A comparison of precipitation and filtration-based SARS-CoV-2 recovery methods and the influence of temperature, turbidity, and surfactant load in urban wastewater

Wastewater-based epidemiology (WBE) has become a complimentary surveillance tool during the SARS-CoV-2 pandemic. Viral concentration methods from wastewater are still being optimised and compared, whilst viral recovery under different wastewater characteristics and storage temperatures remains poorly understood. Using urban wastewater samples, we tested three viral concentration methods; polyethylene glycol precipitation (PEG), ammonium sulphate precipitation (AS), and CP select™ InnovaPrep® (IP) ultrafiltration. We found no major difference in SARS-CoV-2 and faecal indicator virus (crAssphage) recovery from wastewater samples (n = 46) using these methods, PEG slightly (albeit non-significantly), outperformed AS and IP for SARS-CoV-2 detection, as a higher genome copies per litre (gc/l) was recorded for a larger proportion of samples. Next generation sequencing of 8 paired samples revealed non-significant differences in the quality of data between AS and IP, though IP data quality was slightly better and less variable. A controlled experiment assessed the impact of wastewater suspended solids (turbidity; 0–400 NTU), surfactant load (0–200 mg/l), and storage temperature (5–20 °C) on viral recovery using the AS and IP methods. SARS-CoV-2 recoveries were >20% with AS and  0.05), whilst surfactant and storage temperature combined were significant negative correlates (p < 0.001 and p < 0.05, respectively). In conclusion, our results show that choice of methodology had small effect on viral recovery of SARS-CoV-2 and crAssphage in wastewater samples within this study. In contrast, sample turbidity, storage temperature, and surfactant load did affect viral recovery, highlighting the need for careful consideration of the viral concentration methodology used when working with wastewater samples

Shedding of SARS-CoV-2 in feces and urine and its potential role in person-to-person transmission and the environment-based spread of COVID-19

The recent detection of SARS-CoV-2 RNA in feces has led to speculation that it can be transmitted via the fecal-oral/ocular route. This review aims to critically evaluate the incidence of gastrointestinal (GI) symptoms, the quantity and infectivity of SARS-CoV-2 in feces and urine, and whether these pose an infection risk in sanitary settings, sewage networks, wastewater treatment plants, and the wider environment (e.g. rivers, lakes and marine waters). A review of 48 independent studies revealed that severe GI dysfunction is only evident in a small number of COVID-19 cases, with 11 ± 2% exhibiting diarrhea and 12 ± 3% exhibiting vomiting and nausea. In addition to these cases, SARS-CoV-2 RNA can be detected in feces from some asymptomatic, mildly- and pre-symptomatic individuals. Fecal shedding of the virus peaks in the symptomatic period and can persist for several weeks, but with declining abundances in the post-symptomatic phase. SARS-CoV-2 RNA is occasionally detected in urine, but reports in fecal samples are more frequent. The abundance of the virus genetic material in both urine (ca. 102–105 gc/ml) and feces (ca. 102–107 gc/ml) is much lower than in nasopharyngeal fluids (ca. 105–1011 gc/ml). There is strong evidence of multiplication of SARS-CoV-2 in the gut and infectious virus has occasionally been recovered from both urine and stool samples. The level and infectious capability of SARS-CoV-2 in vomit remain unknown. In comparison to enteric viruses transmitted via the fecal-oral route (e.g. norovirus, adenovirus), the likelihood of SARS-CoV-2 being transmitted via feces or urine appears much lower due to the lower relative amounts of virus present in feces/urine. The biggest risk of transmission will occur in clinical and care home settings where secondary handling of people and urine/fecal matter occurs. In addition, while SARS-CoV-2 RNA genetic material can be detected by in wastewater, this signal is greatly reduced by conventional treatment. Our analysis also suggests the likelihood of infection due to contact with sewage-contaminated water (e.g. swimming, surfing, angling) or food (e.g. salads, shellfish) is extremely low or negligible based on very low predicted abundances and limited environmental survival of SARS-CoV-2. These conclusions are corroborated by the fact that tens of million cases of COVID-19 have occurred globally, but exposure to feces or wastewater has never been implicated as a transmission vector

Thermoresponsive Micropatterned Substrates for Single Cell Studies

We describe the design of micropatterned surfaces for single cell studies, based on thermoresponsive polymer brushes. We show that brushes made of poly(N-isopropylacrylamide) grafted at high surface density display excellent protein and cell anti-adhesive properties. Such brushes are readily patterned at the micron scale via deep UV photolithography. A proper choice of the adhesive pattern shapes, combined with the temperature-dependent swelling properties of PNIPAM, allow us to use the polymer brush as a microactuator which induces cell detachment when the temperature is reduced below C

Ionic liquids at electrified interfaces

Until recently, “room-temperature” (<100–150 °C) liquid-state electrochemistry was mostly electrochemistry of diluted electrolytes(1)–(4) where dissolved salt ions were surrounded by a considerable amount of solvent molecules. Highly concentrated liquid electrolytes were mostly considered in the narrow (albeit important) niche of high-temperature electrochemistry of molten inorganic salts(5-9) and in the even narrower niche of “first-generation” room temperature ionic liquids, RTILs (such as chloro-aluminates and alkylammonium nitrates).(10-14) The situation has changed dramatically in the 2000s after the discovery of new moisture- and temperature-stable RTILs.(15, 16) These days, the “later generation” RTILs attracted wide attention within the electrochemical community.(17-31) Indeed, RTILs, as a class of compounds, possess a unique combination of properties (high charge density, electrochemical stability, low/negligible volatility, tunable polarity, etc.) that make them very attractive substances from fundamental and application points of view.(32-38) Most importantly, they can mix with each other in “cocktails” of one’s choice to acquire the desired properties (e.g., wider temperature range of the liquid phase(39, 40)) and can serve as almost “universal” solvents.(37, 41, 42) It is worth noting here one of the advantages of RTILs as compared to their high-temperature molten salt (HTMS)(43) “sister-systems”.(44) In RTILs the dissolved molecules are not imbedded in a harsh high temperature environment which could be destructive for many classes of fragile (organic) molecules

Hemolymph microbiome of Pacific oysters in response to temperature, temperature stress and infection

Microbiota provide their hosts with a range of beneficial services, including defense from external pathogens. However, host-associated microbial communities themselves can act as a source of opportunistic pathogens depending on the environment. Marine poikilotherms and their microbiota are strongly influenced by temperature, but experimental studies exploring how temperature affects the interactions between both parties are rare. To assess the effects of temperature, temperature stress and infection on diversity, composition and dynamics of the hemolymph microbiota of Pacific oysters (Crassostrea gigas), we conducted an experiment in a fully-crossed, three-factorial design, in which the temperature acclimated oysters (8 or 22 °C) were exposed to temperature stress and to experimental challenge with a virulent Vibrio sp. Strain. We monitored oyster survival and repeatedly collected hemolymph of dead and alive animals to determine the microbiome composition by 16s rRNA gene amplicon pyrosequencing. We found that the microbial dynamics and composition of communities in healthy animals (including infection survivors) were significantly affected by temperature and temperature stress, but not by infection. The response was mediated by changes in the incidence and abundance of operational taxonomic units (OTUs) and accompanied by little change at higher taxonomic levels, indicating dynamic stability of the hemolymph microbiome. Dead and moribund oysters, on the contrary, displayed signs of community structure disruption, characterized by very low diversity and proliferation of few OTUs. We can therefore link short-term responses of host-associated microbial communities to abiotic and biotic factors and assess the potential feedback between microbiota dynamics and host survival during disease