Coupling of phycoremediation and phytoremediation technologies to treat tannery effluents with rainwater dilutions.

From tannery effluent (TE) severely polluted with heavy metals (HMs viz., Cr, Cu, Cd, and Pb), hydrophytic phytoextraction remains a challenge as transplanted plants succumb to death on facing acclimatization shock. Current study was aimed at diluting TE with harvested rainwater (HR) for improving HM phytoextraction potential of Phragmites australis (a hydrophyte) assisted with phycoremediation of coupled algae (viz., Oedogonium sp. and Pithophora sp.). The TE:HR dilutions (TEDs) 0, 25, 50, 75, and 100% (v/v) included three sets: set-1 included algae only, set-2 included P. australis only and set-3 included P. australis coupled with combined algal inoculum. Results showed that P. australis assisted with HR dilution and combined algal inoculum showed significantly greater uptake of HMs from each of the TEDs than respective control treatments. Combined algal application in the TEDs proved phycoremediation assistants based on their bioaccumulation factor (BF). The dry biomass of P. australis in TEDs applied with phycoremediation assistants remained greater than uninoculated ones. Overall, HM translocation factor (TF) of P. australis for Cr, Cu, Cd, and Pb remained ≥ 1. The study concludes that HM phytoextraction is substantially increased when concentrated TE is diluted with HR and assisted with phycoremediation of HM tolerant algae.

Characterization of heavy metal-associated bacteria from petroleum-contaminated soil and their resistogram and antibiogram analysis.

The aim of the current study was to screen and identify heavy metal (chromium, cadmium, and lead) associated bacteria from petroleum-contaminated soil of district Muzaffarabad, Azad Jammu and Kashmir, Pakistan to develop ecofriendly technology for contaminated soil remediation. The petroleum-contaminated soil was collected from 99 different localities of district Muzaffarabad and the detection of heavy metals via an atomic absorption spectrometer. The isolation and identification of heavy metals-associated bacteria were done via traditional and molecular methods. Resistogram and antibiogram analysis were also performed using agar well diffusion and agar disc diffusion methods. The isolated bacteria were classified into species, i.e., B. paramycoides, B. albus, B. thuringiensis, B. velezensis, B. anthracis, B. pacificus Burkholderia arboris, Burkholderia reimsis, Burkholderia aenigmatica, and Streptococcus agalactiae. All heavy metals-associated bacteria showed resistance against both high and low concentrations of chromium while sensitive towards high and low concentrations of lead in the range of 3.0 ± 0.0 mm to 13.0 ± 0.0 mm and maximum inhibition was recorded when cadmium was used. Results revealed that some bacteria showed sensitivity towards Sulphonamides, Norfloxacin, Erythromycin, and Tobramycin. It was concluded that chromium-resistant bacteria could be used as a favorable source for chromium remediation from contaminated areas and could be used as a potential microbial filter.

Integrative Evaluation of the Ecological Hazards by Microplastics and Heavy Metals in Wetland Ecosystem.

This study was performed to evaluate the impact of microplastics and heavy metals (Pb, Cd, Cr, Cu, Zn, Ni) on sediments, water, aquatic plants (Pistia stratiotes, Alternanthera philoxeroides, and Ipomoea carnea), and fish (Labeo rohita) samples collected from five different sites in the Bajwat wetlands in Sialkot, Pakistan. The concentrations of Pb, Cd, and Cr were above the permissible limits devised by WHO in all the ecosystem components (i.e. sediments, water, plants, and fish) at all sites. The maximum amount of microplastic particles (2317 microplastic particles per kg of sediments) was recorded at Site 1. The filaments were the most commonly found type of microplastics. Plants and fish samples also showed considerable concentration of metals. The multivariate statistical analysis revealed anthropogenic sources of elevated concentrations of metal elements which could cause adverse biological effects in the ecosystem.

Burkholderia cepacia CS8 improves phytoremediation potential of Calendula officinalis for tannery solid waste polluted soil.

