Search results for: non-toxique assays

Authors: Suruchi Shukla, Shantanu Prakash, Amita Jain

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Introduction: Arboviral diseases are one of the most common causes of viral hemorrhagic fever (VHF). Of which, Dengue and Chikungunya pose a significant health problem in India. Arbovirus has a tendency to cross the territories and emerge in the new region. Considering the above issues, in the current study active surveillance was conducted among viral hemorrhagic fever (VHF) cases reported from Uttar Pradesh (UP), India. We studied the arboviral etiology of VHF; mainly Dengue, Chikungunya, and ZIKA. Methods: Clinical samples of 465 suspected VHF cases referred to tertiary care referral center of UP, India were enrolled in the study during a period from 15th May 2016 to 9th March 2018. Serum specimens were collected and analyzed for the presence of Dengue, Chikungunya, and ZIKA either by serology and/or by molecular assays. Results: Of all tested, 165 (35.4%) cases were positive for either Dengue or Chikungunya. Dengue (21.2%) was found to be the most prevalent, followed by Chikungunya, (6.6%). None of the cases tested positive for ZIKA virus. Serum samples of 35 (7.5%) cases were positive for both Dengue and Chikungunya. DEN-2 serotype was the most predominant serotype. Phylogenetic and sequence analysis of DEN-2 strains showed 100% clustering with the Cosmopolitan genotype strain. Bleeding from several sites, jaundice, abdominal pain, arthralgia, haemoconcentration, and thrombocytopenia were significantly higher in dengue hemorrhagic cases. However, the rash was significantly more common in Chikungunya patients. Most of the Dengue and Chikungunya positive cases (Age group 6-40 years) were seen in post monsoon season (September to November). Conclusion: Only one-third of total VHF cases are positive for either Dengue/Chikungunya or both. This necessitates the screening of other etiologies capable of causing hemorrhagic manifestations.

Keywords: viral hemorrhagic fever, dengue, chikungunya, zika, India

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Authors: Saeid Jafari, Khursheed Ahmad Sheikh, Randy W. Worobo, Kitipong Assatarakul

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In this study, the extraction of cocoa shell powder (CSP) was optimized, and the optimized extracts were spray-dried for encapsulation purposes. Temperature (45-65 ◦C), extraction time (30–60 min), and ethanol concentration (60–100%) were the extraction parameters. The response surface methodology analysis revealed that the model was significant (p ≤ 0.05) in interactions between all variables (total phenolic compound, total flavonoid content, and antioxidant activity as measured by 2,2-Diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP assays), with a lack of fit test for the model being insignificant (p > 0.05). Temperature (55 ◦C), time (45 min), and ethanol concentration (60%) were found to be the optimal extraction conditions. For spray-drying encapsulation, some quality metrics (e.g., water solubility, water activity) were insignificant (p > 0.05). The microcapsules were found to be spherical in shape using a scanning electron microscope. Thermogravimetric and differential thermogravimetric measurements of the microcapsules revealed nearly identical results. The gum arabic + maltodextrin microcapsule (GMM) showed potential antibacterial (zone of inhibition: 11.50 mm; lower minimum inhibitory concentration: 1.50 mg/mL) and antioxidant (DPPH: 1063 mM trolox/100g dry wt.) activities (p ≤ 0.05). In conclusion, the microcapsules in this study, particularly GMM, are promising antioxidant and antibacterial agents to be fortified as functional food ingredients for the production of nutraceutical foods with health-promoting properties.

Keywords: functional foods, coco shell powder, antioxidant activity, encapsulation, extraction

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Authors: Mohammad K. Parvez, Ahmed H. Arbab, Mohammed S. Al-Dosari, Adnan J. Al-Rehaily

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We investigated ex vivo and in vivo antioxidative and hepatoprotective effect of Aerva javanica. Total ethanol extract of A. javanica aerial parts was prepared, and tested on DCFH-toxicated HepG2 cell in CCl4-injured Wistar rats. MTT-assay was used to determine cell viability, and serum biochemical markers of liver injury as well as histopathology were performed. In vitro DPPH and β-carotene free-radical scavenging assay and phytochemical screening of the extract was done. Furthermore, A. javanica total extract was standardized and validated by HPTLC method. While DCFH-injured cells were recovered to about 56.7% by 100 microg/ml of the extract, a 200 microg/ml dose resulted in hepatocytes recovery by about 90.2%. Oral administration of the extract (100 and 200 mg/kg.bw/day) significantly normalized the serum SGOT, SGPT, GGT, ALP, bilirubin, cholesterol, HDL, LDL, VLDL, TG and MDA levels, including tissue NP-SH and TP in CCl4-injured rats. In addition, the histopathology of dissected liver also revealed that A. javanica cured the tissue lesion compared to reference drug, Silymarin. In vitro assays revealed strong free-radical scavenging ability of the extract and presence of alkaloids, flavonoids, tannins, sterols and saponins where Rutin, a well-known antioxidant flavonoid was identified. Our finding therefore, suggests the therapeutic potential of A. javanica in various liver diseases. However, isolation of the active principles, their mechanism of action and other therapeutic contribution remain to be addressed.

