The occurrence of pathogenic Escherichia coli in South African wastewater treatment plants as detected by multiplex PCR

The aim of this study was to investigate the use of PCR to detect commensal and diarrhoeagenic Escherichia coli concentrated from water samples using membrane filtration. To achieve this, culture-based and PCR-based methods were compared for the detection of E. coli in raw sewage and primary, secondary and tertiary effluents from 6 wastewater treatment plants around Johannesburg, Gauteng. E. coli was concentrated from the samples using standard filtration techniques with subsequent incubation on E. coli/coliform chromogenic media to determine the E. coli levels. Bacterial DNA was isolated from bacterial colonies trapped on polyethersulphone membranes after filtration using a celite/guanidium thiocyanate method. A single multiplex PCR (m-PCR) assay was used that targeted the mdh, eaeA, stx1, stx2, st, lt, ial and eagg genes associated with diarrhoeagenic E. coli. The mdh gene was detected in all of the samples even if no culturable E. coli was detected. All the diarrhoeagenic E. coli types were detected in one or more of the raw sewage samples from the various plants. EPEC was present in 20% (2/10) of the samples, EHEC in 50% (5/10), ETEC in 80% (8/10), EIEC in 10% (1/10) and EAEC in 90% (9/10) of the samples. In the case of the primary and secondary treatment only ETEC (5/5; 100%) and EAEC (5/5; 100%) were detected in all of the samples. The results demonstrate that molecular techniques such as PCR have the potential to be used for the monitoring of water samples for the presence of pathogenic E. coli, without the need to culture the organisms.Keywords: E. coli, multiplex PCR, wastewater treatment plant effluen

Detection of Vibrio cholerae O1 in animal stools collected in rural areas of the Limpopo Province

Vibrio cholerae (V. cholerae), the causative agent of cholera, has been responsible for various outbreaks worldwide and may be associated with animal faeces. In an attempt to understand the occurrence of this organism in the environment, 230 faecal samples were collected from pigs, chickens, goats, donkeys, cows and pigeons in rural areas of the Limpopo Province. Bacterial DNA was extracted from the faecal samples using a guanidium thiocyanate-based method. The DNA was screened for the presence of the sodB, rfb, FlaE, 16S rRNA and ctxA genes associated with V. cholerae, V. cholerae O1, V. cholerae O139 using 2 multiplex polymerase chain reactions (m-PCR). The V. cholerae sodB gene was detected in 74 of the 230 samples tested. Detection rates for the faecal samples obtained from individual species were as follows: cows (55/74), chickens (8/74), goats (2/74), donkeys (4/74), pigs (3/74) and pigeons (2/74). V. cholerae O1 was detected in (17/74) cow and (3/74) chicken samples, of which (9/17) cow samples and (3/3) chicken samples tested positive for toxigenic V. cholerae O1. The presence of this organism in faecal samples, taken close to water sources used by the villagers, raises the possibility that the causative V. cholerae O1 strain of the most recent outbreak in South Africa was present in the area 6 months prior to the outbreak.Keywords: Vibrio cholerae, PCR, animal faeces, cholera toxi

Cardiac surgery for the cyanotic infant

No Abstrac

Optimisation of methods for the collection and detection of bacterial pathogens from diarrhoeal human faecal samples using a novel stool collection kit

Bacterial pathogens such as Escherichia coli, Salmonella-, Shigella- and Vibrio species are known to be common causative agents for diarrhoeal disease in humans. This study aimed to develop a culture-independent PCR assay for the detection of bacterial pathogens present in human faecal samples collected using a less intrusive faecal collection technique, the Bio-wipe kit. A multiplex-PCR (m-PCR) was optimised targeting the E. coli mdh gene, the Salmonella IpaB gene, the Vibrio sodB gene, and the Ial and IpaH genes present in entero-invasive E. coli and Shigella spp. The influence of the DNA extraction method, and sensitivity and specificity of the m-PCR and the Bio-wipe storage conditions on the detection of the bacterial pathogens was investigated. A guanidium thiocyanate DNA extraction method used with laboratory-prepared spin columns could successfully extract DNA from 93% of the samples analysed. The m-PCR could successfully identify and differentiate between the various pathogens tested and was specific for the selected pathogens. Faecal matter was successfully recovered from used Bio-wipes and the bacterial DNA could be detected from these samples at concentrations of 10 cfu. Bacterial DNA could be recovered from the Bio-wipes 5 to 10 d after use when the Bio-wipes were stored at 30°C and 14 d after usage when stored at ambient temperature. Thus the Bio-wipe kit, along with the m-PCR, can be used for collection and detection of bacterial pathogens during outbreaks and in rural settings.Keywords: Bio-wipe kit, bacterial pathogens, faecal matter, PCR, DNA extractio