Microbes have shown potential for the bioremediation of tannery waste polluted soil. During our previous study, it was observed that heavy metal resistant Burkholderia cepacia CS8 augmented growth and phytoremediation capability of an ornamental plant. Objective of the present research work was to evaluate the capability of B. cepacia CS8 assisted Calendula officinalis plants for the phytoremediation of tannery solid waste (TSW) polluted soil. The TSW treatment significantly reduced growth attributes and photosynthetic pigments in C. officinalis. However, supplementation of B. cepacia CS8 which exhibited substantial tolerance to the TSW amended soil, augmented growth traits, carotenoid, proline, and antioxidant enzymes level in C. officinalis under toxic and nontoxic regimes. Inoculation of B. cepacia CS8 augmented plant growth (shoot length 13%, root length 11%), physiological attributes (chlorophyll a 14%, chlorophyll b 17%), antioxidant enzyme activities (peroxidase 24%, superoxide dismutase 31% and catalase 19%), improved proline 36%, phenol 32%, flavonoids 14% and declined malondialdehyde (MDA) content 15% and hydrogen peroxide (H2O2) level 12% in C. officinalis at TSW10 stress compared with relevant un-inoculated plants of TSW10 treatment. Moreover, B. cepacia CS8 application enhanced labile metals in soil and subsequent metal uptake, such as Cr 19%, Cd 22%, Ni 35%, Fe 18%, Cu 21%, Pb 34%, and Zn 30%, respectively in C. officinalis plants subjected to TSW10 stress than that of analogous un-inoculated treatment. Higher plant stress tolerance and improved phytoremediation potential through microbial inoculation will assist in the retrieval of agricultural land in addition to the renewal of native vegetation.

During the current study, it was observed that combination of Calendula officinalis and metal tolerant Burkholderia cepacia CS8 not only improved plant growth but also helped phyto-extraction of pollutants present in the tannery solid waste polluted soil. According to our information, research work describing the phytoremediation potential of native metal tolerant microbes and ornamental plants has not been reported in Pakistan.

Metabolic profile and molecular characterization of endophytic bacteria isolated from Pinus sylvestris L. with growth-promoting effect on sunflower.

Endophytic bacteria inhabit plant tissues such as roots, stems, leaves, fruits, and seeds and can multiply inside plant tissue without damaging them. This study involves the isolation, characterization, metabolic profiling, and effect of endophytic bacteria isolated from the roots of Scots pine (Pinus sylvestris), on the growth of sunflower. In the current study, fifteen isolates of endophytic bacteria were obtained from the roots of Scots pine, and their molecular characterization was performed using 16 s rRNA ribotyping. The molecular characterization revealed that the strains belonged to Bacillus spp., Pseudomonas spp., Micrococcus sp., Serratia sp., Enterobacter sp., Pantoea sp., Staphylococcus sp., and Microbacterium sp. Among the isolated strains, 9 strains showed positive results for ammonium production, 12 strains for calcium solubilization, 11 strains for magnesium solubilization, 5 strains for zinc solubilization, 12 strains for phosphate solubilization, 8 strains for potassium solubilization, 10 strains for indole acetic acid (IAA) production, 9 strains for siderophore, and 6 strains for hydrogen cyanide (HCN) production. The greenhouse experiment results demonstrated that all isolated endophytic bacteria improved the shoot length, dry weight, and chlorophyll content of sunflower, whereas a significant increase was observed by PS-3 (Bacillus cereus), PS-6 (Serratia marcescens), and PS-8 (Pseudomonas putida). Besides, the concentration of nitrogen, phosphorus, and potassium were also measured in sunflower shoots, and results asserted that bacterial inoculation increased the bioavailability of these essential nutrients to plants compared to uninoculated control. Thus, these endophytic bacteria could be used as an encouraging option to improve plant growth and performance.

Synergistic impact of two autochthonous saprobic fungi (A. niger and T. pseudokoningii) on the growth, ionic contents, and metals uptake in Brassica juncea L. and Vigna radiata L. under tannery solid waste contaminated soil.

