Molecular and Serological Characterization of the SARS-CoV-2 Delta Variant in Bangladesh in 2021.

Novel SARS-CoV-2 variants are emerging at an alarming rate. The delta variant and other variants of concern (VoC) carry spike (S)-protein mutations, which have the potential to evade protective immunity, to trigger break-through infections after COVID-19 vaccination, and to propagate future waves of COVID-19 pandemic. To identify SARS CoV-2 variants in Bangladesh, patients who are RT-PCR-positive for COVID-19 infections in Dhaka were screened by a RT-PCR melting curve analysis for spike protein mutations. To assess the anti-SARS CoV-2 antibody responses, the levels of the anti-S -proteins IgA and IgG and the anti-N-protein IgG were measured by ELISA. Of a total of 36 RT-PCR positive samples (75%), 27 were identified as delta variants, with one carrying an additional Q677H mutation and two with single nucleotide substitutions at position 23029 (compared to Wuhan-Hu-1 reference NC 045512) in the genome sequence. Three (8.3%) were identified as beta variants, two (5.5%) were identified as alpha variants, three (8.3%) were identified as having a B.1.1.318 lineage, and one sample was identified as an eta variant (B.1.525) carrying an additional V687L mutation. The trend of higher viral load (lower Cp values) among delta variants than in the alpha and beta variants was of borderline statistical significance (p = 0.045). Prospective studies with larger Bangladeshi cohorts are warranted to confirm the emergence of S-protein mutations and their association with antibody response in natural infection and potential breakthrough in vaccinated subjects.

Carbon Nanotube Integrative Sampler (CNIS) for passive sampling of nanosilver in the aquatic environment.

Nanomaterials such as nanosilver (AgNP) can be released into the aquatic environment through production, usage, and disposal. Sensitive and cost-effective methods are needed to monitor AgNPs in the environment. This work is hampered by a lack of sensitive methods to detect nanomaterials in environmental matrixes. The present study focused on the development, calibration and application of a passive sampling technique for detecting AgNPs in aquatic matrixes. A Carbon Nanotube Integrative Sampler (CNIS) was developed using multi-walled carbon nanotubes (CNTs) as the sorbent for accumulating AgNPs and other Ag species from water. Sampling rates were determined in the laboratory for different sampler configurations and in different aquatic matrixes. The sampler was field tested at the Experimental Lakes Area, Canada, in lake water dosed with AgNPs. For a configuration of the CNIS consisting of CNTs bound to carbon fiber (i.e. CNT veil) placed in Chemcatcher® housing, the time weighted average (TWA) concentrations of silver estimated from deployments of the sampler in lake mesocosms dosed with AgNPs were similar to the measured concentrations of "colloidal silver" (i.e. <0.22µm in size) in the water column. For a configuration of CNIS consisting of CNTs in loose powder form placed in a custom made housing that were deployed in a whole lake dosed with AgNPs, the estimated TWA concentrations of "CNIS-labile Ag" were similar to the concentrations of total silver measured in the epilimnion of the lake. However, sampling rates for the CNIS in various matrixes are relatively low (i.e. 1-20mL/day), so deployment periods of several weeks are required to detect AgNPs at environmentally relevant concentrations, which can allow biofilms to develop on the sampler and could affect the sampling rates. With further development, this novel sampler may provide a simple and sensitive method for screening for the presence of AgNPs in surface waters.

The impact of municipal wastewater effluent on field-deployed freshwater mussels in the Grand River (Ontario, Canada).

To examine effects of municipal wastewater effluent (MWWE) on sentinel organisms, the authors deployed caged freshwater mussels (Lasmigona costata) in the Grand River (ON, Canada) upstream and downstream of an MWWE outfall. Passive sampling devices were deployed alongside caged mussels to confirm exposure. Biomarkers of xenobiotic biotransformation, oxidative stress, estrogenicity, and immunomodulation were investigated. Elevated concentrations of selected pharmaceutical and personal care products (PPCPs) and a natural estrogen (estrone) were found at the downstream sites. Mussels caged downstream of the effluent for 2 wk showed minimal evidence of exposure, while those deployed for 4 wk exhibited significantly higher levels of lipid peroxidation (LPO) and glutathione S-transferase (GST) activity, demonstrating that MWWE-exposed mussels exhibit increased activity in xenobiotic conjugation and oxidative stress. With respect to immune responses, a significant increase in plasma lysozyme activity and hemocyte viability was observed in MWWE-exposed mussels. Vitellogenin (vtg)-like protein in male mussels showed a trend toward induction after 4 wk of deployment at the first downstream site, but mean levels were not significantly different. Discriminant function analysis indicated that mussels deployed for 4 wk upstream and downstream of the MWWE discharge could be discriminated on the basis of LPO, GST, plasma lysozyme, and vtg responses. The physiological stress observed in caged mussels indicates that wild mussels chronically exposed to MWWE in this ecosystem would also be negatively impacted.

