Tank Water Samples Research Articles

A Gold Nanoparticle-Based Cortisol Aptasensor for Non-Invasive Detection of Fish Stress.

Cortisol is a key stress biomarker in humans and animals, including fishes. In aquafarming, stress monitoring using cortisol quantification can help to optimize aquaculture practices for welfare and productivity enhancement. However, most current methods for cortisol detection rely on invasive tissue sampling. In this work, we developed a gold nanoparticle (AuNP)-based cortisol sensor to address the demand of detecting picomolar ranges of cortisol from complex fish tank water matrices as a non-invasive alternative for more effective stress monitoring. We first identified a DNA aptamer with effective binding to cortisol and then conjugated the thiol-labelled aptamer to AuNPs together with a blocker molecule (CALNN) to form an Au-Apt-CALNN conjugate that is stable in fish tank water. The cortisol detection principle is based on magnesium chloride (MgCl2)-induced particle aggregation, where the cortisol-bound aptamer on the AuNPs folds into a tertiary structure and provides greater protection for Au-Apt-CALNN against MgCl2-induced aggregation due to steric stabilization. At an optimum MgCl2 concentration, the differential stability of particles with and without cortisol binding offers a limit of detection (LOD) of 100 pM for cortisol within a 35 min reaction. The aptasensor has been validated on recirculating aquaculture system (RAS) fish tank water samples by the HPLC method and was able to detect changes in water cortisol induced by two different stress paradigms. This on-site deployable and non-invasive sensor offers opportunities for more efficient and real-time fish stress monitoring for the optimization of aquaculture practices.

Detection of Free-Living Amoeba in a tertiary care hospital

This study aims to investigate the presence of Free-Living Amoebae (FLA), particularly Acanthamoeba spp., in diverse water systems and body fluids within a hospital. Water and body fluid samples were collected from various locations. Cultivation of FLA was performed using Non-Nutrient Agar (NNA) culture with Escherichia coli, and microscopic observations after staining. Microscopic observations revealed the presence of Acanthamoeba spp. in pre-dialysis tank water and dental flush water samples. The identification of Acanthamoeba in specific water sources underscores the importance of continuous monitoring and preventive strategies to mitigate potential health risks.

Quantification and cultivation of Helicobacter pylori (H. pylori) from various urban water environments: A comprehensive analysis of precondition methods and sample characteristics

Substantial evidence suggests that all types of water, such as drinking water, wastewater, surface water, and groundwater, can be potential sources of Helicobacter pylori (H. pylori) infection. Thus, it is critical to thoroughly investigate all possible preconditioning methods to enhance the recovery of H. pylori, improve the reproducibility of subsequent detection, and optimize the suitability for various water types and different detection purposes. In this study, we proposed and evaluated five distinct preconditioning methods for treating water samples collected from multiple urban water environments, aiming to maximize the quantitative qPCR readouts and achieve effective selective cultivation. According to the experimental results, when using the qPCR technique to examine WWTP influent, effluent, septic tank, and wetland water samples, the significance of having a preliminary cleaning step becomes more evident as it can profoundly influence qPCR detection results. In contrast, the simple, straightforward membrane filtration method could perform best when isolating and culturing H. pylori from all water samples. Upon examining the cultivation and qPCR results obtained from groundwater samples, the presence of infectious H. pylori (potentially other pathogens) in aquifers must represent a pressing environmental emergency demanding immediate attention. Furthermore, we believe groundwater can be used as a medium to reflect the H. pylori prevalence in a highly populated community due to its straightforward analytical matrix, consistent detection performance, and minimal interferences from human activities, temperature, precipitation, and other environmental fluctuations.

Shedding of Viral Haemorrhagic Septicaemia Virus (VHSV) from Rainbow Trout, Oncorhynchus mykiss, and Quantification in Waste from Processing Preclinical Fish