Unrestricted disposal of tannery solid waste (TSW) into agricultural soils has resulted in the contamination of heavy metals (HMs) such as chromium (Cr) cadmium (Cd), Copper (Cu), and Zinc (Zn) along with the severe potential to degrade the environmental quality around the world. In the present study, a combined phyto- and myco-remediation strategy was evaluated to enhance the growth, ionic contents, and phytoextraction potential of Brassica juncea and Vigna radiata for HMs from TSW-contaminated soil. A pot experiment was conducted in the greenhouse using single or combined inoculation of Trichoderma pseudokoningii (Tp) and Aspergillus niger (An) in B. juncea and V. radiata under TSW-contaminated soil at different doses (0, 50, and 100%). The results showed that the growth parameters of both B. juncea and V. radiata were severely affected under 50 and 100% TSW treatment. The combined inoculation of both the fungal species ameliorated the positive impacts of 50 and 100% TSW application on growth and ionic contents accumulation in B. juncea and V. radiata. The combined application of An + Tp at 100% TSW enhanced the shoot length (87.8, 157.2%), root length (123.9, 120.6%), number of leaves (184.2, 175.0%), number of roots (104.7, 438.9%), and dry weight (179.4, 144.8%) of B. juncea and V. radiata, respectively as compared to control with any fungal treatment at 100% TSW. A single application of An at different doses of TSW enhanced the metal concentration in B. juncea, whereas Tp increased the concentration of the metals in V. radiata. The concentration of Cr in roots (196.2, 263.8%), shoots (342.4, 182.2%), Cu in roots (187.6, 137.0%), shoots (26.6, 76.0%), Cd in roots (245.2, 184.6%), shoots (142.1, 73.4%), Zn in roots (73.4, 57.5%), shoots (62.9, 57.6%), in B. juncea were increased by the application of An at 50 and 100% treatment levels of TSW, respectively compared to control (C). Moreover, the HMs (Cr, Cu, Cd, and Zn) uptake was also improved under 50 and 100% TSW with the combined inoculation of Tp + An in both B. juncea and V. radiata. In conclusion, the combined inoculation of Tp + An was more effective in metal removal from TSW-treated soil.NOVELTY STATEMENTLimited studies have been conducted on filamentous fungi systematically under metal-contaminated sites for their diversity, metal tolerance, and their potential in enhancing the phytoremediation potential of different crop plants.In the present study, single and/or combined inoculation of fungal strains was found effective in alleviating different metals stress in tannery solid waste contaminated soil by improving defense mechanisms and plant growth due to the association between fungal strains and plants.The combined application of both fungal strains had an additive effect in enhancing the bioaccumulation capacity of B. juncea and V. radiata compared to their single inoculation.

Autochthonous strains of Trichoderma isolated from tannery solid waste improve phytoextraction potential of heavy metals by sunflower.

This research work was aimed at isolating and demonstrating the significant potential of autochthonous fungi for phytoextraction of hazardous metals in metal polluted soil using Helianthus annuus. Four multi-metal resistant strains of Trichoderma were selected from a total of 21 strains isolated from tannery polluted soil and tannery solid waste. Autochthonous Trichoderma strains were used singly and in the form of consortium (TC). Sunflower was grown in pots for 90 days having eight different amendments of tannery polluted soil with and without Trichoderma inoculation. Growth and biochemical attributes of the plants were observed along with metal content extract by different plant parts. The results revealed that TC enhanced shoot length, shoot dry weight, and metal uptake as compared to single specie inoculation. Similarly, BCF (72.8-118.23%) and TF were significantly pronounced in shoots of H. annuus grown with TC at 40% amended soil. The biochemical analysis of the plants showed that Trichoderma strains boosted the enzymatic (catalase, peroxidase, and superoxide dismutase) antioxidants in the plants. The use of indigenous fungi with metal accumulating plants like sunflower can help to alleviate metal contamination from industrial sites and can make the soil cultivable for energy crops.

The genus Trichoderma is among the most common cosmopolitan soil fungi that enhance phytoextraction capability of plants. Hence, the isolation and identification of diversified and potent Trichoderma strains from contaminated environments is the need of the hour for broad spectrum applications in bioremediation. In the present study, contaminated soil mycoflora was explored and multi-metal resistant strains of Trichoderma were isolated. Their application in myco-assisted phytoextraction with Helianthus annuus was assessed to analyze their impact on the metal removal efficacy and enhancing growth in highly contaminated soil.

Efficacy of cow and buffalo dung on vermiremediation and phytoremediation of heavy metals via Fourier-transform infrared spectroscopy and comet assay.

