Patient-Reported Outcome Measures in Patients with and without Non-Expandable Lung Secondary to Malignant Pleural Effusion-A Single-Centre Observational Study.

BACKGROUND: Malignant pleural effusion (MPE) affects up to 15% of patients with malignancy, and the prevalence is increasing. Non-expandable lung (NEL) complicates MPE in up to 30% of cases. However, it is not known if patients with malignant pleural effusion and NEL are more symptomatic in activities of daily living compared to patients with MPE with expandable lung. METHODS: This was an observational study on consecutively recruited patients with MPE from our pleural clinic. Before thoracentesis, patients completed patient-reported outcomes on cancer symptoms (ESAS), health-related quality of life (5Q-5D-5L), and dyspnoea scores. Following thoracentesis, patients scored dyspnoea relief and symptoms during thoracentesis. Data on focused lung ultrasound and pleural effusion biochemistry were collected. The non-expandable lung diagnosis was made by pleural experts based on radiological and clinical information. RESULTS: We recruited 43 patients, including 12 with NEL (28%). The NEL cohort resembled those from previous studies concerning ultrasonography, pleural fluid biochemistry, and fewer cases with high volume thoracentesis. Patients with and without NEL were comparable concerning baseline demography. The 5Q-5D-5L utility scores were 0.836 (0.691-0.906) and 0.806 (0.409-0.866), respectively, for patients with and without NEL. We observed no between-group differences in symptom burden or health-related quality of life. CONCLUSION: While the presence of NEL affects the clinical management of recurrent MPE, the presence of NEL seems not to affect patients' overall symptom burden in patients with MPE.

Quantification of ß-lactamase producing bacteria in German surface waters with subsequent MALDI-TOF MS-based identification and ß-lactamase activity assay.

Environmental oligotrophic bacteria are suspected to be highly relevant carriers of antimicrobial resistance (AMR). However, there is a lack of validated methods for monitoring in the aquatic environment. Since extended-spectrum ß-lactamases (ESBLs) play a particularly important role in the clinical sector, a culturing method based on R2A-medium spiked with different combinations of ß-lactams was applied to quantify ß-lactamase-producing environmental bacteria from surface waters. In German surface water samples (n = 28), oligotrophic bacteria ranging from 4.0 × 103 to 1.7 × 104 CFU per 100 mL were detected on the nutrient-poor medium spiked with 3rd generation cephalosporins and carbapenems. These numbers were 3 log10 higher compared to ESBL-producing Enterobacteriales of clinical relevance from the same water samples. A MALDI-TOF MS identification of the isolates demonstrated, that the method leads to the isolation of environmentally relevant strains with Pseudomonas, Flavobacterium, and Janthinobacterium being predominant ß-lactam resistant genera. Subsequent micro-dilution antibiotic susceptibility tests (Micronaut-S test) confirmed the expression of ß-lactamases. The qPCR analysis of surface waters DNA extracts showed the presence of ß-lactamase genes (blaTEM, blaCMY-2, blaOXA-48, blaVIM-2, blaSHV, and blaNDM-1) at concentrations of 3.7 (±1.2) to 1.0 (±1.9) log10 gene copies per 100 mL. Overall, the results demonstrate a widespread distribution of cephalosporinase and carbapenemase enzymes in oligotrophic environmental bacteria that have to be considered as a reservoir of ARGs and contribute to the spread of antibiotic resistance.

Ultrasound in the Diagnosis of Non-Expandable Lung: A Prospective Observational Study of M-Mode, B-Mode, and 2D-Shear Wave Elastography.

BACKGROUND: Non-expandable lung (NEL) has severe implications for patient symptoms and impaired lung function, as well as crucial implications for the management of malignant pleural effusion (MPE). Indwelling pleural catheters have shown good symptom relief for patients with NEL; hence, identifying patients early in their disease is vital. With the inability of the lung to achieve pleural apposition following thoracentesis and the formation of a hydropneumothorax, traditionally, chest X-ray and clinical symptoms have been used to make the diagnosis following thoracentesis. It is our aim to investigate whether ultrasound measurement of lung movement during respiration can predict NEL before thoracentesis, thereby aiding clinicians in their planning for the optimal treatment of affected patients. METHODS: A total of 49 patients were consecutively included in a single-centre trial performed at a pleural clinic. Patients underwent protocolled ultrasound assessment pre-thoracentesis with measurements of lung and diaphragm movement and shear wave elastography measurements of the pleura and pleural effusion at the planned site of thoracentesis. RESULTS: M-mode measurements of lung movement provided the best diagnostic ROC-curve results, with an AUC of 0.81. Internal validity showed good results utilising the calibration belt test and Brier test. CONCLUSION: M-mode measurement of lung movement shows promise in diagnosing NEL before thoracentesis in patients with known or suspected MPE. A validation cohort is needed to confirm the results.

