In this study, we evaluated the use of RT-qPCR assays targeting rRNA gene sequences for the detection of fecal bacteria in water samples. We challenged the RT-qPCR assays against RNA extracted from ...sewage effluent (n = 14), surface water (n = 30), and treated source water (n = 15) samples. Additionally, we applied the same assays using DNA as the qPCR template. The targeted fecal bacteria were present in most of the samples tested, although in several cases, the detection frequency increased when RNA was used as the template. For example, the majority of samples that tested positive for E. coli and Campylobacter spp. in surface waters, and for human-specific Bacteroidales, E. coli, and Enterococcus spp. in treated source waters were only detected when rRNA was used as the original template. The difference in detection frequency using rRNA or rDNA (rRNA gene) was sample- and assay-dependent, suggesting that the abundance of active and nonactive populations differed between samples. Statistical analyses for each population exhibiting multiple quantifiable results showed that the rRNA copy numbers were significantly higher than the rDNA counterparts (p < 0.05). Moreover, the detection frequency of rRNA-based assays were in better agreement with the culture-based results of E. coli, intestinal enterococci, and thermotolerant Campylobacter spp. in surface waters than that of rDNA-based assays, suggesting that rRNA signals were associated to active bacterial populations. Our data show that using rRNA-based approaches significantly increases detection sensitivity for common fecal bacteria in environmental waters. These findings have important implications for microbial water quality monitoring and public health risk assessments.
Many factors, including microbiome structure and activity in the drinking water distribution system (DWDS), affect the colonization potential of opportunistic pathogens. The present study aims to ...describe the dynamics of active bacterial communities in DWDS and identify the factors that shape the community structures and activity in the selected DWDSs. Large-volume drinking water and hot water, biofilm, and water meter deposit samples were collected from five DWDSs. Total nucleic acids were extracted, and RNA was further purified and transcribed into its cDNA from a total of 181 water and biofilm samples originating from the DWDS of two surface water supplies (disinfected with UV and chlorine), two artificially recharged groundwater supplies (non-disinfected), and a groundwater supply (disinfected with UV and chlorine). In chlorinated DWDSs, concentrations of <0.02-0.97 mg/l free chlorine were measured. Bacterial communities in the RNA and DNA fractions were analysed using Illumina MiSeq sequencing with primer pair 341F-785R targeted to the 16S rRNA gene. The sequence libraries were analysed using QIIME pipeline, Program R, and MicrobiomeAnalyst. Not all bacterial cells were active based on their 16S rRNA content, and species richness was lower in the RNA fraction (Chao1 mean value 490) than in the DNA fraction (710). Species richness was higher in the two DWDSs distributing non-disinfected artificial groundwater (Chao1 mean values of 990 and 1 000) as compared to the two disinfected DWDSs using surface water (Chao1 mean values 190 and 460) and disinfected DWDS using ground water as source water (170). The difference in community structures between non-disinfected and disinfected water was clear in the beta-diversity analysis. Distance from the waterworks also affected the beta diversity of community structures, especially in disinfected distribution systems. The two most abundant bacteria in the active part of the community (RNA) and total bacterial community (DNA) belonged to the classes Alphaproteobacteria (RNA 28 %, DNA 44 %) and Gammaproteobacteria (RNA 32 %, DNA 30 %). The third most abundant and active bacteria class was Vampirovibrionia (RNA 15 %), whereas in the total community it was Paceibacteria (DNA 11 %). Class Nitrospiria was more abundant and active in both cold and hot water in DWDS that used chloramine disinfection compared to non-chlorinated or chlorine-using DWDSs. Thirty-eight operational taxonomic units (OTU) of Legionella, 30 of Mycobacterium, and 10 of Pseudomonas were detected among the sequences. The (RT)-qPCR confirmed the presence of opportunistic pathogens in the DWDSs studied as Legionella spp. was detected in 85 % (mean value 4.5 × 10
gene copies/100 ml), Mycobacterium spp. in 95 % (mean value 8.3 × 10
gene copies/100 ml), and Pseudomonas spp. in 78 % (mean value 1.6 × 10
gene copies/100 ml) of the water and biofilm samples. Sampling point inside the system (distance from the waterworks and cold/hot system) affected the active bacterial community composition. Chloramine as a chlorination method resulted in a recognizable community composition, with high abundance of bacteria that benefit from the excess presence of nitrogen. The results presented here confirm that each DWDS is unique and that opportunistic pathogens are present even in conditions when water quality is considered excellent.