Heavy metal contamination raised significant concerns throughout the world. The current research aimed to evaluate the impact of organic manure (cow dung and buffalo dung) on vermiremediation and phytoremediation and to remediate heavy metals, i.e., cadmium, lead, and chromium, from artificial contaminated soil via both remediation techniques. The impact of livestock manure was evaluated for the first time which could be effective in in situ as well as ex situ studies. Eisenia fetida, Pheretima lignicola, and Spinacia oleracea were used for the remediation process. Results revealed that E. fetida tolerated lead at 280 mg, cadmium at 150 mg, and chromium at 860 mg compared to P. lignicola. The growth and reproduction of E. fetida were efficient in the cow dung manure compared to buffalo dung. Similarly, seed germination and growth of Spinacia oleracea were better in cow dung media compared to buffalo dung. Bioaccumulation factor showed that E. fetida showed a higher accumulation of heavy metals in their tissues when vermi + phytoremediation was jointly applied (9.50 mg/l of Pb, 24.166 of Cd, and 6.695 of Cr). Fourier-transform infrared spectroscopy indicated that heavy metals had no drastic effects on E. fetida and S. oleracea. Similarly, comet assay revealed that heavy metals had no genotoxic effect on the E. fetida and S. oleracea. It was concluded that both E. fetida and S. oleracea are appropriate for heavy metals remediation in cow dung manure.

Role of composted tannery solid waste and its autochthonous microbes in enhancing phytoextraction of toxic metals and stress abatement in sunflower.

The excessive concentration of multiple heavy metals in the tannery solid waste (TSW) needs integrated process solutions for its decontamination. This study is aimed at deriving TSW compost and autochthonous microbe synergies for improving phytoextraction potential of sunflower. In-vessel composting of TSW was carried out by using fruit waste as an inoculum to achieve the optimized conditions. Autochthonous strains of Trichoderma viride and Bacilllus sp. isolated from TSW were utilized individually as well as in combination with TSWC amendments of 2.5, 5 and 10% (w/w) prepared in our pilot scale experiment. Analyses of TSW compost based on FTIR and SEM illustrated the wide range of functionality and porosity along the mesh of fungal hyphae and inorganic moieties present on the compost surface. Plant biomass and TMs uptake (Cr 540 mg kg-1 > Cd 330 mg kg-1 > Pb 285 mg kg-1) were significantly pronounced in shoots of sunflower under combined treatments at 10% TSWC amended soils. However, in seeds, TMs were found below detection limit (BDL) through atomic absorption spectrophotometry. Biochemical assays of sunflower including total chlorophyll content (18%), total soluble protein (45%), superoxide dismutase (80%) and catalase (75%) activities were also increased significantly at higher level of amendment in combination with microbes than in the control. Despite being high in TMs, high biomass in sunflower and associated elevation in biochemical products demonstrate the potential of TSW for valorization.Novelty statement: This study identifies the cost-effective management of multi metal contaminated tannery solid waste through deriving its compost along with autochthonous microbes as phytoextraction assistants by yielding higher plant biomass. This study suggests the use of composted TSW inoculated with selected autochthonous fungi and bacteria for enhancing sunflower's biomass and enhancing the bioavailable fractions of toxic metals for phytoextraction.

Ameliorative role of foliar Zn-lysine application on wheat (Triticum aestivum L.) stressed by Tannery Wastewater.

Tannery industries discharge a high concentration of chromium (Cr) along with other heavy metals, which are hazardous for all life forms. With increasing shortage of freshwater, tannery effluent is frequently used for crop an irrigation, causing damage to plants' health. In order to address this challenge, amino acid chelate fertilizer was used to investigate the impact on wheat crops against tannery waste water. Tannery wastewater (TW) was used at different levels such as 0%, 25%, 50%, and 100% with an amendment of foliar Zn-lysine (Zn-lys) at30 mg/L. This research highlighted the positive correlation of Zn-lysine on the morpho-physiological, biochemical, and gas exchange traits under different levels of tannery wastewater. The findings of this study showed that the application of Cr-rich tannery wastewater at different treatment levels resulted in a significant reduction in plant height (23%, 31%, and 36%), the number of tillers (21%, 30%, and 43%), spike (19%, 36%, and 55%) and dry weight (DW) of grains (10%, 25%, and 49%) roots DW (17%, 41%, 56%), and shoots DW (22%, 32%, and 47%) as compared to control. Foliar-applied Zn-lys positively enhanced photosynthetic attributes, antioxidant enzymes activities and gas exchange traits by reducing the oxidative stress alone and under Cr stress. The concentration of Cr in roots (21%, 37%, 38%) and shoots (11%, 36%, 37%) was reduced by the foliar application of Zn-lys at different treatment levels. These findings conclude that Zn-lys served as a protector for the growth and development of wheat and has an incredible potential to inhibit the phytotoxicity induced by excess Cr. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-022-01265-6.