Safety and feasibility of oesophageal ultrasound for the work-up of thoracic malignancy in patients with respiratory impairment.

Biopsying lung tumours with endobronchial access in patients with respiratory impairment is challenging. However, fine needle aspiration with the endobronchial ultrasound-endoscope via the oesophagus (EUS-B-FNA) makes it possible to obtain tissue samples without entering the airways. Safety of EUS-B-FNA in these patients has not earlier been investigated prospectively. Therefore, this study aimed at assessing feasibility and safety of EUS-B-FNA from centrally located tumours suspected of thoracic malignancy in patients with respiratory insufficiency. The study is a prospective observational study. Patients with indication of EUS-B-FNA of centrally located tumours and respiratory impairment defined as modified Medical Research Council (mMRC) dyspnoea scale score of ≥3, saturation ≤90% or need of continuous oxygen supply were included prospectively in three centres. Any adverse events (AEs) were recorded during procedure and 1-hour recovery. AEs were defined as hypoxemia (saturation <90% or need for increased oxygen supply) or any kind of events needing intervention. Late procedure-related events were recorded during 30-day follow-up. Between April 1, 2020 and January 30, 2021, 16 patients were included. No severe AEs (SAEs) occurred, but AEs were seen in 50% (n=8) and 13% (n=2) of the patients during procedure and recovery respectively. AEs included hypoxemia corrected with increased oxygen supply and in two cases reversal of sedation. Late procedure-related events were seen in 13% (n=2) and included prolonged need of oxygen and one infection treated with oral antibiotics. In this cohort, EUS-B-FNA of centrally located tumours was safe and feasible in patients with respiratory impairment, when examined in the bronchoscopy suite. A variety of mostly mild and manageable complications may occur, a few even up to 30 days post-procedure.

Massive solitary fibrous intrathoracic tumor presented as primary lung cancer.

Solitary fibrous tumours (SFT) are rare soft tissue tumours with a primarily benign course. Complete surgical resection is the mainstay treatment. In this case report, a 75-year-old man had a massive intrathoracic SFT which was subsequently surgically resected without complications. Although the clinical presentation and CT features of these tumours can mimic lung cancer, the clinical course is significantly more favourable. Diagnostic examination and surgical treatment of intrathoracic SFT should be considered even in patients with increased post-operative risk.

Decay of Cryptosporidium parvfum DNA in cowpats in subtropical environments determined using qPCR.

Cryptosporidium oocysts pose a significant threat to public health due to its ability to contaminate environmental waters, leading to outbreaks of waterborne diseases and emphasizing the crucial need for effective water treatment and monitoring systems. This study aimed to investigate the decay of Cryptosporidium oocyst DNA in cow fecal matter under different environmental conditions prevalent in sub-tropical Southeast Queensland (SEQ) during summer and winter seasons. The effects of ambient sunlight and shaded conditions on the decay rates of C. parvum DNA in cow fecal samples were evaluated. The results showed that measurable levels of C. parvum DNA were observed for up to 60 days during the summer experiments, with a slower decay rate on the surface (k = -0.029) and sub-surface (k = -0.043) of the cowpat under shaded conditions than those on the surface (k = -0.064) and sub-surface (k = -0.079) under sunlight conditions. The decay rates of C. parvum DNA on the surface and sub-surface of the cowpat under shaded conditions were significantly slower (p = 0.004; p = 0.004) than those on the surface and sub-surface under sunlight conditions during summer experiments. During the winter treatments, measurable levels of C. parvum DNA were observed for up to 90 days, and the decay rates were slower on the surface (k = -0.036) and sub-surface (k = -0.034) of the cowpat under shaded conditions than those under sunlight conditions (k = -0.067 for surface and k = -0.057 for sub-surface). The decay rates of C. parvum DNA on the surface and sub-surface of the cowpat under shaded conditions were significantly slower than those on the surface (p = 0.009) and sub-surface (p = 0.041) under sunlight conditions during winter experiments. Moreover, the decay rate in the summer sunlight surface treatment (k = -0.064) was significantly faster from those in the winter shaded surface (k = -0.036; p = 0.018) and sub-surface (k = -0.034; p = 0.011) treatments. Similar results were also observed for summer sunlight sub-surface (k = -0.079), which was significantly faster than winter shaded surface (k = -0.036; p = 0.0008) and sub-surface (k = -0.034; p = 0.0005) treatments. Overall, these findings are important to enhance our understanding on the degradation of C. parvum DNA in cow fecal matter in SEQ, particularly in relation to seasonal variations and environmental conditions.

