Tracing microplastic pollution in Mahi River estuary, Gulf of Khambhat, Gujarat, and their influence on functional traits of macrobenthos.

Most maritime habitats contain microplastic (MPs) contamination. The quality of the benthic ecosystem's habitat is declining as MPs accumulate in marine system. The contamination of MPs must therefore be investigated. We studied MPs pollution in the Mahi River, estuary, and macrobenthos. In the present study, the abundance of MPs fragments gradually decreased from the high tide zone to the low tide zone and muddy sediment has high MPs concentrations due to sediment characteristics and particle size. The majority of sediment and biota MPs were fibrous and black. MPs in both silt and biota have identical chemical compositions (modified cellulose), shapes, and colors. A significant source of pollutants and MPs fluxing into the ocean is well within the river system. Perinereis aibuhitensis ingested the most MPs out of 11 species, whereas Amphipods did not show any presence of MPs. Our findings showed that functional characteristics are essential for macrobenthos MPs intake. MPs in macrobenthos are high due to biological functions such as feeding, ecological groups, feeding mechanisms, body size, and bioturbation. MPs in marine sediment and organisms are tracked down to the Mahi River exceeding 50 km. The present work has investigated the idea that the macrobenthos that live in the sediment are ingesting the MPs that are building up there and this ingestion relies on the macrobenthos' functional characteristics.

Development of a laboratory-based model to study the interaction between nutrients and Vibrio cholerae and predicting the spread of cholera outbreaks in the Indian subcontinent.

Cholera is an infectious disease that is transmitted through contaminated water. The disease includes a long back history of epidemics. Despite the numerous hygiene and prevention techniques that have been developed for Cholera, outbreaks of cholera are still reported worldwide. The resolution to this issue lies in promptly identifying the area susceptible to cholera outbreaks, a matter that continues to perplex scientists and medical professionals. It has been reported that Vibrio is effective in nitrogen digestion because it contains the nasA gene. In this study, initially the impact of nutrients (nitrate and nitrite) on growth of Vibrio cholerae was determined, subsequently a relationship was developed between nutrient substrates and V. cholerae growth rate, using Monod model. Subsequently, the model was applied to large national river quality data set (2012-2014) developed by Central Pollution Control Board (CPCB) and a possible cholerae outbreak zone was predicted. This work will definitely help the policy makers to develop management strategy for keeping rivers safe from future cholera outbreak.

Market-ready U-AST kit: simple, fast, cost-effective solution for concurrently detecting urinary tract infection and antibiotic resistance.

There is an increasing demand for an inexpensive, quick, accessible, and simple method for the detection of urinary tract infection (UTI) together with the antibiotic-resistance profile of the infection-causing bacteria. Our primary goal is to assist doctors in prescribing antibiotics that will quickly treat infections and reduce the likelihood of antibiotic resistance spreading throughout the community. To this end, a urinary tract infection antibiotic-sensitivity test (U-AST) kit was developed for the validation of bacterial infection in the urinary tract and determination of the antibiotic-resistance profile of the bacteria in a short time. The U-AST kit was standardized using standard strains of bacteria, specifically Escherichia coli, Enterococcus faecalis, Pseudomonas aeruginosa, Vibrio cholerae, and Pseudomonas species. Further, the kit was validated using 50 clinical urine samples with variation in their physical and chemical parameters, and the resistance pattern against five therapeutically important antibiotics were tested. The results acquired using the U-AST kit showed a 100% similarity to those acquired using the laboratory-based gold standard method. Interestingly, the U-AST kit required a maximum of 9 h to understand the bacterial contamination and resistance profile of the bacterial community, which was observed by a simple color change. The same result can be obtained using the gold standard method but requires 36-72 h, a sophisticated microbiology method, and skilled microbiologists. Other methods can also predict infection quickly with the aid of sophisticated instrumentation; however, understanding the antibiotic-resistance pattern is not possible. To the best of our understanding, this is a unique technique for the quick, easy, and inexpensive detection of UTI with antibiotic sensitivity testing and does not require a special laboratory set-up or expert personnel. The commercialization of the developed clinically validated U-AST kit is currently underway.

Time series (2003-15) analysis of selected physicochemical parameters in Indian Ocean: Cumulative impacts prediction on coral bleaching using machine learning.