Viral haemorrhagic septicaemia virus (VHSV) is a fish disease notifiable to the World Organisation for Animal Health. The United Kingdom is currently free from VHSV, and the introduction and onward spread of this disease could cause major economic losses in aquaculture facilities. Legislation in Great Britain requires that imports of live fish for aquaculture purposes into declared disease-free areas are of equivalent disease-free status. However, conditions on fish products are less stringent, whereby eviscerated fish or fillets can be transported from areas with disease to areas declared disease-free. Market-size rainbow trout were experimentally infected with VHSV to investigate two important factors relevant for pathogen introduction and transmission: (1) VHSV shedding, quantified by daily assessment of viral titres in tank water samples, and (2) VHSV concentrations in liquid and solid processing waste. Evisceration and filleting preclinical fish, maceration, and wastewater separation processes within a facility were mimicked, and VHSV was quantified in each fraction of the wastewater. Shedding was detected 25 hr post-challenge. Levels increased daily to peak on day 5 post-challenge, with a calculated average titre of 1.35 × 103 TCID50 mL−1 kg−1 fish, 1 day before clinical signs of disease. Preclinical fish contained virus levels in their kidney, skin, and muscle >107 TCID50 g−1. The fish had significantly higher levels in the kidney, and evisceration led to higher VHSV concentrations in the waste compared to filleting. However, there was no significant difference in levels in wastewater released from the two processes after the removal of solids, even when macerated; average titres were >104 TCID50 mL−1. The quantities of VHSV from shedding and processing can be utilised when modelling transmission and undertaking more accurate risk assessments for imports and processing of commodities, with the ultimate aim of reducing the global risk of disease from international trade and processing.

Hidden risks: Simulated leakage of domestic sewage to secondary water supply systems poses serious microbiological risks

There are continuous reports about the pollution of the secondary water supply systems (SWSSs), among which domestic sewage leakage is the most serious. In this study, a pilot experiment lasting 70 days was conducted to explore the changes in physicochemical water quality and the microbial profiles in SWSSs polluted by different doses of domestic sewage through qPCR and high-throughput sequencing methods. The results showed that when domestic sewage entered the simulated water storage tank, a large amount of organic matter brought by domestic sewage quickly consumed chlorine disinfectants. High pollution levels (pollution index ≥ 1/1000) were accompanied by significant increases in turbidity and ammonia nitrogen concentration (p < 0.05) and by abnormal changes in sensory properties. Although different microbial community structures were found only at high pollution levels, qPCR results showed that the abundance of the bacterial 16S rRNA gene and some pathogenic gene markers in the polluted tank increased with the pollution level, and the specific gene marker of pathogens could be detected even at imperceptible pollution levels. In particular, the high detection frequency and abundance of Escherichia coli and Enterococcus faecails in polluted tank water samples demonstrated that they can be used for early warning. Moreover, it seems that the microorganisms that came with the domestic sewage lost their cultivability soon after entering SWSSs but could recover their activities during stagnation. In addition, the biofilm biomass in the polluted tank with high pollution levels increased faster at the initial stage, while after a longer contact time, it tended to remain at the same level as the control tank. This study emphasized the high microbial risk introduced by sewage water leakage even at imperceptible levels and could provide scientific suggestions for early warning and prevention of pollution to SWSSs.

A single-center experience with microbiologic surveillance of LivaNova 3T heater-cooler devices (HCDs)

Background: The global outbreak of Mycobacterium chimaera infections associated with HCDs resulted in new maintenance recommendations. Since 2018, HCDs have been disinfected according to instructions for use (IFU), including twice-monthly bleach disinfection and monitoring hydrogen peroxide (H2O2) to maintain a minimum daily concentration of 100 ppm. In February 2020, the IFU added the recommendation to perform microbiologic surveillance of HCD tank water to ensure effectiveness of disinfection to levels of &lt;1 colony forming unit per milliliter (CFU/mL) of nontuberculous mycobacterium (NTM). We report our experience with this microbiologic surveillance as well as that of culturing the HCD environment to investigate modes of transmission. Methods: In 2022, we began culturing tank water in 10 HCDs for NTM. For a subset of 6 HCDs, quantitative NTM culturing of tank water before and after bleach disinfection was done. After initial results indicated widespread-contamination of HCDs with M. chimaera, we performed fill water cultures from 5 sinks in 4 HCD maintenance rooms. We also conducted 20 two-hour NTM settle-plate cultures of a cardiac operating room (OR) at different sites both inside (n = 7) and outside (n = 3) the OR: 10 with the HCD (located outside the OR) turned off (controls) and 10 with HCD turned on (exposure). A paired t test was used to evaluate differences in mean recovery of NTM in tank water samples. Results: Cultures from 7 (70%) of 10 HCDs were positive, with a mean of 13.6 CFU/mL M. chimaera (Table 1). There was no significant difference between the 10 pairs of pre- and postdisinfection NTM cultures done according to the IFU from 6 HCDs: mean predisinfection cultures (15.5 CFU/mL) versus mean postdisinfection cultures (12 CFU/mL) (P = .90) (Table 2). For fill water, 1 of 7 sink samples in 1 of 4 rooms was positive for M. chimaera ( &lt;1 CFU/mL) from a specimen from a fresh 0.2-µm filter that had been stored in the fill-sink splash zone. OR settle-plate cultures showed 0 (0%) of 10 control sites and 1 (10 %) of 10 exposure sites inside the OR positive for NTM, with a single CFU of M. avium–intracellulare complex. Conclusions: Our data cannot clearly refute either of 2 possible scenarios for HCD contamination: cross contamination during device maintenance versus at the point of manufacture. Despite the IFU guidance or disinfection being implemented, disinfection procedures failed to suppress NTM contamination, and tank water within most HCDs was contaminated with M. chimaera regardless of age or whether it was deep cleaned or upgraded with an aerosol containment device.Disclosures: None