Assessing the effectiveness of a biological recovery of nickel for tailing dumps management

The mobilization of nickel from sulphide minerals using sulfuric acid and heterotrophic microorganism (Bacillus subtilis) was independently examined. The influences of parameters such as the concentration of acid and bacteria as well as reaction time were considered. Results of the monod-type kinetic study showed faster recovery of nickel from tailings (20 ppm/h) than from ore (8.07 ppm/h) by biological mobilization and similar trend with sulfuric acid

Free-living amoebae isolated from a hospital water system in South Africa: A potential source of nosocomial and occupational infection

Abstract: This study investigated the occurrence of free-living amoebae (FLA) in a public hospital in South Africa. A total of 97 water and biofilm samples from the municipal water inlet of the hospital, theatres, theatre sterilization service unit, central sterilization service unit, endoscopy/gastroscopy unit, intensive care unit and the renal unit were collected and examined for the presence of FLA using an amoebal co-culture and molecular techniques. Of the 97 samples, 77 (79.4 %), 40 (52%) water and 37 (48.1%) biofilm, contained FLA. The genera Acanthamoeba, Vermamoeba (formerly Hartmanella) and Naegleria were detected by morphology, 18S rRNA PCR and sequence analyses. Further sequence analysis of the five Acanthamoeba positive isolates revealed a close resemblance with the potentially pathogenic T20 genotype. These results show a potential health risk to immuno-compromised patients and health care workers as some of the species detected are pathogenic and may harbour potential intracellular bacteria responsible for nosocomial infections. To date, this is the first report on the detection of potentially pathogenic amoebae from South African hospital water systems

Applicability of Bio-wipes for the collection of human faecal specimens for detection and characterisation of enteric viruses

OBJECTIVE : To determine whether gastroenteritis viruses and other enteric viruses could be detected in faecal specimens collected with Bio-wipes. METHODS : Faecal specimens, self-collected with Bio-wipes, from 190 individuals (94 diarrhoeal, 93 non-diarrhoeal, 3 unknown) were screened for eight human enteric viruses (enterovirus, hepatitis A virus, adenovirus, astrovirus, norovirus GI and GII, sapovirus and rotavirus) by real-time (reverse transcription)-polymerase chain reaction. Rotaviruses and noroviruses from positive specimens were genotyped. results At least one enteric virus could be detected in 82.6% (157/190) of faecal specimens. Mixed infections of up to four different viruses could be detected in both diarrhoeal and non-diarrhoeal specimens. Enteroviruses were detected most frequently (63.7%), followed by adenoviruses (48.4%) and noroviruses (32.2%). Genotyping was successful for 78.6% of rotaviruses and 44.8% of noroviruses. CONCLUSIONS : Bio-wipes provide a user friendly, easier method for stool collection that facilitates enteric virus detection and genetic characterisation.http://onlinelibrary.wiley.com/journal/10.1111/(ISSN)1365-3156hb201

Modelling thirty-day mortality in the acute respiratory distress syndrome (ARDS) in an adult ICU