Coral bleaching is an important ecological threat worldwide, as the coral ecosystem supports a rich marine biodiversity to survive. Sea surface temperature was considered a major culprit; however, later it was observed that other water parameters like pH, tCO2, fCO2, salinity, dissolved oxygen, etc. also play a significant role in bleaching. In the present study, all these parameters of the Indian Ocean area for 15 years (2003-2017) were collected and analysed using machine learning language. The main aim is to see the cumulative impacts of various ocean parameters on coral bleaching. Introducing machine learning in environmental impact assessment studies is a new approach, and the prediction of coral bleaching using simulation of physico-chemical parameters interactions shows 94.4 % accuracy for the prediction of the future bleaching event. This study can be probably the first step in the application of the machine learning language for the prediction of coral bleaching in the field of marine science.

Evaluation of a synbiotic formulation for water remediation in a shrimp pond.

In recent years, the use of probiotic bacteria has attracted the interest of the marine shrimp farming industry. However, there are certain limitations pertaining to the practical application of many commercially available probiotics. Here, a thoroughly screened optimal consortium of three indigenous sulfur probiotics was tested for antibiotic susceptibility and was found to be safe, with each culture being sensitive to all the tested antibiotics. Further, de-potash vinasse (DPV), an environmental hazard, was tested for its prebiotic potential, and its 1% (w/v) concentration was found to be effective for long-term viability (> 66 days) of the probiotic cultures and safe for Artemia. The synbiotic formulation was tested first in a lab-scale microcosm setup successfully and subsequently tried on a shrimp farm; it was observed that the product was congruent to the efficiency of a commercial probiotic regarding almost all physicochemical parameters, sulfide, nitrate-N, nitrite-N, phytoplankton sustenance, Pseudomonas count, coliform count, and heterotrophic count. In addition, it was significantly efficient in maintaining pH, reducing ammonia-N and phosphate-P, Vibrio and Aeromonas count, and a net increase in the yield of shrimp biomass by 625 kg, thus proving to be a better alternative than one of the already available remediation methods.

Interaction of physico-chemical parameters with Shannon-Weaver Diversity Index based on phytoplankton diversity in coastal water of Diu, India.

Phytoplankton acts as carbon sinks due to photosynthetic efficacy and their diversity is expressed by SWDI (Shannon-Weaver Diversity Index), which depends on water quality parameters. The coastal water of Diu was studied for three seasons, and the relationship between different parameters and SWDI was established. Subsequently, an attempt was made to build up a prediction model of SWDI based on multilayer perceptron Artificial neural network (ANN) using the R programme. Analysis shows interrelationship between the water quality parameters and phytoplankton diversity is same in linear principal component analysis (PCA) and neural network model. Variations of different parameters depend on seasonal changes. The ANN model shows that ammonia and phosphate are key parameters that influence the SWDI of phytoplankton. Seasonal variation in SWDI is related to variation in water quality parameters, as explained by both ANN and PCA. Hence, the ANN model can be an important tool for coastal environmental interaction study.

Pre-to-post Diwali air quality assessment and particulate matter characterization of a western coastal place in India.

Diwali has become an occasion of air and noise pollution, and the release of particulate matter and toxic gases has chronic and acute effects on people and their environment. Thus, an air quality assessment study was done by CSIR-CSMCRI covering the pre-to-post Diwali 2021 period (5 days) in the three locations (traffic, residential, and control) of Bhavnagar. The average 24-h concentration of PM10 (380 µg/m3), PM2.5 (182.2 µg/m3), and SPM (403 µg/m3) was significantly higher during Diwali, exceeding the National Ambient Air Quality Standards (NAAQS). The concentrations of SO2 and NO2 were 121.8 µg/m3 and 102.1 µg/m3. Metals like Zn, Al, Pb, and Mn were found in higher concentrations during the study. The air quality index (AQI) was maximum on Diwali, resembling very poor air quality. More elements and oxides were detected in PM2.5 (S, Al, Mg, Ba, and Zn and their oxides) than in PM10 (Fe and S) through WDXRF. Water-soluble anions like SO42-, Cl-, and NO3- were observed during the study, with a higher SO42- (64%) on Diwali. The PM10 morphology and mapping of elements were done using SEM-EDX. Emerging contaminants, specifically phthalate groups, were detected through GCMS. The enrichment factor (EF) showed Zn and Pb originating from anthropogenic activities. The air quality data was validated using a variance test, least significance difference (LSD), correlation, and principal component analysis (PCA). This paper is the first to highlight the air quality assessment during Diwali for a western coastal place in India. It is time to implement regulations on burning firecrackers for pollution reduction, aiming to achieve a sustainable atmosphere.