To close or open the tank input water valve: Secondary water-supply systems with double tanks will induce a higher microbial risk

Secondary water supply systems (SWSSs) are widely used to supply water to high-rise households in urban residential buildings. A special mode of double tanks with one used while another was spared was noted in SWSSs, which would facilitate microbial growth due to longer water stagnation in the spare tank. There are limited studies on the microbial risk of water samples in such SWSSs. In this study, the input water valves of the operational SWSSs consisting of double tanks were artificially closed and opened on time. Propidium monoazide-qPCR and high-throughput sequencing were performed to systematically investigate the microbial risks in water samples. After closing the tank input water valve, it may take several weeks to replace the bulk water in the spare tank. The residual chlorine concentration in the spare tank decreased by up to 85 % within 2–3 days compared with that in the input water. The microbial communities in the spare and used tank water samples clustered separately. High bacterial 16S rRNA gene abundance and pathogens-like sequences were detected in the spare tanks. Most antibiotic-resistant genes (11/15) in the spare tanks showed an increase in their relative abundance. Moreover, when both tanks within one SWSS were in use, the water quality of the used tank water samples deteriorated to varying degrees. Overall, running SWSSs with double tanks will reduce the replacement rate of water in one storage tank, and consumers who use taps served by the presented SWSSs may have a higher microbial risk.

MLVSS / MLSS ratio’s standard value obtained from different aeration tank samples of different capacity sewage treatment plant - A case study

Waste water treatment system plays a vital role in controlling pollution of natural water bodies like lake, pond, river etc., by Municipal and Industrial effluents. Different industrial effluents play its own role in contaminating the water bodies which in turn creates huge impact for aquatic and terrestrial life. From past studies we understood that, all the sewage treatment systems have a secondary treatment step which are mainly driven by Bacterial oxidation or in other terms can be pronounced as Biological augmentation, Biological calcification etc., In environmental engineering term this bacterial growth will be pronounced as MLSS (Mixed Liquor Suspended Solids) and MLVSS (Mixed Liquor Volatile Suspended Solids). MLVSS will be an inclusive part of the MLSS and also can be sorted as live bacterial cells which can really does the oxidation process in the secondary treatment step of Sewage treatment plant. Hence, this study was performed to evaluate the percentage or concentration of MLVSS available in the total value of MLSS. For this study aeration tank water samples were collected from 6 different STP capacities from 6 different areas. All the samples were tested for MLSS and MLVSS concentration with the available standard method of drying. Drying with 105° C in oven gives the value of MLSS and drying with 550° C in furnace gives the value of MLVSS. With all the tested samples the concentration of MLVSS from the total MLSS was evaluated and standardized.

Evaluating Stress-Mediated Microbial Pathogenesis in Golden Shiners, Notemigonus crysoleucas.