Failures in the drinking water distribution system cause gastrointestinal outbreaks with multiple pathogens. A water distribution pipe breakage caused a community-wide waterborne outbreak in Vuorela, ...Finland, July 2012. We investigated this outbreak with advanced epidemiological and microbiological methods. A total of 473/2931 inhabitants (16%) responded to a web-based questionnaire. Water and patient samples were subjected to analysis of multiple microbial targets, molecular typing and microbial community analysis. Spatial analysis on the water distribution network was done and we applied a spatial logistic regression model. The course of the illness was mild. Drinking untreated tap water from the defined outbreak area was significantly associated with illness (RR 5.6, 95% CI 1.9-16.4) increasing in a dose response manner. The closer a person lived to the water distribution breakage point, the higher the risk of becoming ill. Sapovirus, enterovirus, single Campylobacter jejuni and EHEC O157:H7 findings as well as virulence genes for EPEC, EAEC and EHEC pathogroups were detected by molecular or culture methods from the faecal samples of the patients. EPEC, EAEC and EHEC virulence genes and faecal indicator bacteria were also detected in water samples. Microbial community sequencing of contaminated tap water revealed abundance of Arcobacter species. The polyphasic approach improved the understanding of the source of the infections, and aided to define the extent and magnitude of this outbreak.
Knowledge of the decay characteristics of health-related microbes in surface waters is important for modeling the transportation of waterborne pathogens and for assessing their public health risks. ...Although water temperature and light exposure are major factors determining the decay characteristics of enteric microbes in surface waters, such effects have not been well studied in subarctic surface waters. This study comprehensively evaluated the effect of temperature and light on the decay characteristics of health-related microbes Escherichia coli, enterococci, microbial source tracking markers (GenBac3 & HF183 assays), coliphages (F-specific and somatic), noroviruses GII and Legionella spp. under simulated subarctic river water conditions. The experiments were conducted in four different laboratory settings (4 °C/dark, 15 °C/dark, 15 °C/light, and 22 °C/light). The T90 values (time required for a 90 % reduction in the population of a target) of all targets were higher under cold and dark (2.6–51.3 days depending upon targets) than under warm and light conditions (0.6–3.5 days). Under 4 °C/dark (simulated winter) water conditions, F-specific coliphages had 27.2 times higher, and coliform bacteria had 3.3 times higher T90 value than under 22 °C/light (simulated summer) water conditions. Bacterial molecular markers also displayed high variation in T90 values, with the greatest difference between 4 °C/dark and 22 °C/light recorded for HF183 DNA (20.6 times) and the lowest difference for EC23S857 RNA (6.6 times). E. coli, intestinal enterococci, and somatic coliphages were relatively more sensitive to light than water temperature, but F-specific coliphages, norovirus, and all bacterial rDNA and rRNA markers were relatively more sensitive to temperature than light exposure. Due to the slow microbial decay in winter under subarctic conditions, the microbial quality of river water might remain low for a long time after a sewage spill. This increased risk associated with fecal pollution during winter may deserve more attention, especially when river waters are used for drinking water production.
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•Coliphages decayed 22–27 times, and E. coli 4.8 times slower in 4 °C than 22 °C.•E. coli, enterococci, and som. coliphages were more sensitive to light than temperature.•Bacterial DNA/RNA, and Legionella were more sensitive to temperature than light.•Norovirus GII and F-specific coliphages were also more sensitive to temperature.•Indication power of fecal indicators can vary between seasons of the year.
The latest developments in oncological therapies for malignant melanoma, and the discovery that complete lymph node dissection offers no survival benefit, are changing the landscape of melanoma ...surgery. There is a need for more information on health-related quality of life (HRQoL) consequences of melanoma surgery.
This longitudinal cohort study was carried out from 2004 to 2009 in the Helsinki and Uusimaa Hospital District and patients were followed-up at 6, 12 and 24 months. The patients were asked to fill in the generic 15D questionnaire and the cancer-specific European Organization for Research and Treatment of Cancer Quality of Life Questionnaire (EORTC QLQ-30). In addition, they were asked selected questions from the EORTC Item Library regarding upper and lower limb edema.
A total of 169 (64.5%) patients with local or locally advanced melanoma referred for surgical treatment responded, of whom 161 were included in the final analysis. For the whole patient group, distress, depression and emotional function improved over time. Worse HRQoL in some of the dimensions were associated with female sex, skin transplant versus direct wound closure and complications 30 days or more after surgery, but none was associated with worse overall HRQoL. Postoperative complications, type of wound closure or lymph node surgery had no effect on overall HRQoL. Patient-reported limb edema was associated with worse overall HRQoL at baseline and during follow-up by both instruments. Patients reporting limb edema reported worse mobility and more pain throughout the study.
Patient-reported limb edema, regardless of the cause, seems to be an important predictor of worse HRQoL among patients with melanoma.
Resection of colorectal cancer (CRC) metastases provides good survival but is probably underused in real-world practice.