Determination of antibiotic resistance profile of bacterial community from environmental water using antibiotic-resistant bacterial contamination detection (ABCD) kit.

Quick and easy monitoring approaches are needed to assess sources, clinical relevance, and incidences of antibiotic resistance in environmental bacterial communities. In the present work, antibiotic-resistant bacterial contamination detection (ABCD) kits were developed for the same. The method was standardized with strains of Escherichia coli TS7, Staphylococcus arlettae HWI8, Enterococcus faecalis HWI19, and Aerococcus viridans HWII16 with known antibiogram using six clinically important antibiotics. The method was verified with different water sources having different physicochemical parameters successfully. Only 1 ml of sample water is required to be mixed with an optimized concentration of the antibiotic solution and incubated for 6h; subsequently, a color change to pink may be observed within a specified amount of time upon the addition of the bacterial detection PVDF membrane to monitor the presence of resistant bacteria. There is no color change in the case of antibiotic-susceptible bacterial communities or the absence of a resistant community. Moreover, the time taken for color change is inversely related to the magnitude of the antibiotic-resistant communities in terms of enumeration. Up to our understanding, this is the 1st report which can determine an antibiotic-resistance profile of any water source by observing only color change within a maximum of 7 h (6 h for co-culture of bacteria and antibiotics + 1 h for color change detection) time without the aid of any microbiology laboratory or skilled manpower.

Monitoring and assessment of Dracaena-based constructed vertical flow wetlands treating textile dye wastewater.

The monitoring and assessment of multiple constructed vertical flow wetlands (CVFWs) treating textile dye wastewater (metanil yellow as dye) are studied covering three seasons. Three CVFWs (CVFW-1, dye-5 mg/l; CVFW-2, dye-50 mg/l; and CVFW-3, dye-100 mg/l) and a control (dye-5 mg/l) were used. The CVFWs with Dracaena (an ornamental plant) efficiently removed contaminants like dye, COD, NH4+-N, and PO43--P from the wastewater under varying inlet dye concentrations, indicating its dependence on meteorological conditions. Substantial dye removal was observed to be maximum in summer (control, 44.3%; CVFW-1, 75.1%; CVFW-2, 76.1%; CVFW-3, 46%), but lesser in winter (control, 45%; CVFW-1, 73.1%; CVFW-2, 76.8%; CVFW-3, 42.6%) and minimum in monsoon (control, 40.8%; CVFW-1, 63.5%; CVFW-2, 51.6%; CVFW-3, 37.1%), respectively. Efficiency was less in CVFW-3 as it observed plant stress due to higher inlet dye concentration. COD removal was higher in winter, followed by summer and monsoon. A first-order kinetic model was used to investigate the efficiency of the CVFW system w.r.t. contaminant removal. Various functional groups were characterized using Fourier transform infrared spectroscopy (FTIR) from the inlet and outlet water samples of different CVFWs. The Dracaena accumulated various elements and oxides during the treatment with no stress on its health. No effects on plant health highlight the suitability of Dracaena for textile wastewater treatment. The results were validated using statistical tools like the Mann-Whitney U test and principal component analysis (PCA).

A novel technique for the preparation of curd (Dahi) by PVDF membrane-based strip.

Preparation of curd at home is quite challenging as it requires skills like addition of proper amount of starter culture, maintenance and storage of inoculum for the preparation of good and consistent quality curd. The present work utilized bacterial attachment property of PVDF (Polyvinylidene fluoride) membrane to prepare a strip which can be dipped into milk for the preparation of consistent quality curd. Shelf-life of the strip is around 100 days. The strip prepared curd was well comparable with the curd prepared by the commercial inoculum based on their pH, % lactic acid, % syneresis and bacterial load. Strip of size 5 × 5 cm2 was enough for preparation of 500 mL curd. It was proved by different analytical techniques like AFM, SEM and FTIR that PVDF was not having any leaching property during curd preparation. It can also be used in repeated contact with food products, as it is FDA (Food and Drug Administration) compliant and non-toxic. The curd strip has significant industrial relevance as it is a cost-effective alternative of any commercial inoculum (very expensive) and also meets the demand of consumers with the rising health awareness and busy lifestyles. Further, it is spillage proof, portable, ready-to-use. Supplementary Information: The online version contains supplementary material available at 10.1007/s13197-021-05339-3.