Flavobacterium covae (columnaris) is a microbial pathogen of the Golden Shiner (Notemigonus crysoleucas), a principal bait species. We investigated the effects of density and water temperature on the survival of fish subjected to a columnaris challenge and whether flow cytometry (FCM) could be a fast and reliable method to distinguish and enumerate F. covae populations from water and fish in experimental tanks. Juvenile Golden Shiners averaging 2.62 (±0.78 S.D.) g (negative for F. covae) were used in simultaneous trials at 22°C and 28°C in two ultra-low flow-through systems: each consisting of four treatments and five replicates per treatment. Treatments were fish stocked at either 600 fish/m3 or 2,400 fish/m3 and either challenged with F. covae or not; survival was observed for 48 h after challenge. Samples of water and fish tissue were obtained for FCM enumerations and validation by qPCR. No significant differences in survival were recorded between density treatments; however, high temperature and columnaris challenge treatments showed significantly higher mortality. Bacterial enumeration (number/mL) by FCM highly correlated with bacterial counts r = 0.81 (p = 0.001) in the water samples. Higher water temperatures may have increased columnaris infections and mortality in Golden Shiners. Flow cytometry is a reliable method of enumerating F. covae from experimental tank water samples.

Detection and population genomics of sea turtle species via noninvasive environmental DNA analysis of nesting beach sand tracks and oceanic water.

Elusive aquatic wildlife, such as endangered sea turtles, are difficult to monitor and conserve. As novel molecular and genetic technologies develop, it is possible to adapt and optimize them for wildlife conservation. One such technology is environmental (e)DNA - the detection of DNA shed from organisms into their surrounding environments. We developed species-specific green (Chelonia mydas) and loggerhead (Caretta caretta) sea turtle probe-based qPCR assays, which can detect and quantify sea turtle eDNA in controlled (captive tank water and sand samples) and free ranging (oceanic water samples and nesting beach sand) settings. eDNA detection complemented traditional in-water sea turtle monitoring by enabling detection even when turtles were not visually observed. Furthermore, we report that high throughput shotgun sequencing of eDNA sand samples enabled sea turtle population genetic studies and pathogen monitoring, demonstrating that noninvasive eDNA techniques are viable and efficient alternatives to biological sampling (e.g., biopsies and blood draws). Genetic information was obtained from sand many hours after nesting events, without having to observe or interact with the target individual. This greatly reduces the sampling stress experienced by nesting mothers and emerging hatchlings, and avoids sacrificing viable eggs for genetic analysis. The detection of pathogens from sand indicates significant potential for increased wildlife disease monitoring capacity and viral variant surveillance. Together, these results demonstrate the potential of eDNA approaches to ultimately help understand and conserve threatened species such as sea turtles.

Microbiological quality of roof tank water in an urban village in southeastern China

Urban villages are unique residential neighborhoods in urban areas in China. Roof tanks are their main form of water supply, and water quality deterioration might occur in this system because of poor hygienic conditions and maintenance. In this study, water samples were seasonally collected from an urban village to investigate the influence of roof tanks as an additional water storage device on the variation in the microbial community structure and pathogenic gene markers. Water stagnation in the roof tank induced significant decreases in chlorine (p < 0.05), residual chlorine was as low as 0.02 mg/L in spring. Propidium monoazide (PMA)-qPCR revealed a one-magnitude higher level of total viable bacterial concentration in roof tank water samples (2.14 ± 1.81 × 105 gene copies/mL) than that in input water samples (3.57 ± 2.90 × 104 gene copies/mL, p < 0.05), especially in spring and summer. In addition, pathogenic fungi, Mycobacterium spp., and Legionella spp. were frequently detected in the roof tanks. Terminal users might be exposed to higher microbial risk induced by high abundance of Legionella gene marker. Spearman's rank correlation and redundancy analysis showed that residual chlorine was the driving force that promoted bacterial colonization and shaped the microbial community. It is worth noted that the sediment in the pipe will be agitated when the water supply is restored after the water outages, which can trigger an increase in turbidity and bacterial biomass. Overall, the findings provide practical suggestions for controlling microbiological health risks in roof tanks in urban villages.

