Spatial patterns in host-associated and free-living bacterial communities across six temperate estuaries.

A major goal of microbial ecology is to establish the importance of spatial and environmental factors in driving community variation. Their relative importance likely varies across spatial scales, but focus has primarily been on free-living communities within well-connected aquatic environments rather than less connected island-like habitats such as estuaries, and key host-associated communities within these systems. Here we sampled both free-living (seawater and sediment) and host-associated (estuarine fish hindgut microbiome, Pelates sexlineatus) communities across six temperate Australian estuaries spanning ∼500 km. We find that spatial and environmental factors have different influences on these communities, with seawater demonstrating strong distance-decay relationships (R = -0.69) and significant associations with a range of environmental variables. Distance-decay relationships were weak for sediment communities but became stronger over smaller spatial scales (within estuaries, R = -0.5), potentially reflecting environmental filtering across biogeochemical gradients or stochastic processes within estuary sediments. Finally, P. sexlineatus hindgut microbiome communities displayed weak distance-decay relationships (R = -0.36), and limited variation explained by environmental variables, indicating the significance of host-related factors in driving community variation. Our findings provide important ecological insights into the spatial distributions and driving forces of both free-living and host-associated bacterial patterns across temperate estuarine systems.

eDNA metabarcoding reveals shifts in sediment eukaryote communities in a metal contaminated estuary.

Metal contamination is a global issue impacting biodiversity in urbanised estuaries. Traditional methods to assess biodiversity are time consuming, costly and often exclude small or cryptic organisms due to difficulties with morphological identification. Metabarcoding approaches have been increasingly recognised for their utility in monitoring, however studies have focused on freshwater and marine systems despite the ecological significance of estuaries. We targeted estuarine eukaryote communities within the sediments of Australia's largest urbanised estuary, where a history of industrial activity has resulted in a metal contamination gradient. We identified specific eukaryote families with significant correlations with bioavailable metal concentrations, indicating sensitivity or tolerance to specific metals. While polychaete families Terebellidae and Syllidae demonstrated tolerance to the contamination gradient, members of the meio- and microfaunal communities including diatoms, dinoflagellates and nematodes displayed sensitivities. These may have high value as indicators but are frequently missed in traditional surveys due to sampling limitations.

Elevated estuary water temperature drives fish gut dysbiosis and increased loads of pathogenic vibrionaceae.

Marine water temperatures are increasing globally, with eastern Australian estuaries warming faster than predicted. There is growing evidence that this rapid warming of coastal waters is increasing the abundance and virulence of pathogenic members of the Vibrionaceae, posing a significant health risk to both humans and aquatic organisms. Fish disease, notably outbreaks of emerging pathogens in response to environmental perturbations such as heatwaves, have been recognised in aquaculture settings. Considerably less is known about how rising sea surface temperatures will impact the microbiology of wild fish populations, particularly those within estuarine systems that are more vulnerable to warming. We used a combination of Vibrio-specific quantitative PCR and amplicon sequencing of the 16S rRNA and hsp60 genes to examine seawater and fish (Pelates sexlineatus) gut microbial communities across a quasi-natural experimental system, where thermal pollution from coal-fired power stations creates a temperature gradient of up to 6 °C, compatible with future predicted temperature increases. At the warmest site, fish hindgut microbial communities were in a state of dysbiosis characterised by shifts in beta diversity and a proliferation (71.5% relative abundance) of the potential fish pathogen Photobacterium damselae subsp. damselae. Comparable patterns were not identified in the surrounding seawater, indicating opportunistic proliferation within estuarine fish guts under thermal stress. A subsequent evaluation of predicted future warming-related risk due to pathogenic Vibrionaceae in temperate estuarine fish demonstrated that warming is likely to drive opportunistic pathogen increases in the upper latitudinal range of this estuarine fish, potentially impacting adaptations to future warming. These findings represent a breakthrough in our understanding of the dynamics of emerging pathogens in populations of wild aquatic organisms within environments likely to experience rapid warming under future climate change.

Legacy metal contamination is reflected in the fish gut microbiome in an urbanised estuary.

Estuaries are critical habitats subject to a range of stressors requiring effective management. Microbes are gaining recognition as effective environmental indicators, however, the response of host associated communities to stressors remains poorly understood. We examined microbial communities from seawater, sediments and the estuarine fish Pelates sexlineatus, in Australia's largest urbanised estuary, and hypothesised that anthropogenic contamination would be reflected in the microbiology of these sample types. The human faecal markers Lachno3 and HF183 were not detected, indicating negligible influence of sewage, but a gradient in copy numbers of the class 1 integron (intI-1), which is often used as a marker for anthropogenic contamination, was observed in sediments and positively correlated with metal concentrations. While seawater communities were not strongly driven by metal contamination, shifts in the diversity and composition of the fish gut microbiome were observed, with statistical links to levels of metal contamination (F2, 21 = 1.536, p < 0.01). Within the fish gut microbiome, we further report increased relative abundance of amplicon sequence variants (ASVs; single inferred DNA sequences obtained in sequencing) identified as metal resistant and potentially pathogenic genera, as well as those that may have roles in inflammation. These results demonstrate that microbial communities from distinct habitats within estuarine systems have unique response to stressors, and alterations of the fish gut microbiome may have implications for the adaptation of estuarine fish to legacy metal contamination.

Tracking the impacts of nutrient inputs on estuary ecosystem function.