Identification and determination of optimum growth condition with respect to selected environmental parameters for open sea cultivation of Gracillaria edulis in Andaman water.

Gracilaria edulis is one of India's most widely cultivated seaweeds. Pilot scale cultivation of Gracilaria edulis was initiated at Andaman, India for the first time. In the present study attempt has been made to identify how different water quality parameters influence the growth. Total 11 physicochemical parameters and 9 microbiological parameters, as well as chlorophyll and zooplankton, phytoplankton parameters were studied for two different seasons to evaluate which parameters influence seaweed growth. Six (nitrate, nitrite, ammonia, silicate, chlorophyll, photosynthetic active radiation) have a positive impact on seaweed growth, while some of the bacterial species showed negative impact. Lowess 3D curve fit model showed pH range from 7.59 to 7.82, N/P ratio of 2.046, rainfall 23.85-24 mm, and Photosynthetic active radiation of 376.6 W/m2 are optimum for Gracillaria growth. This model can be applied to future mass culture.

Pre-to-post COVID-19 lockdown and their environmental impacts on Ghoghla beach and Somnath beach, India.

Environmental impact of COVID-19 imposed lockdown (2020) and the new normal condition (2021) on two different beaches of India (Ghoghla beach, Diu and Somnath beach, Veraval) were compared with the pre-lockdown era, 2013. The lockdown phase favored the natural restoration of the beaches and showed infinitesimal values of the parameters tested when compared with the pre-lockdown regime. However, the new normal situation in 2021 opened up the accessibility of these beaches to the tourists and pilgrims resulting in significant changes of water quality. The release of diluted sewage mixed with freshwater from the Somnath town to the sea has led to the drastic change in beach water quality. The mean cadmium concentration increased drastically in beach waters (Ghoghla: 1.35, 0.28 and 7.09 µg/L; Somnath: 0.45, 0.28 and 0.58 µg/L) during pre-to-post lockdown, respectively. However, post-lockdown resulted in the rise of toxic heavy metals in the sediments of Somnath beach but Ghoghla beach remained to be pristine which may be due to the Blue Flagship status. The total number of marine bacteria was higher during 2013 and 2021 when compared during lockdown describing greater human interventions. For instance, Vibrio spp. count in Ghoghla beach water during pre-lockdown phase was 7733 CFU/mL and this value reduced to 70 and 5 CFU/mL in the lockdown and post-lockdown phases. Interestingly, the diversity of planktonic and benthic components showed a different trend from pre-to-post lockdown due to significant change in the inorganic nutrients and metal bioaccumulation. To our knowledge, this will be the first comprehensive assessment to report the environmental and ecological health of Ghoghla beach and Somnath beach during the pre-to-post lockdown.

Isolation and screening of sulfur-oxidizing bacteria from coast of Bhavnagar, India, and formulation of consortium for bioremediation.

Reduced sulfur compounds are a nuisance in coastal industries causing heavy economical as well as ecological loss. One such compound, hydrogen sulfide, is proven toxic to aquatic animals as it interferes with their respiration and metabolism as well as overall development, thereby causing direct increase in mortality. Typically, 96-h LC50 values to freshwater and marine fishes are 0-25µM and 525-700µM, respectively. Management of sulfide and other reduced sulfur compounds from aquaculture water and sediment using bioremediating sulfur-oxidizing bacteria as probiotics has attracted attention in recent decades due to its efficiency and minimized environmental effects. In the present study, 201 native and indigenous probiotic candidates were isolated, from various coastal environments. The prospective candidates were screened based on pH reduction and 19 sulfur-oxidizing bacteria were selected and tested for salt tolerance. Further screening was done based on biosafety, ability to produce sulfate by oxidizing thiosulfate, and 16S rRNA-based identification to obtain nine probiotic candidates. Three strains (Enterobacter ludwigii HS1-SOB, Pseudomonas stutzeri B6-SOB, and Cytobacillus firmus C8-SOB) exerting highest sulfate-ion production were selected for formulating a probiotic consortium using mixture design matrix. The optimal composition was determined to be equal ratios of the three isolates that yielded 0.083 mM of sulfate from thiosulfate broth medium at room temperature in 7 days. This is a standalone report of sulfur-oxidizing probiotic consortium composed of the said bacteria. The consortium may be used as a strong tool for remediation of reduced sulfur in aquaculture and associated coastal environments.