Biosorption potential and molecular characterization of metal-resistant autochthonous microbes from tannery solid waste.

This study encompasses isolation and screening of heavy metal-resistant fungal and bacterial strains from tannery solid waste (TSW). Twelve fungal strains and 25 bacterial strains were isolated from TSW. The growth of fungal strains was observed against different heavy metals ranging from 10 to 1050 mg L-1 and the growth of bacteria was observed in metal concentrations ranging from 10 to 1200 mg L-1. Five multi-metal-resistant fungal isolates belonging to the genus Trichoderma and ten bacterial isolates belonging to the genus Bacillus showed good metal resistance and biosorption potential. They were identified through molecular techniques, fungi based on ITS region ribotyping, and bacteria based on 16S rRNA ribotyping. The fungal strains were characterized as T. hamatum (TSWF-06), T. harzianum (TSWF-11), T. lixii (TSWF-02), and T. pseudokoningii (TSWF-03, TSWF-10). The bacterial strains were characterized as Bacillus xiamenensis (TSW-02), B. velezensis (TSW-05), B. piscis (TSW-06), B. safensis (TSW-10), B. subtilis (TSW-14, TSW-15, TSW-17) B. licheniformis (TSW-19), B. cereus (TSW-20), and B. thuringiensis (TSW-22). The fungal strains, namely, T. pseudokoningii (TSWF-03) and T. harzianum, proved to be two multi-metal-resistant strains with good biosorption efficiency. Unlike fungi, bacterial strains showed metal-specific resistance. The strains Bacillus xiamenensis, B. subtilis (TSW-14), and B. subtilis (TSW-15) showed good biosorption efficiency against Cr, B. safensis against Cu, B. piscis, and B. subtilis (TSW-17) against Pb and B. licheniformis and B. thuringiensis against Zn. The autochthonous fungal and bacterial strains can therefore be employed to clean metal-contaminated environments.

Autochthonous Arthrospira platensis Gomont Driven Nickel (Ni) Phycoremediation from Cooking Oil Industrial Effluent.

Nickel (Ni) leftovers arise from both catalyst application interventions and Ni alloy piping of the cooking oil industry (COI) being wasted as pollutants of freshwater bodies via discharged effluent. The current study assessed one of the indigenously feasible Ni removal systems comprising autochthonous Arthrospira platensis Gomont (AP)-driven Ni phycoremediation cells (NPCs). After screening AP for hyperaccumulation in the Ni spiked solution, AP was transferred to the NPCs. Propagation of the AP inoculum was proportionate to the pollution load drop of COI with 22.97 and 55.07% drops in the biochemical (BOD) and chemical oxygen demand (COD), respectively. With the 0.11 bioconcentration factor, there was an uptake of 14.24 g mineral with 16.22% Ni removal and a 36.35 desorption ratio. The experimental data closely fitted with the Langmuir and Freundlich isotherms, respectively. The study concluded that A. platensis could be taken for treatment of Ni-loaded industrial effluents at the microcosmic level.

Application of microbe-impregnated tannery solid waste biochar in soil enhances growth performance of sunflower.

Synergistic effect of biochar and microbes in soil enhances performance of plants. Hazardous tannery solid waste can be reduced by one-third in volume by conversion to biochar. A greenhouse trial was set up with soil having different doses of metal resistant microbe-impregnated biochar (MIBC) prepared from tannery solid waste. Consortia of autochthonous strains of Trichoderma and Bacillus were inoculated on BC and the behavior and fate of metals were evaluated for their bioavailability to sunflower. Sunflower was grown in pots for 80 days having six different amendments of tannery solid waste biochar (0-10% w/w) with and without Trichoderma and Bacillus consortia and its morphological and biochemical attributes as well as metal uptake were observed. The results illustrated that application of BC at 2% rate without inoculation increased the shoot length and dry biomass by 19.8% and 77.4%, respectively, while plant growth and performance were reduced at higher amendments of BC. However, application of MIBC with Trichoderma or/and Bacillus consortium significantly improved the plant attributes at all levels of amendment. The results indicated that MIBC having Trichoderma and Bacillus consortia at 10% rate increased shoot length and dry biomass by 65.3% and 516% compared to control without BC. Application of BC without inoculation reduced the uptake of Cu, Fe, and Ni and increased the mobilization of all other metals for uptake in sunflower. Mobilization and uptake of Cd, Cr, Cu, Ni, Pb, and Zn decreased with MIBC having Trichoderma and Bacillus consortia whereas that of Fe and Mg were noted. A considerable decrease in proline and total phenolic content was demonstrated by MIBC-grown sunflower. The data of metal fractionation in BC also supported the above findings. Therefore, MIBC can be used as a promising option for enhancing growth performance and ensuring the physiological safety of sunflower as an energy crop.