Endoscopic ultrasound-guided pancreas biopsy in the hands of a chest physician.

This is the first paper to report sampling of pancreatic lesions by EUS-B-FNA. A 76 year old patient suspected of primary lung cancer presented with a 36 × 24 mm lesion in the pancreas. Thoracentesis showed malignant cells suggestive of mucinous adenocarcinoma, but immunohistochemistry was inconclusive. Due to rapid deterioration of performance status of this frail patient, the program was shortened to EUS-B-FNA of the pancreatic lesion, which showed mucinous adenocarcinoma suggestive of primary pancreatic cancer. We conclude that EUS-B-FNA from a pancreatic lesion in the hands of a chest physician is feasible and diagnostic of a tumor in pancreas.

Microbiological characterization of stormwater in a high-income neighborhood in Rio de Janeiro, Brazil.

Stormwater harvesting and reuse in the urban environment is emerging as an alternative water source, despite human pathogens in the stormwater may represent a hazard to public health. This study presents the results of 1-year monitoring to evaluate the quality of stormwater obtained in a high-income neighborhood in Rio de Janeiro for a set of microbiological parameters as total coliforms, Escherichia coli (E. coli), human adenovirus (HAdV), human JC polyomavirus (JCPyV), Group A rotavirus (RVA), and norovirus GI and GII. Forty-eight stormwater samples obtained from two multiplex units presented total coliforms and E. coli in 91.7% (n = 44) and 58.3% (n = 28) of samples, while HAdV and JCPyV were detected in 20.8% (n = 10) and 12.5% (n = 6), respectively. Viral quantification ranged from 103 to 104 genomic copies/liter (GC/L) for HAdV and from 101 to 104 GC/L for JCPyV. Neither RVA nor norovirus GI and GII was detected. Fifteen out of sixteen (93.8%) samples containing viruses were compliant as per fecal indicator bacteria (FIB) according to Brazilian standards for rainwater reuse and US EPA Guidelines for Water Reuse, suggesting that viruses monitoring should complement the study of bacterial indicators.

Assessment of competence in local anaesthetic thoracoscopy: development and validity investigation of a new assessment tool.

BACKGROUND: The aims of the study were to develop an assessment tool in local anaesthetic thoracoscopy (LAT), investigate validity evidence, and establish a pass/fail standard. METHODS: Validity evidence for the assessment tool was gathered using the unified Messick framework. The tool was developed by five experts in respiratory medicine and medical education. Doctors with varying experience performed two consecutive procedures in a standardized, simulation-based setting using a newly developed thorax/lung silicone model. Performances were video-recorded and assessed by four expert raters using the new tool. Contrasting groups' method was used to set a pass/fail standard. RESULTS: Nine novices and 8 experienced participants were included, generating 34 recorded performances and 136 expert assessments. The tool had a high internal consistency (Cronbach's alpha =0.94) and high inter-rater reliability (Cronbach's alpha =0.91). The total item score significantly correlated with the global score (rs=0.86, P<0.001). Participants' first performance correlated to second performance (test-retest reliability) with a Pearson's r of 0.93, P<0.001. Generalisability (G) study showed a G-coefficient of 0.92 and decision (D) study estimated that one performance assessed by two raters or four performances assessed by one rater are needed to reach an acceptable reliability, i.e., G-coefficient >0.80. The tool was able to discriminate between the two groups in both performances: experienced mean score =30.8±4.2; novice mean score =15.8±2.3, P<0.001. Pass/fail standard was set at 22 points. CONCLUSIONS: The newly developed assessment tool showed solid evidence of validity and can be used to ensure competence in LAT.