In-depth performance study of an innovative decentralized multistage constructed wetland system treating real institutional wastewater.

The performance of an innovative decentralized multistage constructed wetland (DMCW) treating institutional wastewater is studied covering three seasons. The DMCW system with Canna lily efficiently removed organics contaminants like COD and BOD, and nutrients from the wastewater, showing its dependency on meteorological factors. Overall the performance is maximum in summer and least in monsoon, with a COD removal of 85.6% in summer followed by 82.5% in winter and 61.2% in monsoon. Removal of TSS (67.7-85.5%), PO43--P (52.1-64.4%), NH4+-N (56.6-71.6%), NO3--N (47.3-63.4%) and NO2--N (62-75.4%) were achieved along with heavy metals like Cr, Mn, Fe, Ni, Cu, Zn, As, Cd, Hg and Pb. Removal of pathogens like Vibrio is >98%, E. coli 95%, Pseudomonas 99%, and Aeromonas 63% was observed. Mass removal rate of COD was maximum in summer (97.3 g/m2/d) followed by winter (78.7 g/m2/d) and monsoon (43.5 g/m2/d). Majority of organics removal during the treatment was highlighted through Gas Chromatography-Mass Spectrometry (GCMS) and Fourier Transform Infrared Spectroscopy (FTIR) confirmed wastewater to be complex. The Canna lily accumulated various elements and oxides during the treatment with no stress on its health. The treated water quality is within the permissible limits and stands suitable for irrigational purposes. Better plant health and increased microbial diversity in the garden proves the suitability of treated water for irrigational activities. The results were validated using statistical tools like Mann-Whitney U test and principal component analysis.

Impact of 2nd wave of COVID-19-related lockdown on coastal water quality at Diu, western coast of India and role of total alkalinity on bacterial loads.

A detailed coastal water monitoring near Diu coast, western part of India was performed from October, 2020 to May, 2021 covering the 2nd lockdown time. Average monthly fluctuation from 7 different sampling stations of total 9 physico-chemical parameters such as pH, salinity, turbidity, nitrite (NO2), nitrate (NO3), ammonia (NH3), phosphate (PO4), total alkalinity and silicate were recorded. Initially, Mann-Kendall trend test for all the 9 parameters showed non-zero trend, which may be either linear or non-linear. During 2nd lockdown period, there was a fluctuation of value for parameters like pH, salinity, nitrate, nitrite and phosphate. Average total bacterial count and differential bacterial count also gradually decreased from March, 2021 sampling. Principal component analysis (PCA) plot covering all the physico-chemical parameters as well as the differential bacterial count showed a distinct cluster of all bacterial count with total alkalinity value. Subsequently, mathematical equation was formulated between total alkalinity value and all differential bacterial count. Upto our knowledge, this is the first report where mathematical equation was formulated to obtain value of different bacterial load based on the derived total alkalinity value of the coastal water samples near Diu, India.

Simple, rapid and on spot dye-based sensor for the detection of Vibrio load in shrimp culture farms.

For the detection of Vibrio bacteria, a kit involving two-step method was developed. In the in first step, a specific media was added in the water sample which selectively promote the growth of vibrios and inhibit the growth of other bacteria. The second step involved addition of dye-based sensor (already developed in our previous work) in the sample which detect the active Vibrio and changed the colour of the sample to red/pink. The vibrio detection kit was optimised on five different species of Vibrio (V. cholerae, V. parahaemolyticus, V. campbellii, V. harveyi and V. proteolyticus) and two negative control bacteria (Escherichia coli and Bacillus subtilis). The kit was further evaluated on aquaculture pond water and probiotics used in aquaculture farms. It successfully estimated Vibrio concentration of all the five strains in aquaculture ponds. The negative control bacteria and probiotics were not sensed by the kit. Hence, the kit developed here is perfect for the detection of Vibrio, especially in aquaculture farms.

New normal baseline data during nationwide lock down due to Covid 19 pandemic in the world's largest ship recycling yard at Alang, India.