A survey on heavy metal concentrations in residential neighborhoods: The influence of secondary water supply systems

Water quality deterioration often occurs in secondary water supply systems (SWSSs), and increased heavy metal concentrations can be a serious problem. In this survey, twelve residential neighborhoods were selected to investigate the influence of SWSSs on the seasonal changes in heavy metal concentrations from input water to tank and tap water. The concentrations of nine evaluated heavy metals in all groups of water samples were found to be far below the specified standard levels in China. The concentrations of Fe, Mn, and Zn increased significantly from the input water samples to the tank and tap water samples in spring and summer (p < 0.05), especially for the water samples that had been stagnant for a long time. Negative correlations were found between most of the heavy metals and residual chlorine (Fe, Cu, Zn, and As, r = -0.186 to -0.519, p < 0.05). In particular, a high negative correlation was observed between Fe and residual chlorine (r = -0.489 to -0.519, p < 0.01) in spring and summer. Fe and Mn displayed positive correlations with turbidity (r = 0.672 and 0.328, respectively; p < 0.05). In addition, Cr and As were found to be positively associated with some nutrients (NO3−, TN, and SO42−; r = 0.420-0.786, p < 0.01). The material of the storage tanks had little influence on the difference in heavy metal concentrations. Overall, this survey illustrated that SWSSs may pose a chronic threat to water quality and could provide useful information for practitioners.

Zebrafish, Danio rerio, Skin Mucus Harbors a Distinct Bacterial Community Dominated by Actinobacteria.

The skin mucus of teleost fish harbors a complex microbial community that is continually interacting with the aquatic environment. Despite zebrafish, Danio rerio, serving as a model organism in a myriad of research fields, very little is known about the composition and role of the skin mucus microbiome. The purpose of this study was to determine a simple sampling method for the skin mucus microbiome, identify prominent bacterial members, and compare its composition to the microbial community of the surrounding environment. Next-generation sequencing of the V3-V4 region of the 16S rRNA gene was performed on skin mucus and filtered tank water samples. Results show that prominent bacterial members of the skin mucus in zebrafish include Actinobacteria (Mycobacteriaceae) and Gammaproteobacteria (Aeromonadaceae), followed by Alphaproteobacteria and Betaproteobacteria. The tank water contained much higher bacterial diversity and was clearly different from the skin mucus microbiome, despite continuous interaction. This study identifies a straightforward sampling method for the zebrafish skin mucus microbiome, enabling hypothesis generation on the role of ectosymbionts on host and microbiome health.

Determining Bacterial Load and Water Quality Parameters of Chlorinated Tomato Flume Tanks in Florida Packinghouses

Monitoring and maintenance of water quality in dump tanks or flume systems is crucial to maintaining proper sanitizer levels to prevent pathogen cross-contamination during postharvest washing of tomatoes, but there is limited information on how organic matter influences sanitizer efficacy in the water. The main objective of this study was to monitor water quality in flume tanks and evaluate the efficacy of postharvest washing of tomatoes in commercial packinghouses. Flume tank water samples (n = 3) were collected on an hourly basis from three packinghouses in Florida and analyzed for pH, total dissolved solids (TDS), free chlorine, chemical oxygen demand (COD), oxidation-reduction potential, and turbidity. Additionally, three flume-water samples were collected and tested for total aerobic plate count (APC), total coliforms (TC), and Escherichia coli. Fresh tomatoes (n = 3), both before and after washing, were collected and analyzed for the same bacterial counts. Turbidity, COD, and TDS levels in flume water increased over time in all packinghouses. Correlations observed include COD and turbidity (r = 0.631), turbidity and TDS (r = 0.810), and oxidation-reduction potential and chlorine (r = 0.660). APC for water samples had an average range of 0.0 to 4.7 log CFU/mL and TC average range of 0.0 to 4.7 log CFU/mL. All water samples were negative for E. coli. The average APC for pre- and postflume tomatoes from the three packinghouses was 6.0 log CFU per tomato and ranged from 2.2 to 7.4 log CFU per tomato. The average TC count was <1.5 and 7.0 log CFU per tomato for pre- and postwash tomatoes, respectively. There was no significant effect (P > 0.05) of postharvest washing on the microbiological qualities of tomatoes. Water quality in flume tanks deteriorated over time in all packinghouses during a typical operational day of 4 to 8 h.