Virological Characterization of Roof-Harvested Rainwater of Densely Urbanized Low-Income Region.

Roof-harvested rainwater (RHRW) is considered relatively clean water, even though the possible presence of pathogens in the water may pose human health risks. In this study, we investigated the occurrence of enteric viruses in the first flush (10 mm) of RHRW from a densely populated and low-income urbanized region of Rio de Janeiro. One hundred samples (5 L) were collected from 10 rainfall events between April 2015 and March 2017. RNA and DNA viruses were concentrated using the skimmed milk flocculation method and analyzed using the TaqMan® quantitative RT-qPCR and qPCR. Human adenoviruses, noroviruses, rotaviruses A, and avian parvoviruses were detected in 54%, 31%, 12%, and 12% of the positive samples. JC polyomavirus, also targeted, was not detected. Virus concentrations ranged from 1.09 × 101 to 2.58 × 103 genome copies/Liter (GC/L). Partial nucleotide sequence confirmed the presence of HAdV type 41, norovirus genotype GII.4, and avian parvovirus 1. The results suggest that the first flush diversion devices may not adequately remove enteric virus from the rainwater. Additional treatment of RHRW is required to mitigate potential health risks from potable use of captured water.

Characterization of Lung Tumors that the Pulmonologist can Biopsy from the Esophagus with Endosonography (EUS-B-FNA).

BACKGROUND: According to guidelines, it is possible to biopsy lung tumors "immediately adjacent to the esophagus" with EUS-B-FNA. However, it is unknown what "immediately adjacent" exactly means. OBJECTIVE: to investigate the possibility of achieving EUS-B-FNA biopsies from a lung tumor depending on the distance from the esophagus and to establish the maximal allowable distance between the tumor and the esophagus. METHODS: In a prospective observational study, we included patients with a lung tumor located maximum 6 cm from the esophagus and indication of EUS-B-FNA from the tumor. The tumors were of different sizes. In a plot presenting the tumor size-distance relationship in cases with (biopsy) versus without (non-biopsy) successful EUS-B-FNA, a separation line representing the threshold between the groups were identified and a biopsy-index equation established. The maximal tumor-size corrected distance (TSCD) was calculated using the residuals to the separation line. RESULTS: In total, 70 patients were included. EUS-B-FNA from the lung tumor was possible in 46 patients. All tumors with a distance from the esophagus below 19 mm could be biopsied. The maximal allowable esophagus-tumor distance depended on tumor size. From the separation line, a biopsy-index equation was established with the sensitivity of 93.5%, a specificity of 100%, and total accuracy of 95.7%. The TSCD was 31 mm (sensitivity: 95.7%, specificity 75.0%, and accuracy: 88.6%). CONCLUSION: We established a biopsy-index equation to predict the achievability of a lung tumor using EUS-B-FNA depending on distance to esophagus and tumor size. A general maximal TSCD was 31 mm.

EUS-B for suspected left adrenal metastasis in lung cancer.

BACKGROUND: Several studies have reported the efficacy of esophageal ultrasound-guided fine needle aspiration (EUS-FNA) for the detection of metastases in the left adrenal gland (LAG) in patients with lung cancer. Currently we have only limited evidence based on small studies on the usefulness of EUS-B [endobronchial ultrasound (EBUS) scope into the esophagus] to provide tissue proof of suspected LAG metastases. The objectives of this study are to investigate feasibility, safety and diagnostic yield of EUS-B-FNA in LAG analysis in patients with proven or suspected lung cancer. METHODS: In two Danish hospitals, a systematic search in the electronic database for patients who underwent EUS-B-FNA of the LAG for suspected or proven lung cancer was performed retrospectively between January 1st, 2015 and December 31st, 2017. Computed tomography (CT), positron emission tomography-CT, endoscopy, pathology and follow-up data were acquired. RESULTS: One hundred and thirty-five patients were included; the prevalence of biopsy proven LAG malignancy was 30% (40/135). A total of 87% (117/135) of EUS-B-FNA samples were adequate (i.e., containing adrenal or malignant cells). No complications were observed. CONCLUSIONS: We present the largest cohort of patients ever reported showing that EUS-B-FNA of the LAG is a safe and feasible procedure and should therefore be used for staging purposes in patients with lung cancer and a suspicious LAG.