Estuaries are one of the most impacted coastal environments globally, subjected to multiple stressors from urban, industry and coastal development. With increasing anthropogenic activity surrounding estuarine systems, sewage inputs have become a common concern. Stable isotope analysis provides a well-established tool to investigate the incorporation of nitrogen into marine organisms and identify major nutrient sources. Benthic macroinvertebrate communities are often used as bioindicators in ecological studies as they typically display predictable responses to anthropogenic pressures, however have a suite of limitations and costs associated with their use. 16S rDNA amplicon sequencing techniques allow for investigation of the microbial communities inhabiting complex environmental samples, with potential as a tool in the ecological assessment of pollution. These communities have not yet been adequately considered for ecological studies and biomonitoring, with a need to better understand interactions with environmental stressors and implications for ecosystem function. This study used a combination of stable isotope analysis to trace the uptake of anthropogenic nitrogen in biota, traditional assessment of benthic macroinvertebrate communities, and 16S rDNA genotyping of benthic microbial communities. Stable isotope analysis of seagrass and epiphytes identified multiple treated and untreated sewage inputs, ranges of 5.2-7.2‰ and 1.9-4.0‰ for δ15N respectively, as the dominant nitrogen source at specific locations. The benthic macroinvertebrate community reflected these inputs with shifts in dominant taxa and high abundances of polychaetes at some sites. Microbial communities provided a sensitive indication of impact with a breadth of information not available using traditional techniques. Composition and predicted function reflected sewage inputs, particularly within sediments, with the relative abundance of specific taxa and putative pathogens linked to these inputs. This research supports the growing body of evidence that benthic microbial communities respond rapidly to anthropogenic stressors and have potential as a monitoring tool in urban estuarine systems.

Comparison between the two systems to evaluate the appropriateness of endoscopy of the upper digestive tract.

OBJECTIVES: The aim of this study was to compare the diagnostic performance of the two systems for the evaluation of the appropriateness of upper digestive endoscopy suggested by the American Society of Gastrointestinal Endoscopy (ASGE) and by the European Panel on the Appropriateness of Gastrointestinal Endoscopy (EPAGE). METHODS: Patients referred for the upper digestive endoscopy (EGD) to a University Outpatients Clinic of Northeastern Italy were consecutively included in this prospective observational study. Before the EGD, the endoscopist assigned the patients to one of the ASGE appropriateness classes; another endoscopist then identified the detailed clinical scenario for the patients, which corresponds to scenarios examined by EPAGE by using a nine-point scale: 1-3 inappropriate; 4-6 uncertain; and 7-9 appropriate. The relationship between the appropriateness of use and the presence of relevant endoscopic lesions (neoplasms, ulcers, esophagitis, erosive gastritis/duodenitis, stenosis, and varices) was assessed, calculating the sensitivity and the specificity for each of the ASGE criteria, and each of the EPAGE scores, and plotting them to form a receiver operating characteristic (ROC) curve. The area under the ROC curve (AUC) provides a summary measure of test performance, and can vary from a minimum of 0.5 to a maximum of 1.0. We compared the AUC of the ROC curve derived from the ASGE criteria against that derived from the EPAGE criteria. RESULTS: A total of 2,300 consecutive patients were included in the study (42% men; mean age: 57.3; range: 12-99); comparison of appropriateness criteria according to the ASGE and EPAGE could be made for 2,000 patients. The AUC of the ROC curve derived from the ASGE criteria was 0.553 (95% CI: 0.527-0.579), significantly higher than the AUC of the ROC curve derived from the EPAGE score: 0.523 (95% CI: 0.497-0.549; p < 0.05). CONCLUSIONS: We suggest that the diagnostic yield for relevant endoscopic findings obtained by both the systems (ASGE and EPAGE) is low; slightly better results could be accomplished by the ASGE criteria.

ASGE guidelines for the appropriate use of upper endoscopy: association with endoscopic findings.

BACKGROUND: This prospective study examined the appropriate use of EGD in an open-access system with the American Society for Gastrointestinal Endoscopy (ASGE) guidelines and determined whether the ASGE guidelines were associated with relevant endoscopic findings. METHODS: In a cohort of 1777 consecutive patients referred for open-access EGD, the proportion of patients who underwent EGD for appropriate indications was prospectively assessed. The relationship between appropriateness and the presence of clinically relevant endoscopic diagnoses was assessed by calculating (1) the likelihood ratio, positive and negative, of the indications; and (2) the change in the probability of relevant endoscopic diagnoses in the presence of the ASGE criteria. RESULTS: The rate for EGDs "generally not indicated" was 15.6%. Relevant endoscopic diagnoses were present in 47.4% of cases with ASGE indications versus 28.8% of patients without appropriate indications as defined by the ASGE criteria (OR: 2.23; 99% CI [1.55, 3.22]; p < 0.01). A similar difference was observed for erosive gastritis (OR: 1.86; 99% CI [1.17, 2.95]; p < 0.01), erosive esophagitis (OR: 1.48; 99% CI [0.87, 2.52]; p < 0.05), and Barrett's esophagus (OR: 9.76; 99% CI [0.72, 132]; p < 0.05). The pretest probability of finding a relevant endoscopic diagnosis was modified slightly when an ASGE indication(s) was present and decreased markedly when ASGE criteria were absent. CONCLUSIONS: The use of the ASGE guideline for appropriate indications for EGD can improve patient selection for the procedure. However, to avoid missed diagnoses of serious disease, use of the guidelines must be tailored to the specific clinical setting.