A comprehensive investigation of the microbial risk of secondary water supply systems in residential neighborhoods in a large city

Secondary water supply systems (SWSSs) are characterized by long water stagnation and low levels of chlorine residuals, which may pose a high microbial risk to terminal users. In this study, the SWSSs of 12 residential neighborhoods in a metropolitan area of 5 million people in southeastern China were seasonally investigated to assess their microbial risks by determining more than 30 physicochemical and biological parameters. Although the microbiological quality of SWSS water met the requirements of the standards for drinking water quality of China, it did deteriorate in various aspects. The heterotrophic plate counts with R2A media were high (> 100 CFU/mL) in some SWSS tank and tap water samples. Propidium monoazide (PMA)-qPCR revealed a one magnitude higher abundance of viable bacteria in the tank and tap water samples (average 103.63±1.10 and 103.65±1.25 gene copies/mL, respectively) compared with the input water samples, and Enterococcus, Acanthamoeba, and Hartmannella vermiformis were only detected in the tanks. In particular, the high detection frequency of Legionella in 35% tank and 21% tap water samples suggested it is a supplementary microbial safety indicator in SWSSs. The microbial regrowth potential was more obvious in summer, and Illumina sequencing also demonstrated distinct seasonal changes in the relative abundance of bacterial gene sequences at the genus level. Turbidity and residual chlorine were closely connected with total bacterial biomass, and the latter seemed responsible for microbial community structure alteration. The extremely low chlorine residuals associated with a high abundance of total bacteria (as high as 106.48 gene copies/mL) and Legionella (as high as 106.71 gene copies/100 mL) in the closed valve tanks highlighted the high microbial risk increased by mishandling the operation of SWSSs. This study found that SWSSs possessed a higher microbial risk than the drinking water network, which suggested that the frequency and scope of monitoring the microbial risk of SWSSs in megacities should be strengthened for the purpose of waterborne epidemic disease prevention and control.

Filtration, concentration and detection of salmonid alphavirus in seawater during a post-smolt salmon (Salmo salar) cohabitant challenge

Currently, the prevalence of salmonid alphavirus (SAV) in Norwegian Atlantic salmon farms is largely surveyed via sacrificing fish and sampling of organ tissue on a monthly basis. However, a more cost-efficient, straightforward, rapid, reliable, reproducible and animal welfare friendly method based on the detection of SAV in water could be considered as an alternative method. In the present study, such a method was developed and optimized through a 6 wk cohabitant challenge trial, using post-smolt Atlantic salmon Salmo salar L challenged with high or low doses of SAV subtype 3 (SAV3). Tank water and tissue samples from cohabitant fish were collected at 16 time points. SAV3 was concentrated from the water by filtration, using either electronegative or electropositive membrane filters, which were subsequently rinsed with one of 4 different buffer solutions. SAV3 was detected first in tank water (7 d post-challenge, DPC), and later in cohabitant fish organ tissue samples (12 DPC). The electronegative filter (MF-Millipore™) and rinsing with NucliSENS® easyMAG® Lysis Buffer presented the best SAV3 recovery. A significant positive correlation was found between SAV3 in the tank water concentrates and the mid-kidney samples. Based on these results, detection of SAV3 in filtrated seawater is believed to have the potential to serve as an alternative method for surveillance of SAV in Atlantic salmon farms.

Filtration, concentration and detection of salmonid alphavirus in seawater during a post-smolt salmon (Salmo salar) cohabitant challenge.

Currently, the prevalence of salmonid alphavirus (SAV) in Norwegian Atlantic salmon farms is largely surveyed via sacrificing fish and sampling of organ tissue on a monthly basis. However, a more cost-efficient, straightforward, rapid, reliable, reproducible and animal welfare friendly method based on the detection of SAV in water could be considered as an alternative method. In the present study, such a method was developed and optimized through a 6 wk cohabitant challenge trial, using post-smolt Atlantic salmon Salmo salar L challenged with high or low doses of SAV subtype 3 (SAV3). Tank water and tissue samples from cohabitant fish were collected at 16 time points. SAV3 was concentrated from the water by filtration, using either electronegative or electropositive membrane filters, which were subsequently rinsed with one of 4 different buffer solutions. SAV3 was detected first in tank water (7 d post-challenge, DPC), and later in cohabitant fish organ tissue samples (12 DPC). The electronegative filter (MF-Millipore™) and rinsing with NucliSENS® easyMAG® Lysis Buffer presented the best SAV3 recovery. A significant positive correlation was found between SAV3 in the tank water concentrates and the mid-kidney samples. Based on these results, detection of SAV3 in filtrated seawater is believed to have the potential to serve as an alternative method for surveillance of SAV in Atlantic salmon farms.

The effectiveness of treating irrigation water using ultraviolet radiation or sulphuric acid fertilizer for reducing generic Escherichia coli on fresh produce-a controlled intervention trial.