Changes in bacterial community structure after exposure to silver nanoparticles in natural waters.

Silver nanoparticles (AgNPs) are widely used in commercial products as antibacterial agents, but AgNPs might be hazardous to the environment and natural aquatic bacterial communities. Our recent research demonstrated that AgNPs rapidly but temporarily inhibited natural bacterioplankton production. The current study investigates the mechanism for the observed bacterial reaction to AgNPs by examining how AgNPs impact bacterial abundance, metabolic activity (5-cyano-2,3-ditolyl tetrazolium chloride (CTC+) cells), and 16S rRNA community composition. Natural bacterioplankton communities were dosed with carboxy-functionalized AgNPs at four concentrations (0.01-1 mg-Ag/L), incubated in triplicate, and monitored over 5 days. Ionic silver (AgNO(3)) and Milli-Q water treatments were used as a positive and negative control, respectively. Four general AgNP exposure responses, relative to the negative control, were observed: (1) intolerant, (2) impacted but recovering, (3) tolerant, and (4) stimulated phylotypes. Relationships between cell activity indicators and bacterial phylotypes, suggested that tolerant and recovering bacteria contributed the most to the community's productivity and rare bacteria phylotypes stimulated by AgNPs did not appear to contribute much to cell activity. Overall, natural bacterial communities tolerated single, low level AgNP doses and had similar activity levels to the negative control within five days of exposure, but bacterial community composition was different from that of the control.

Depth-profiling of environmental pharmaceuticals in biological tissue by solid-phase microextraction.

The parallel in vivo measurement of chemicals at various locations in living tissues is an important approach furthering our understanding of biological uptake, transportation, and transformation dynamics. However, from a technical perspective, such measurements are difficult to perform with traditional in vivo sampling techniques, especially in freely moving organisms such as fish. These technical challenges can be well addressed by the proposed depth-profiling solid-phase microextraction (DP-SPME) technique, which utilizes a single soft, flexible fiber with high spatial resolution. The analytical accuracy and depth-profiling capability of DP-SPME was established in vitro within a multilayer gel system and an onion artificially contaminated with pharmaceuticals. In vivo efficacy was demonstrated by monitoring pharmaceutical distribution and accumulation in fish muscle tissue. The DP-SPME method was validated against pre-equilibrium SPME (using multiple small fibers), equilibrium SPME, and liquid extraction methods; results indicated DP-SPME significantly improved precision and data quality due to decreased intersample variation. No significant adverse effects or increases in mortality were observed in comparisons of fish sampled by DP-SPME relative to comparable fish not sampled by this method. Consequently, the simplicity, effectiveness, and improved precision of the technique suggest the potential for widespread application of DP-SPME in the sampling of heterogeneous biotic and abiotic systems.

Detection and characterization of silver nanoparticles in aqueous matrices using asymmetric-flow field flow fractionation with inductively coupled plasma mass spectrometry.

Engineered nanomaterials (EN) may be released into the environment as a result of their use in various consumer products. Silver nanoparticles (nAg) are widely used as an antimicrobial agent in personal care and household products, and in textiles. Since there is high potential for nAg to be released into municipal wastewater and then discharged into the aquatic environment, there is a need to develop methods for the analysis of these materials in aqueous matrices. Asymmetric-flow field flow fractionation (AF4) with on-line detection by ultra violet-visible (UV-Vis) spectroscopy or inductively coupled plasma mass spectrometry (ICP-MS) was used to detect and characterize nAg in aqueous matrices. Analysis of a mixture of 20, 40 and 60 nm nAg standards suspended in water resulted in a well resolved fractogram. Retention times of nAg separated by AF4 were correlated with the particle sizes of the standards. The limit of detection (LOD) for analysis of nAg using the on-line AF4/ICP-MS method was 0.80 ng mL(-1). Two calibration approaches (i.e., external calibration and standard addition) were used to quantify nAg concentrations, and both methods gave similar results. Using the on-line AF4/ICP-MS analytical method, nano-sized Ag was detected and quantified in untreated wastewater (i.e., influent) collected from a wastewater treatment plant. The concentration and the modal size of nAg in the influent were 1.90 ng mL(-1) and 9.3 nm respectively.

