex and cardiac conduction.

The menopause symptom profile of Maori and non-Maori women in New Zealand.

OBJECTIVES: To describe menopause symptoms in postmenopausal Maori and non-Maori New Zealand women, and explore relationships between symptoms, sociodemographic profile and hormone replacement therapy (HRT) use. METHODS: Cross-sectional analysis of data collected during recruitment of women from 27 primary-care practices into an observational study and the international WISDOM trial of postmenopausal HRT. RESULTS: A total of 3616 women aged 49-70 years (mean 58.9 years) were recruited between 1999 and 2004. Maori and non-Maori participants differed in demographic and clinical characteristics, but few differences were observed in the frequency of menopause-related symptoms. Vasomotor symptoms (hot flushes and/or night sweats) were reported by 34.4% of women, with no statistically significant difference between Maori and non-Maori (controlling for demographic variables, p > 0.05). Compared to non-Maori, Maori were less likely to have ever used HRT (24% vs. 54%), and only 5% of Maori reported current HRT use compared to 30% of non-Maori (p < 0.05). CONCLUSIONS: Maori and non-Maori have the same rate of self-reported vasomotor symptoms despite differences in factors thought to influence the experience of menopause symptoms. Differences in HRT use between Maori and non-Maori need to be further explored to identify contributing factors and whether, in turn, these factors impact on quality of life.

Physical mapping of the IGF2 locus in the South American opossum Monodelphis domestica.

The South American opossum Monodelphis domestica has been a model organism for marsupials for many years and has recently been the subject of a large-scale genome sequencing effort that will provide the foundation for comparative studies of gene function and regulation. Genomic imprinting is one mechanism of gene regulation that has received increasing attention due to the impact of imprinting defects on development and disease. We have mapped the imprinted insulin-like growth factor II (IGF2) gene of M. domestica as a first step in understanding the regulatory mechanisms involved in genomic imprinting in this marsupial.

The ototoxicity of styrene: a review of occupational investigations.

OBJECTIVES: The objective of this study was to review critically a number of occupational investigations of the exposure and effect relation between inhaled styrene vapour and hearing loss. There is concern that workers' hearing may be impaired by exposure to styrene, as used in industries making plastics and fibreglass-reinforced products. METHODS: Seven occupational studies, each dealing with the ototoxicity of styrene, were examined. Factors assessed included the experimental design and number of subjects within exposure groups, measurement of the styrene-in-air concentration, confirmation of the styrene exposure by blood or urine analysis, determination of the hearing threshold levels for the exposure and control groups, and measurement of any occupational noise in the subjects' workplaces. Consideration was also given to statistical relations between high-frequency hearing loss and lifetime exposure indices for styrene and noise. RESULTS: The results are equivocal. Four investigations failed to find any effect of styrene on hearing thresholds. In contrast, other investigations claimed to have demonstrated styrene-induced hearing loss in industrial populations, with synergism between styrene and noise. However, these reports exhibited shortcomings of experimental design and data analysis. CONCLUSIONS: Considering the body of evidence as a whole, hearing deficits due to occupational exposure to styrene at low concentrations have not been demonstrated by scientifically reliable argument. There is some suggestion of an association between styrene exposure, occupational noise, and hearing dysfunction. Further studies in humans are necessary to clarify this question.

Variation of young normal-hearing thresholds measured using different audiometric earphones: implications for the acoustic coupler and the ear simulator.

This paper questions the necessity for two calibration devices to measure the acoustic output from different types of audiometric earphones. International standards give the audiometric zero for TDH39 earphones on the IEC 60318-3 acoustic coupler; the IEC 60318-1 ear simulator is intended for other supra-aural earphone types. If hearing threshold samples from young, healthy ears were found to be more variable using TDH39 earphones, then that earphone and its coupler might be taken out of service. The audiological literature yielded threshold survey results for over 5100 otologically normal ears of subjects aged 31 years or less. These independent samples showed smaller variation for TDH39 samples than for samples using other earphones; this finding does not support abandoning the TDH39 and its coupler. Nevertheless, benefits accrue from calibrating TDH39 output to the audiometric zero as measured on the ear simulator.

Maori women and menopause.

Maori are the indigenous people of New Zealand who in total make up 14.5% of the population. Although this group has a significantly lower life expectancy than non-Maori, coupled with increased rates of mortality and morbidity, very little is known about the menopausal health needs of older Maori women. As the first step in addressing the health needs of this group, older Maori women's definitions, attitudes, symptoms, expectations and health needs at menopause need to be identified and described. The study Nga Ruahine or "Maori in Menopause" is the foundation study of the Aotearoa Women's Health Initiative (AWHI). AWHI is a women's health programme being developed by the Wellington School of Medicine, which involves a suite of studies. The objective is to describe the journey of older Maori women through menopause and beyond and to compare and contrast the experience of Maori women from both traditional and contemporary upbringings, with reference to the Pakeha (European) population. It is hoped that this work could lead to further studies such as, for example, a longitudinal observational study looking at older New Zealand women. The potential significance of this approach is discussed.