Fluoroscopy guided percutaneous biopsy in combination with bronchoscopy and endobronchial ultrasound in the diagnosis of suspicious lung lesions - the triple approach.

Flexible bronchoscopy and endobronchial ultrasound guided transbronchial needle aspiration (EBUS-TBNA) are the pulmonologists´ basic procedures for the biopsy of suspicious lung lesions. If inconclusive, other guiding-modalities for tissue sampling are needed, computed tomography performed by a radiologist, or - if available - radial EBUS or electromagnetic navigation biopsy. We wanted to investigate if same-day X-ray fluoroscopy-guided transthoracic fine-needle aspiration biopsy (F-TTNAB) performed by the pulmonologist immediately after bronchoscopy and EBUS is a feasible alternative. We retrospectively identified consecutive patients in whom F-TTNAB followed a bronchoscopy and EBUS in the same séance. Patients in whom the suspicion of malignancy was invalidated after complete work up were followed for six months to identify false-negative cases. In total 125 patients underwent triple approach (bronchoscopy, EBUS and F-TTNAB) during the same séance. Malignancy was diagnosed in 86 (69%), and 77 of these (90%) were primary lung cancers. The diagnostic yield of F-TTNAB for malignancy was 77%, and sensitivity was 90%. Pneumothorax occurred in 35 (28%) patients, and was administered with pleural drainage in 22 (18% of all patients). No cases of prolonged haemoptysis were observed. The risk of pneumothorax differed insignificantly with lesion size ≤2.0 cm (27%) versus >2.0 cm (29%). We conclude that it is feasible for pulmonologist to perform F-TTNAB immediately after endoscopy as a combined triple approach in a fast-track workup of suspected lung cancer.

EUS-B-FNA for Diagnosing Liver and Celiac Metastases in Lung Cancer Patients.

BACKGROUND: In patients with suspected or proven lung cancer, assessment of regional nodal and distant metastases is key before treatment planning. By introducing the endobronchial ultrasound (EBUS)-guided scope into the esophagus and stomach (EUS-B), liver lesions and celiac nodes can be visualized. To date, the utility of EUS-B in diagnosing liver lesions and retroperitoneal lymph nodes is unknown. OBJECTIVES: To assess the feasibility, safety, and diagnostic yield of sampling of liver lesions and retroperitoneal nodes by EUS-B fine-needle aspiration (FNA) in a lung cancer staging setting. METHOD: Consecutive patients suspected of lung cancer in 2 Danish centers between 1 January 2015 and 31 December 2017 were included retrospectively when a lesion in the liver or a retroperitoneal lymph node was visualized and biopsied with EUS-B-FNA. RESULTS: 23 left liver lobe lesions and 19 retroperitoneal lymph nodes were sampled by EUS-B-FNA. Sensitivity and diagnostic yield of sampled liver lesions were 86 and 83%, respectively. In 19/23 patients, there was a cytopathological diagnosis of malignancy. Sensitivity and diagnostic yield from retroperitoneal lymph node samples were 83 and 63%, respectively. In 10/19 patients, the diagnosis was malignancy. No complications were observed. CONCLUSION: EUS-B-FNA enables safe sampling of left liver lobe lesions and retroperitoneal lymph nodes. EUS-B should be considered as a minimally invasive technique to provide tissue proof of distant metastases lung cancer patients.

Endoscopic Ultrasound with Bronchoscope-Guided Fine Needle Aspiration for the Diagnosis of Paraesophageally Located Lung Lesions.