Effects of silver nanoparticles on bacterial activity in natural waters.

Silver nanoparticles (AgNPs) may be introduced into aquatic ecosystems because of their widespread use as antimicrobial agents. However, few studies have investigated the impacts of AgNPs on natural aquatic microbial activity in an environmentally relevant context. In this study, bacterioplankton were collected from nine aquatic habitats and exposed to six concentrations of carboxy-functionalized AgNP (ViveNano, 10-nm particle size, 20% Ag w/w) over 48 h. After 1 h of exposure, bacterial production and extracellular alkaline phosphatase affinity were significantly reduced in all AgNP-exposed samples. However, across a 48-h exposure, extracellular aminopeptidase affinity was not consistently impacted by AgNPs. After 48 h, bacterial production recovered by 40 to 250% at low AgNP nominal concentrations (0.05 and 0.1 mg/L) but remained inhibited at the two highest AgNP nominal concentrations (1 and 10 mg/L). In contrast, AgNO(3) additions between 0.01 to 2 mg Ag/L fully inhibited bacterial production over the 48-h exposure. At 48-h exposure, the lowest observed effective concentrations and average median effective concentration for bacterial production ranged from 8 to 66 and 15 to 276 µg Ag/L, respectively. Thus, in natural aquatic systems, AgNP concentrations in the nanogram per liter range are unlikely to negatively impact aquatic biogeochemical cycles. Instead, exposures in the low microgram per liter range would likely be required to negatively impact natural aquatic bacterioplankton processes.

Determination of tranexamic acid concentration by solid phase microextraction and liquid chromatography-tandem mass spectrometry: first step to in vivo analysis.

A solid phase microextraction (SPME) method followed by LC-MS/MS analysis was developed to determine the concentration of tranexamic acid (TA) in plasma. The use of a new biocompatible C18 coating allowed the direct extraction from complex biological samples without prior sample preparation; no matrix effect was observed. The results revealed that SPME was suitable for the analysis of polar drugs such as TA; such an application was previously inaccessible because of the limited availability of SPME coatings that can extract polar molecules. The proposed method was validated according to the bioanalytical method validation guidelines. LOD and LLOQ were 0.5 and 1.5 µg/ml, respectively. The recovery for the method was 0.19%, and the accuracy and precision of the method were <9 and <11%, respectively, with the exception of LLOQ, where the values were <16 and <13%, respectively. The performance of the proposed method was also compared against that of the standard techniques of protein precipitation and ultrafiltration. A statistical analysis indicated a clinically significant agreement among all assays. Another advantage of SPME over conventional techniques was the easy automation and feasibility of in vivo analysis; this advantage makes it possible to use the proposed method for an on-site analysis in clinical practice.

Pre-equilibrium solid-phase microextraction of free analyte in complex samples: correction for mass transfer variation from protein binding and matrix tortuosity.

The accurate measurement of free analyte concentrations within complex sample matrixes by pre-equilibrium solid-phase microextraction (SPME) has proven challenging due to variations in mass uptake kinetics. For the first time, the effects of the sample binding matrix and tortuosity on the kinetics of analyte extraction (from the sample to the SPME fiber) are demonstrated to be quantitatively symmetrical with those of the desorption of preloaded deuterated standards (from the fiber to the sample matrix). Consequently, kinetic calibration methods can be employed to correct for variation in SPME sampling kinetics, facilitating the application of pre-equilibrium SPME within complex sample systems. This approach was applied ex vivo to measure pharmaceuticals in fish muscle tissues, with results consistent with those obtained from equilibrium SPME and microdialysis. The developed method has the inherent advantages of being more accurate, precise, and reproducible, thus providing the framework for applications where rapid measurement of free analyte concentrations (within complicated sample matrixes such as biological tissues, sediment, and surface water) are required.