Concept of the notional person in the assessment of hearing disability.

Medicolegal assessment in relation to a claim for hearing damage requires an equitable baseline against which to compare the state of hearing of the claimant. This baseline, or 'notional person', should match the claimant in every respect save for the cause of the hearing loss. The amount of compensation is determined from the difference between the percentage disability of the claimant and that of the 'notional person'. These percentages are, in turn, calculated from the respective hearing threshold levels. In the particular case of occupational noise-induced hearing loss there are, in principle, two ways of proceeding. One way is to estimate directly the noise-induced threshold shift which would result from the noise exposure and to subtract this from the claimant's measured threshold level; in practice the noise exposure is rarely known with the necessary accuracy. The alternative method is to construct the notional person's threshold level by adding together all the non-compensable components of hearing loss known to exist in the claimant. These may include: conductive hearing loss together with any sensorineural sequelae, sensorineural loss specifically attributable to identifiable accident or disease, pure biological ageing, any pathological overlay due to deficiencies of aural or general health, the effect of non-occupational noise, and the socioeconomic status of the claimant. Once these components are summed, whatever remains unaccounted for is deemed to be due to the occupational noise. An International Standard (ISO 7029) exists for the biological ageing component but it is specific to 'otologically normal' persons and on its own does not account for all the time-dependent hearing losses found in a typical claimant. Moreover, the term 'otologically normal', although defined in the relevant Standards, leads to varying interpretations, even to misunderstandings. In practice, the equitable baseline for the disability assessment is most often a 'typical' rather than an 'otologically normal' person in the Standards sense. This paper offers guidance for estimating the combined time-dependent threshold shift in these commonly occurring cases, based on the use of published survey data. It is recommended that a presumption of 'typical hearing' should be made unless the claimant in question can be specifically verified as 'otologically normal'.

Substrate and inhibitor studies with human gastric aspartic proteinases.

The separation of pepsin isoenzymes 1, 2, 3 and 5 (gastricsin) in human gastric juice was effected by chromatography on Mono Q ion-exchanger, and slow-moving proteinase was purified to homogeneity by using a modified procedure incorporating a novel affinity-chromatography step. The pH-activity profiles of these enzymes with mucus glycoprotein and basement-membrane substrates were determined; the profiles for pepsin 2 were noticeably different, and, in general, the pH optima for the hydrolysis of basement membrane were more acidic. Pepsin 1 expressed larger specificity constants (kcat./Km) than pepsin 3 with a series of synthetic peptide substrates, reflecting greater binding (smaller Km) by pepsin 1. Inhibitor studies at pH 1.7 and 4.5 with a series of P2-substituted lactoyl-pepstatins implied that valine at position P2 was optimal for inhibiting pepsins 1, 2 and 3 but detrimental for pepsin 5, whereas lysine at position P2 was tolerated well by pepsin 5 but not by pepsins 1, 2 and 3. The potency of lactoyl-pepstatin with lysine at position P2 did not increase as a function of pH. P2-substituted lactoyl-pepstatins failed to show any inhibitory selectivity among pepsins 1, 2 and 3.

Specification of the geometry of the human ear canal for the prediction of sound-pressure level distribution.

The geometry of 15 human ear canals has been studied. Silicone rubber molds were made of the ear canals of human cadavers, and a mechanical probe system was used to obtain approximately 1000 coordinate points over the surface of each mold. The data points were accurate to about 0.03 mm in each of the three space directions, allowing ample resolution of surface detail. The measurements have been summarized as individual ear canal area functions, the area of cross-sectional slices normal to a curved central axis following the bends of the canal. Large intersubject differences were found, but several overall trends were evident in the area functions. Accurate specification of the canal geometry has lead to improved predictions of the sound-pressure distribution along the human ear canal at frequencies greater than 8 kHz. Such predictions are relevant to the development of high-frequency audiometric methods, high-fidelity hearing aids, and to the interpretation of experiments in physiological and psychological acoustics.

The EAP and workplace psychiatric injury.

The author provides brief historical and current overviews of EAP development, describes such programs' tasks and goals, and discusses how a professionally staffed EAP can address psychiatric injury in the workplace. Based on his experience in a major financial corporation, he describes strategies and interventions used by the EAP he oversees.