AimsThe aims of this study were to: (i) estimate the effectiveness of ultraviolet radiation (UV) and sulphuric acid‐based fertilizer (SA), at reducing levels of generic Escherichia coli in surface irrigation water and on produce and surface soil in open produce fields; and (ii) describe the population dynamics of generic E. coli in produce fields.Methods and ResultsSpinach and cantaloupe plots were randomly assigned to control, UV or SA treatment groups. Irrigation water was inoculated with Rifampicin‐resistant E. coli prior to treatment. More than 75% of UV‐ and SA‐treated tank water samples had counts below the detection limit, compared to a mean count of 3·3 Log10 CFU per ml before treatment. Levels of Rifampicin‐resistant E. coli in soil and produce both increased and decreased over 10–15 days after irrigation, depending on the plot and time‐period.ConclusionsUV and SA treatments effectively reduce the levels of E. coli in surface irrigation water. Their effectiveness at reducing contamination on produce was dependent on environmental conditions. Applying wait‐times after irrigation and prior to harvest is not a reliable means of mitigating against contaminated produce.Significance and Impact of the StudyThe results are of timely importance for the agricultural industry as new FSMA guidelines require producers to demonstrate a low microbial load in irrigation water or allow producers to apply a wait‐time to mitigate the risk of contaminated produce.

Messenger RNA typing of environmental RNA (eRNA): A case study on zebrafish tank water with perspectives for the future development of eRNA analysis on aquatic vertebrates

AbstractAs an alternative/supplement to conventional biodiversity survey methods, environmental DNA (eDNA) analysis has developed rapidly during the past decade, and is widely used for the biomonitoring of wildlife. However, the interpretation of eDNA results has been limited to the presence/absence or biomass/abundance of the target species because of uncertainty regarding the dynamics of eDNA in natural environments. This limitation may be ameliorated by targeting environmental RNA (eRNA). RNA is more prone to degrade than DNA and the pattern of messenger RNA (mRNA) expression changes depending on physiological conditions, meaning that the presence or concentration of mRNA could reflect the organism's presence with higher temporal resolution and provide information beyond simple presence/absence. Technical developments in the detection of eRNA focusing on mRNA with these distinct features could permit the advanced usage of genetic materials in water. In advancing this technique, we initiated this study asking that if we can detect elevated levels of eRNA whose genes are specific to a tissue source (e.g., gills or skin), then could not we infer the tissue origin of the genetic material detected. To this end, we developed gene‐specific primer sets for the target genes with biased expression in the gills, skin, and intestine, and conducted reverse transcription–polymerase chain reaction on zebrafish breeding tank water samples, obtaining positive results for all assays. The result of our experiment confirmed that the specific target tissues can be the source of genetic materials detected in water and with that we offer a proof of concept for eRNA analysis targeting specific mRNAs of aquatic vertebrates. In this commentary, we provide information on the experimental steps used for mRNA typing of eRNA from zebrafish as well as the limitations and challenges of this technique and the prospect of mRNA typing of eRNA in the future.

Pharmacokinetic profile of a single dose of an oral pradofloxacin suspension administered to eastern long-necked turtles (Chelodina longicollis).

The pharmacokinetics of fluoroquinolones in chelonians are well described but this does not extend to pradofloxacin, a broad-spectrum veterinary fluoroquinolone available as an oral suspension for cats and dogs. The aim of this study was to investigate the single-dose pharmacokinetic profile of pradofloxacin oral suspension at 7.5mg/kg in eastern long-necked turtles (Chelodina longicollis). Eight treated turtles were sampled at multiple time points up to 168hr. Plasma concentrations were measured using high-performance liquid chromatography. Pradofloxacin was quantifiable for up to 48hr after drug administration. The Tmax (9.0hr) and T½ to 48hr (13.16hr) were longer, and the Cmax (0.2μg/ml) and AUC0-24 (2.2hr*μg/ml) lower, than previously reported in cats and dogs. Pradofloxacin was measurable in tank water samples for up to 48hr. No adverse effects were observed in six turtles administered 7.5mg/kg sid for 7days. Using mammalian MIC data, the AUC0-24 /MIC ratios for a range of bacterial isolates suggest that this dose of pradofloxacin in turtles is unlikely to be effective against many bacterial pathogens.