Publisher's copy made available with the permission of the publisher © Australian Society of AnaesthetistsVariables predicting thirty-day outcome from Acute Respiratory Distress Syndrome (ARDS) were analysed using Cox regression structured for time-varying covariates. Over a three-year period, 1996-1998, consecutive patients with ARDS (bilateral chest X-ray opacities, PaO₂/FiO₂ ratio of <200 and an acute precipitating event) were identified using a prospective computerized data base in a university teaching hospital ICU. The cohort, 106 mechanically ventilated patients, was of mean (SD) age 63.5 (15.5) years and 37% were female. Primary lung injury occurred in 45% and 24% were postoperative. ICU-admission day APACHE II score was 25 (8); ARDS onset time from ICU admission was 1 day (median: range 0-16) and 30 day mortality was 41% (95% CI: 33%-51%). At ARDS onset, PaO₂/FiO₂ ratio was 92 (31), 81% had four-quadrant chest X-ray opacification and lung injury score was 2.75 (0.45). Average mechanical ventilator tidal volume was 10.3 ml/ predicted kg weight. Cox model mortality predictors (hazard ratio, 95% CI) were: APACHE II score, 1.15 (1.09-1.21); ARDS lag time (days), 0.72 (0.58-0.89); direct versus indirect injury, 2.89 (1.45-5.76); PaO₂/FiO₂ ratio, 0.98 (0.97-0.99); operative versus non-operative category, 0.24 (0.09-0.63). Time-varying effects were evident for PaO₂/FiO₂ ratio, operative versus non-operative category and ventilator tidal volume assessed as a categorical predictor with a cut-point of 8 ml/kg predicted weight (mean tidal volumes, 7.1 (1.9) vs 10.7 (1.6) ml/kg predicted weight). Thirty-day survival was improved for patients ventilated with lower tidal volumes. Survival predictors in ARDS were multifactorial and related to patient-injury-time interaction and level of mechanical ventilator tidal volume.J. L. Moran, P. J. Solomon, V. Fox, M. Salagaras, P. J. Williams, K. Quinlan, A. D. Berstenhttp://www.aaic.net.au/Article.asp?D=200332

Energetic particle influence on the Earth's atmosphere

This manuscript gives an up-to-date and comprehensive overview of the effects of energetic particle precipitation (EPP) onto the whole atmosphere, from the lower thermosphere/mesosphere through the stratosphere and troposphere, to the surface. The paper summarizes the different sources and energies of particles, principally galactic cosmic rays (GCRs), solar energetic particles (SEPs) and energetic electron precipitation (EEP). All the proposed mechanisms by which EPP can affect the atmosphere are discussed, including chemical changes in the upper atmosphere and lower thermosphere, chemistry-dynamics feedbacks, the global electric circuit and cloud formation. The role of energetic particles in Earth’s atmosphere is a multi-disciplinary problem that requires expertise from a range of scientific backgrounds. To assist with this synergy, summary tables are provided, which are intended to evaluate the level of current knowledge of the effects of energetic particles on processes in the entire atmosphere

Tocilizumab in patients admitted to hospital with COVID-19 (RECOVERY): a randomised, controlled, open-label, platform trial

Background: In this study, we aimed to evaluate the effects of tocilizumab in adult patients admitted to hospital with COVID-19 with both hypoxia and systemic inflammation. Methods: This randomised, controlled, open-label, platform trial (Randomised Evaluation of COVID-19 Therapy [RECOVERY]), is assessing several possible treatments in patients hospitalised with COVID-19 in the UK. Those trial participants with hypoxia (oxygen saturation &lt;92% on air or requiring oxygen therapy) and evidence of systemic inflammation (C-reactive protein ≥75 mg/L) were eligible for random assignment in a 1:1 ratio to usual standard of care alone versus usual standard of care plus tocilizumab at a dose of 400 mg–800 mg (depending on weight) given intravenously. A second dose could be given 12–24 h later if the patient's condition had not improved. The primary outcome was 28-day mortality, assessed in the intention-to-treat population. The trial is registered with ISRCTN (50189673) and ClinicalTrials.gov (NCT04381936). Findings: Between April 23, 2020, and Jan 24, 2021, 4116 adults of 21 550 patients enrolled into the RECOVERY trial were included in the assessment of tocilizumab, including 3385 (82%) patients receiving systemic corticosteroids. Overall, 621 (31%) of the 2022 patients allocated tocilizumab and 729 (35%) of the 2094 patients allocated to usual care died within 28 days (rate ratio 0·85; 95% CI 0·76–0·94; p=0·0028). Consistent results were seen in all prespecified subgroups of patients, including those receiving systemic corticosteroids. Patients allocated to tocilizumab were more likely to be discharged from hospital within 28 days (57% vs 50%; rate ratio 1·22; 1·12–1·33; p&lt;0·0001). Among those not receiving invasive mechanical ventilation at baseline, patients allocated tocilizumab were less likely to reach the composite endpoint of invasive mechanical ventilation or death (35% vs 42%; risk ratio 0·84; 95% CI 0·77–0·92; p&lt;0·0001). Interpretation: In hospitalised COVID-19 patients with hypoxia and systemic inflammation, tocilizumab improved survival and other clinical outcomes. These benefits were seen regardless of the amount of respiratory support and were additional to the benefits of systemic corticosteroids. Funding: UK Research and Innovation (Medical Research Council) and National Institute of Health Research