BACKGROUND: Diagnosing centrally located lung tumors without endobronchial abnormalities and not located near the major airways is a diagnostic challenge. Tumors near or adjacent to the esophagus can be aspirated and detected with esophageal ultrasound (EUS) using gastrointestinal endoscopes. OBJECTIVE: To assess the feasibility and diagnostic yield of endoscopic ultrasound with bronchoscope-guided fine needle aspiration (EUS-B-FNA) in paraesophageally located lung tumors and its added value to bronchoscopy and endobronchial ultrasound (EBUS). METHODS: Retrospective, multicenter international study (from January 1, 2015 until January 1, 2018) of patients with suspected lung cancer, undergoing bronchoscopy, EBUS, and endoscopic ultrasound bronchoscopy (EUS-B) in one session by a single operator (pulmonologist), in whom the primary lung tumor was detected and aspirated by EUS-B. In the absence of malignancy following endoscopy, transthoracic ultrasound needle aspiration, clinical and radiological follow-up of at least 6 months was performed. The yield and sensitivity of EUS-B-FNA and its added value to bronchoscopy and EBUS was assessed. RESULTS: 58 patients were identified with the following diagnosis: non-small-cell lung cancer (n = 43), small-cell lung cancer (n = 6), mesothelioma (n = 2), metastasis (n = 1), nonmalignant (n = 6). The yield and sensitivity of EUS-B-FNA for detecting lung cancer was 90%. In 26 patients (45%), the intrapulmonary tumor was exclusively detected by EUS-B. Adding EUS-B to conventional bronchoscopy and EBUS increased the diagnostic yield for diagnosing lung cancer in para-esophageally located lung tumors from 51 to 91%. No EUS-B-related complications were observed. CONCLUSION: EUS-B-FNA is a feasible and safe technique for diagnosing centrally located intrapulmonary tumors that are located near or adjacent to the esophagus. EUS-B should be considered in the same endoscopy session following nondiagnostic bronchoscopy and EBUS.

Quantification of hookworm ova from wastewater matrices using quantitative PCR.

A quantitative PCR (qPCR) assay was used to quantify Ancylostoma caninum ova in wastewater and sludge samples. We estimated the average gene copy numbers for a single ovum using a mixed population of ova. The average gene copy numbers derived from the mixed population were used to estimate numbers of hookworm ova in A. caninum seeded and unseeded wastewater and sludge samples. The newly developed qPCR assay estimated an average of 3.7×103 gene copies per ovum, which was then validated by seeding known numbers of hookworm ova into treated wastewater. The qPCR estimated an average of (1.1±0.1), (8.6±2.9) and (67.3±10.4) ova for treated wastewater that was seeded with (1±0), (10±2) and (100±21) ova, respectively. The further application of the qPCR assay for the quantification of A. caninum ova was determined by seeding a known numbers of ova into the wastewater matrices. The qPCR results indicated that 50%, 90% and 67% of treated wastewater (1L), raw wastewater (1L) and sludge (~4g) samples had variable numbers of A. caninum gene copies. After conversion of the qPCR estimated gene copy numbers to ova for treated wastewater, raw wastewater, and sludge samples, had an average of 0.02, 1.24 and 67 ova, respectively. The result of this study indicated that qPCR can be used for the quantification of hookworm ova from wastewater and sludge samples; however, caution is advised in interpreting qPCR generated data for health risk assessment.

Optimization of sampling strategy to determine pathogen removal efficacy of activated sludge treatment plant.

Large-scale wastewater schemes rely on multi-barrier approach for the production of safe and sustainable recycled water. In multi-barrier wastewater reclamation systems, conventional activated sludge process (ASP) often constitutes a major initial treatment step. The main aim of this research was to determine most appropriate sampling approach to establish pathogen removal efficacy of ASP. The results suggest that ASP is capable of reducing human adenovirus (HAdV) and polyomavirus (HPyV) by up to 3 log10. The virus removal data suggests that HAdV removal is comparable to somatic bacteriophage belonging to Microviridae family. Due to the high removal of Escherichia coli (>3 log10) and very poor correlation with the enteric virus, it is not recommended that E. coli be used as a surrogate for enteric virus removal. The results also demonstrated no statistically significant differences (t test, P > 0.05) in calculated log removal values (LRVs) for HAdV, HPyV, and Microviridae from samples collected on hydraulic retention time (HRT) or simultaneous paired samples collected for influent and effluent. This indicates that a more practical approach of simultaneous sampling for influent and effluent could be used to determine pathogen removal efficiency of ASP. The results also suggest that a minimum of 10, preferably 20 samples, are required to fully capture variability in the removal of virus. In order to cover for the potential seasonal prevalence of viruses such as norovirus and rotavirus, sampling should be spread across all seasons.

Comparative prevalence of Escherichia coli carrying virulence genes and class 1 and 2 integrons in sub-tropical and cool temperate freshwater.