A prospective Finnish nationwide study enrolled treatable metastatic CRC ...patients. The intervention was the assessment of resectability upfront and twice during first-line therapy by the multidisciplinary team (MDT) at Helsinki tertiary referral centre. The primary outcome was resection rates and survival.
In 2012–2018, 1086 patients were included. Median follow-up was 58 months. Multiple metastatic sites were present in 500 (46%) patients at baseline and in 820 (76%) during disease trajectory. In MDT assessments, 447 (41%) were classified as resectable, 310 (29%) upfront and 137 (18%) after conversion therapy. Six-hundred and ninety curative intent resections or local ablative therapies (LAT) were performed in 399 patients (89% of 447 resectable). Multiple metastasectomies for multisite or later developing metastases were performed in 148 (37%) patients. Overall, 414 liver, 112 lung, 57 peritoneal, and 107 other metastasectomies were performed. Median OS was 80·4 months in R0/1-resected (HR 0·15; CI95% 0·12–0·19), 39·1 months in R2-resected/LAT (0·39; 0·29–0·53) patients, and 20·8 months in patients treated with “systemic therapy alone” (reference), with 5-year OS rates of 66%, 40%, and 6%, respectively.
Repeated centralized MDT assessment in real-world metastatic CRC patients generates high resectability (41%) and resection rates (37%) with impressive survival, even when multisite metastases are present or develop later.
The funders had no role in the study design, analysis, and interpretation of the data or writing of this report.
There is a growing need for information regarding the Health-Related Quality of Life (HRQoL) of cancer survivors. This study aimed to assess the HRQoL of patients treated for cutaneous malignant ...melanoma between 1980 and 2004 in the Helsinki and Uusimaa Hospital district and compare the results to the general population.
HRQoL of 981 cutaneous melanoma patients (aged 13 to 97 years, 56.1% female) was assessed using the generic 15D instrument and compared to the general population. The association between demographic and clinical factors and HRQoL was analyzed using oneway ANOVA, student's t-test and multivariate regression.
The mean 15D score of melanoma patients was slightly lower (0.904) than that of the general population (0.911, p=0.027), but the difference was not statistically significant. HRQoL deteriorates with age and metastatic disease and improves with time.
No evidence was found that long-term HRQoL of melanoma survivors was worse than the general population.
ABSTRACT
A major goal of evolutionary science is to understand how biological diversity is generated and altered. Despite considerable advances, we still have limited insight into how phenotypic ...variation arises and is sorted by natural selection. Here we argue that an integrated view, which merges ecology, evolution and developmental biology (eco evo devo) on an equal footing, is needed to understand the multifaceted role of the environment in simultaneously determining the development of the phenotype and the nature of the selective environment, and how organisms in turn affect the environment through eco evo and eco devo feedbacks. To illustrate the usefulness of an integrated eco evo devo perspective, we connect it with the theory of resource polymorphism (i.e. the phenotypic and genetic diversification that occurs in response to variation in available resources). In so doing, we highlight fishes from recently glaciated freshwater systems as exceptionally well‐suited model systems for testing predictions of an eco evo devo framework in studies of diversification. Studies on these fishes show that intraspecific diversity can evolve rapidly, and that this process is jointly facilitated by (i) the availability of diverse environments promoting divergent natural selection; (ii) dynamic developmental processes sensitive to environmental and genetic signals; and (iii) eco evo and eco devo feedbacks influencing the selective and developmental environments of the phenotype. We highlight empirical examples and present a conceptual model for the generation of resource polymorphism – emphasizing eco evo devo, and identify current gaps in knowledge.
The aim of this study was to develop and test a generic framework to enable international collaboration for producing and sharing results of health technology assessments (HTAs).
Ten international ...teams constructed the HTA Core Model, dividing information contained in a comprehensive HTA into standardized pieces, the assessment elements. Each element contains a generic issue that is translated into practical research questions while performing an assessment. Elements were described in detail in element cards. Two pilot assessments, designated as Core HTAs were also produced. The Model and Core HTAs were both validated. Guidance on the use of the HTA Core Model was compiled into a Handbook.
The HTA Core Model considers health technologies through nine domains. Two applications of the Model were developed, one for medical and surgical interventions and another for diagnostic technologies. Two Core HTAs were produced in parallel with developing the model, providing the first real-life testing of the Model and input for further development. The results of formal validation and public feedback were primarily positive. Development needs were also identified and considered. An online Handbook is available.
The HTA Core Model is a novel approach to HTA. It enables effective international production and sharing of HTA results in a structured format. The face validity of the Model was confirmed during the project, but further testing and refining are needed to ensure optimal usefulness and user-friendliness. Core HTAs are intended to serve as a basis for local HTA reports. Core HTAs do not contain recommendations on technology use.
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