A detailed study to generate the new normal baseline data has been carried out during nationwide lockdown (May 12 to May 16, 2020) covering sampling for ambient air, coastal water, coastal sediments, fish and bioaccumulation of heavy metals, in an around Alang, the world's biggest ship recycling yard. The lockdown data were compared with 2018 and 2019 observed data. PM10 values during lockdown were reduced by 3.75 to 4.5 times as compared with previous 2 years. Similarly, four-fold reduction of PM2.5 and SPM values was observed during lockdown. The gaseous pollutants like NO2 and O3 are within safe limit. Overall air quality index (AQI) improved significantly during lockdown. Similarly, there was drastic reduction in the majority of the nutrient parameters in the coastal water. Different heavy metal concentration in the coastal sediments samples also showed strong reduction during lockdown sampling in comparison with other two sampling. This proves that the coastal environment has its efficient self-cleaning potentials if there is considerable reduction in the anthropogenic as well as industrial activities. Diversity of phytoplankton and zooplankton also increased. The results were validated using statistical techniques like analysis of variance and least significance difference (LSD).

Characterization of VopJ by modelling, docking and molecular dynamics simulation with reference to its role in infection of enteropathogen Vibrio parahaemolyticus.

Vibrio parahaemolyticus is a renowned enteropathogen known for infecting humans. The infection usually involves various genes which help bacterium bypass the immune system of the host. Type III secretion system (T3SS) is an essential factor for the infection. The present study introduces the probable structure of VopJ, a T3SS effector of V. parahaemolyticus. The vopJ gene was amplified and sequenced from V. parahaemolyticus. The model generated through homology modelling showed a Z score of around 2.5, which fits quite near in the standard model available in the databases. The model has only a couple of outlier amino acids, which indicate a good fit of the model. Docking studies with small molecules like Acetyl-CoA, Inositol hexakisphosphate, GTP, and AMP have shown negative ΔG - 10.49, -52.80, -8.36 and -9.02, respectively, which indicates spontaneous binding. The molecular simulation studies have also supported the binding with a low RMSD value of less than 0.5 nm. The RMSF values obtained using the modelling were also quite low (>0.35 nm), which indicates the consistency achieved using the docking studies. These small molecules are very crucial in the MAPK pathways, which is essential for the immune response from the host cell. This effector can thus have an ability to highjack the immune system and help the bacterium in the potent infection. Up to our understanding, this is the first report which describes the in-silico model to understand the mode of infection of T3SS in enteropathogen V. parahaemolyticus. Communicated by Ramaswamy H. Sarma.

Characterization of phenazine-1-carboxylic acid by Klebsiella sp. NP-C49 from the coral environment in Gulf of Kutch, India.

Coral-associated microbes from Marine National Park (MNP), Gulf of Kutch (GoK), Gujarat, India, were screened for siderophore production. Maximum siderophore-producing isolate NP-C49 and its compound were identified and characterized. The isolate was identified as Klebsiella sp. through 16S rRNA genes sequencing (GenBank accession nos. KY412519 and MTCC 25160). Antibiotic susceptibility profile against 20 commercial antibiotics showed its more sensitivity compared to human pathogenic strain, i.e., Klebsiella pneumonia. The compound was identified as phenazine-1-carboxylic acid (PCA) using the multinuclear ID (1H and 13C) and 2D (1H-1H COSY and 1H-13C HETCOR) NMR along with high-resolution mass spectrometry. No significant difference in the bacterial growth in the presence of PCA, FeCl3 and Fe(OH)3 indicated involvement of factors other than PCA in bacterial growth. The study first reports the identification and characterization of PCA from Klebsiella sp. both from terrestrial and marine sources.

Estimation of bisulfate in edible plant foods, dog urine, and drugs: picomolar level detection and bio-imaging in living organisms.

In order to explore the properties of any species in solution, the actual, i.e. equilibrium concentration of the free species should be taken into account. Researchers have not paid attention to the deprotonation equilibrium between HSO4- and SO42- while probing bisulfate ion. In this study, we have addressed this concern and developed two zwitterions, CG (coumarin-integrated glycine) and CA (coumarin-integrated alanine), for the selective detection of HSO4- at a picomolar level (50 to 325 pM) with very high binding affinity (∼108 M-1) in pure water at physiological pH. The principle of HSO4- recognition was established via UV-vis and fluorescence techniques. DFT calculations suggested that the H-bonding interactions between the probes and HSO4- are the driving force for this unforeseen selectivity. The membrane penetration ability and nontoxicity of CG/CA enable them to function as staining agents in living brine shrimps and bacteria. The use of these probes for the estimation of HSO4- in various day-to-day edible foods and drugs along with urine samples is unprecedented. The significance and novelty of this study lies in the application and development of assays for estimating bisulfate in several real-world samples that are predominantly aqueous in nature, which are the first of their kind.