Fungal biostimulant-driven phytoextraction of heavy metals from tannery solid waste contaminated soils.

Two of the multiple limitations of phytoextraction efficiency (PE %) of TSW polluted soils are: (i) low growth of plant performance, (ii) poor bioavailability of excessive essential and heavy metals (ascribed as Category-I and II metals respectively) The current study reports biostimulant role of allochthonous Trichoderma harzianum (F1) and autochthonous Trichoderma pseudokoningii (F2) in growth of Tagetes patula L. and uptake of Category-I & II metals from TSW-soil (0, 5 & 10%). Significantly higher growth (27.5-47.8% dry wt. than Control) and highly significantly higher uptake of Category-I & II metals (72-80% Ca, 32-69% K, 72-76% Na & 73-86% Cd, 63-100% Cr, 72-77% Cu, 73-78% Fe, 43-77% Mg, 22-33% Ni, 70-73% Zn) was observed in T. patula applied with F1 + F2 treatment. The PE (%) parameters viz.specific extraction yield, tolerance and translocation index of Category-I & II metals were higher in plants cultivated on fungal inoculated TSW:soil. The Trichoderma spp. acted as strong biostimulants for enhancing plant growth and conc. of catalase (CAT, 44-52% than control), superoxide dismutase (SOD, 37-43%), soluble proteins (37-68%) and total chlorophyll (10-26%) in T. patula during metal phytoextraction of TSW:soil. Novelty statement Due to multiple socio-economic constraints for effective management of tannery solid waste (TSW), the heavy metal phytoextraction seems to be one of the promising approaches. However, due to complex composition of TSW, that is, with more than 37 components, high pH, multiple types and high conc. of metals; there lies huge challenge of enhancing phytoextraction efficiency (PE %). This can be done by enhancing growth of hyperaccumulator plants and increasing bioavailable fraction of metals. The current study suggests application of selected fungal biostimulants for increasing growth of T. patula while improving bioavailable fraction of the total metal contents of the TSW: soil.

Modeling of Cr contamination in the agricultural lands of three villages near the leather industry in Kasur, Pakistan, using statistical and GIS techniques.

Kasur is one of the hubs of leather industry in the Punjab, Pakistan, where chrome tanning method of leather processing is extensively being used. Chromium (Cr) accumulation levels in the irrigation water, soil, and seasonal vegetables were studied in three villages located in the vicinity of wastewater treatment plant and solid waste dumping site operated by the Kasur Tanneries Waste Management Agency (KTWMA). The data was interpreted using analysis of variance (ANOVA), clustering analysis (CA), and principal component analysis (PCA). Interpolated surface maps for Cr were generated using the actual data obtained for the 30 sampling sites in each of the three villages for irrigation water, soil, and seasonal vegetables. The level of contamination in the three villages was directly proportional to their distance from KTWMA wastewater treatment plant and the direction of water runoff. The highest level of Cr contamination in soil (mg kg-1) was observed at Faqeeria Wala (37.67), intermediate at Dollay Wala (30.33), and the least in Maan (25.16). A gradational variation in Cr accumulation was observed in the three villages from contaminated wastewater having the least contamination level (2.02-4.40 mg L-1), to soil (25.16-37.67 mg kg-1), and ultimately in the seasonal vegetable crops (156.67-248.33 mg kg-1) cultivated in the region, having the highest level of Cr contamination above the permissible limit. The model used not only predicted the current situation of Cr contamination in the three villages but also indicated the trend of magnification of Cr contamination from irrigation water to soil and to the base of the food chain. Among the multiple causes of Cr contamination of vegetables, soil irrigation with contaminated groundwater was observed to be the dominant one.

The Pakistan Risk of Myocardial Infarction Study: a resource for the study of genetic, lifestyle and other determinants of myocardial infarction in South Asia.