Aquatic environments are now recognized secondary habitat of potentially pathogenic Escherichia coli. In this study, PCR-based analyses were used to determine the phylogenetic composition and frequency of occurrence of eight clinically significant virulence genes (VGs) in E. coli isolates from sub-tropical Brisbane and cool temperate Tasmania freshwater in Australia. In Brisbane, non-commensal E. coli isolates belonging to the B2 and D phylogenetic group were dominant (72%). A significantly higher number (P < 0.05) of E. coli carrying VGs were detected in the sub-tropical freshwaters compared to the cool temperate water. Furthermore, diarrheagenic pathotype (EHEC) was also observed in the sub-tropical freshwater. The genes east1 and eaeA were significantly more common (P < 0.00001) than other VGs. The eaeA gene which codes for intimin protein along with toxin genes east1, stx 1 , stx 2 , and LT1 were mostly detected in phylogenetic groups B2 and D. The ANOVA results also suggested a statistically significant difference (P < 0.016) between the VGs carried by phylogenetic groups B2 and D. Class 1 integrase (intl1) and class 2 integrase (intl2) genes were detected in 38 (24.83%) and 23 (15.03%) of E. coli isolates, respectively. The Gretna site (Tasmania) with known fecal input from bovine and ovine sources had the highest number of E. coli carrying intl1 (29%) and intl2 (13%) genes. In addition, class 2 integron was more commonly detected in the phylogenetic group B2. The results of this study highlight the need to better understand sources and reasons for the high prevalence of E. coli carrying clinically significant VGs in a sub-tropical environment and its public health implications.

Pesticide occurrence and spatio-temporal variability in urban run-off across Australia.

Stormwater is a major driving factor of aquatic ecosystem degradation as well as one of the largest untapped urban freshwater resources. We present results from a long-term, multi-catchment study of urban stormwater pesticides across Australia that addresses this dichotomous identity (threat and resource), as well as dominant spatial and temporal patterns in stormwater pesticide composition. Of the 27 pesticides monitored, only 19 were detected in Australian stormwater, five of which (diuron, MCPA, 2,4-D, simazine, and triclopyr) were found in >50% of samples. Overall, stormwater pesticide concentrations were lower than reported in other countries (including the United States, Canada and Europe), and exceedances of public health and aquatic ecosystem standards were rare (<10% of samples). Spatio-temporal patterns were investigated with principal component analysis. Although stormwater pesticide composition was relatively stable across seasons and years, it varied significantly by catchment. Common pesticide associations appear to reflect 1) user application of common registered formulations containing characteristic suites of active ingredients, and 2) pesticide fate properties (e.g., environmental mobility and persistence). Importantly, catchment-specific occurrence patterns provide opportunities for focusing treatment approaches or stormwater harvesting strategies.

Amplicon-based profiling of bacteria in raw and secondary treated wastewater from treatment plants across Australia.

In this study, we investigated the use of Illumina high-throughput sequencing of 16S ribosomal RNA (rRNA) amplicons to explore microbial diversity and community structure in raw and secondary treated wastewater (WW) samples from four municipal wastewater treatment plants (WWTPs A-D) across Australia. Sequence reads were analyzed to determine the abundance and diversity of bacterial communities in raw and secondary treated WW samples across the four WWTPs. In addition, sequence reads were also characterized to phenotypic features and to estimate the abundance of potential pathogenic bacterial genera and antibiotic-resistant genes in total bacterial communities. The mean coverage, Shannon diversity index, observed richness (S obs), and abundance-based coverage estimate (ACE) of richness for raw and secondary treated WW samples did not differ significantly (P > 0.05) among the four WWTPs examined. Generally, raw and secondary treated WW samples were dominated by members of the genera Pseudomonas, Arcobacter, and Bacteroides. Evaluation of source contributions to secondary treated WW, done using SourceTracker, revealed that 8.80-61.4% of the bacterial communities in secondary treated WW samples were attributed to raw WW. Twenty-five bacterial genera were classified as containing potential bacterial pathogens. The abundance of potentially pathogenic genera in raw WW samples was higher than that found in secondary treated WW samples. Among the pathogenic genera identified, Pseudomonas and Arcobacter had the greatest percentage of the sequence reads. The abundances of antibiotic resistance genes were generally low (<0.5%), except for genes encoding ABC transporters, which accounted for approximately 3% of inferred genes. These findings provided a comprehensive profile of bacterial communities, including potential bacterial pathogens and antibiotic-resistant genes, in raw and secondary treated WW samples from four WWTPs across Australia and demonstrated that Illumina high-throughput sequencing can be an alternative approach for monitoring WW quality in order to protect environmental and human health.