The burden of coronary heart disease (CHD) is increasing at a greater rate in South Asia than in any other region globally, but there is little direct evidence about its determinants. The Pakistan Risk of Myocardial Infarction Study (PROMIS) is an epidemiological resource to enable reliable study of genetic, lifestyle and other determinants of CHD in South Asia. By March 2009, PROMIS had recruited over 5,000 cases of first-ever confirmed acute myocardial infarction (MI) and over 5,000 matched controls aged 30-80 years. For each participant, information has been recorded on demographic factors, lifestyle, medical and family history, anthropometry, and a 12-lead electrocardiogram. A range of biological samples has been collected and stored, including DNA, plasma, serum and whole blood. During its next stage, the study aims to expand recruitment to achieve a total of about 20,000 cases and about 20,000 controls, and, in subsets of participants, to enrich the resource by collection of monocytes, establishment of lymphoblastoid cell lines, and by resurveying participants. Measurements in progress include profiling of candidate biochemical factors, assay of 45,000 variants in 2,100 candidate genes, and a genomewide association scan of over 650,000 genetic markers. We have established a large epidemiological resource for CHD in South Asia. In parallel with its further expansion and enrichment, the PROMIS resource will be systematically harvested to help identify and evaluate genetic and other determinants of MI in South Asia. Findings from this study should advance scientific understanding and inform regionally appropriate disease prevention and control strategies.

A novel haplotype in ABCA1 gene effects plasma HDL-C concentration.

BACKGROUND AND OBJECTIVES: ATP-binding cassette transporter 1 (ABCA1) is a trans-membrane protein responsible for the efflux of cholesterol and phospholipids across the cell membrane, an essential step in the reverse cholesterol transport system. This study investigates the effect of five non-synonymous SNPs of ABCA1 gene on plasma HDL-C levels in Pakistani individuals free of ischemic heart disease and stroke. METHODS: Five non-synonymous SNPs were selected after sequencing ABCA1 gene in patients of Hypoalphalipoproteinemia. The presence of these SNPs was then checked in 200 individuals by using PCR-RFLP. Plasma glucose and lipid fractions were measured in fasting state. Ethical approval was obtained from the Ethical Review Committee, Aga Khan University and informed consent was obtained from all subjects. RESULTS: LL genotype of V825L polymorphism was associated with decreased levels of HDL-C [-0.17 (-0.32 to -0.19); P=0.02] and P774 allele showed a significant increase in HDL-C levels as compared to T774 allele [-0.15 (-0.18 to -0.02); P=0.01]. R219K, A399V and V771M polymorphisms did not show any association with levels of HDL-C, LDL-C, cholesterol and triglycerides. Haplotype analysis between R219K and V825L polymorphisms showed a unique interaction between R219 allele and L825 allele. The RL haplotype was found to be associated with decreased levels of HDL-C [-0.12 (-0.22 to -0.03); P=0.001]. CONCLUSIONS: ABCA1 polymorphisms are associated with varying levels of HDL-C in Pakistani individuals. These results warrant further investigations as ABCA1 polymorphisms may have a major role in the high incidence of cardiovascular disorders in South Asians.

Association of phosphodiesterase 4D gene with ischemic stroke in a Pakistani population.

BACKGROUND AND OBJECTIVES: Identification of STRK1 locus by the deCODE group followed by the discovery of phosphodiesterase 4D (PDE4D) gene in strong association with ischemic stroke patients has provided useful insights toward understanding the genetic etiology of the disease. In this study, we aimed at investigating the association between 3 polymorphisms of the PDE4D gene and ischemic stroke in the Pakistani population. METHODS: Three polymorphisms in PDE4D gene were analyzed in 200 patients of ischemic stroke and 250 controls of Pakistani origin using polymerase chain reaction-restriction fragment length polymorphism method. Data were coded and entered in SPSS Windows (version 12.0). Odds ratios and 95% CIs were calculated using multivariate logistic regression analysis. RESULTS: Marker SNP83(rs966221) was found significantly associated with ischemic stroke on univariate and multivariate analysis (P<0.005; odds ratio, 1.64 [1.13 to 2.40]). Haplotype analysis for markers in linkage disequilibrium failed to show any association with the disease. CONCLUSIONS: The association of PDE4D variation with ischemic stroke extends to the Pakistani population and supports a role for phosphodiesterases in stroke pathogenesis.