Keywords: Aerva javanica, antioxidant, hepatoprotection, rutin

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Authors: Famuwagun A. A., Abiona O. O., Gbadamosi S.O., Adeboye O. A., Adebooye O. C.

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The need to provide more information on the perceived bioactive status of the peel of plantain fruit informed the design of this research. Matured Plantain fruits were harvested, and fruits were allowed to ripen spontaneously. Samples of plantain fruit were taken every fortnight, and the peels were removed. The peels were dried using two different drying techniques (Oven drying and sun drying) and milled into powdery forms. Other samples were picked and processed in a similar manner on the first, third, seventh and tenth day until the peels of the fruits were fully ripped, resulting in eight different samples. The anti-oxidative properties of the samples using different assays (DPPH, FRAP, MCA, HRSA, SRSA, ABTS, ORAC), inhibitory activities against enzymes related to diabetes (alpha-amylase and glucosidase) and inhibition against angiotensin-converting enzymes (ACE) were evaluated. The result showed that peels of plantain fruits on the 7th day of ripening and sundried exhibited greater inhibitions against free radicals, which enhanced its antioxidant activities, resulting in greater inhibitions against alpha-amylase and alpha-glucosidase enzymes. Also, oven oven-dried sample of the peel of plantain fruit on the 7th day of ripening had greater phenolic contents than the other samples, which also resulted in higher inhibition against angiotensin converting enzymes when compared with other samples. The results showed that even though the unripe peel of plantain fruit is assumed to contain excellent bioactive activities, consumption of the peel should be allowed to ripen for seven days after maturity and harvesting so as to derive maximum benefit from the peel.

Keywords: functional ingredient, diabetics, hypertension, functional foods

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Authors: Chengcao Sun, Cuili Yang, Shujun Li, Ruilin Xue, Yongyong Xi, Liang Wang, Dejia Li

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Backgrounds: Inflammation is widely distributed in patients with Duchenne muscular dystrophy (DMD), and ultimately leads to progressive deterioration of muscle function with the co-effects of chronic muscle damage, oxidative stress, and reduced oxidative capacity. NF-E2-related factor 2 (Nrf2) plays a critical role in defending against inflammation in different tissues via activation of phase II enzymes, heme oxygenase-1 (HO-1). However, whether Nrf2/HO-1 pathway can attenuate muscle inflammation on DMD remains unknown. The purpose of this study was to determine the anti-inflammatory effects of Sulforaphane (SFN) on DMD. Methods: 4-week-old male mdx mice were treated with SFN by gavage (2 mg/kg body weight per day) for 4 weeks. Gastrocnemius, tibial anterior and triceps brachii muscles were collected for related analysis. Immune cell infiltration in skeletal muscles was analyzed by H&E staining and immuno-histochemistry. Moreover, the expressions of inflammatory cytokines,pro-inflammatory cytokines and Nrf2/HO-1 pathway were detected by western blot, qRT-PCR, immunohistochemistry and immunofluorescence assays. Results: Our results demonstrated that SFN treatment increased the expression of muscle phase II enzymes HO-1 in Nrf2 dependent manner. Inflammation in mdx skeletal muscles was reduced by SFN treatment as indicated by decreased immune cell infiltration and lower expressions of the inflammatory cytokines CD45, pro-inflammatory cytokines tumour necrosis factor-α and interleukin-6 in the skeletal muscles of mdx mice. Conclusions: Collectively, these results show that SFN can ameliorate muscle inflammation in mdx mice by Nrf2/HO-1 pathway, which indicates Nrf2/HO-1 pathway may represent a new therapeutic target for DMD.

Keywords: sulforaphane, Nrf2, HO-1, inflammation

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Authors: Leila Najafian

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Sarcoplasmic protein hydrolysates (SPHs) and myofibrillar protein hydrolysates (MPHs) from patin (Pangasius sutchi) were produced using two types of proteases: Papain and Alcalase. 1,1-diphenyl-2-picrylhydrazyl (DPPH), 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) diammonium salt (ABTS) radical scavenging activities and metal chelating activity assays for antioxidant activities were carried out on the SPHs and MPHs. The hydrolysates were isolated and purified by ultrafiltration, gel filtration and reverse phase high-performance liquid chromatography (RP-HPLC) and liquid chromatography with tandem mass spectrometry detection (LC-MS/MS) was used in identifying peptide sequences. The results showed that when the DH of MPHs increased, the protein solubility increased, while the highest amount of the protein solubility of SPHs was after 60 min incubation. The effect of DH on antioxidant activities of SPHs and MPHs was investigated. Among the hydrolysates, papain-MPH and Alcalase-SPH, which had the highest antioxidant activities, were purified. The potent fractions obtained from RP-HPLC of sarcoplasmic (SI 3 fraction) and myofibrillar (MI 4 fraction) hydrolysates showed the highest DPPH radical scavenging activity. The FVNQPYLLYSVHMK peptide for MPH and the LVVDIPAALQHA peptide for SPH exhibited the highest antioxidant activity. The presence of hydrophobic and hydrophilic amino acids, namely leucine (L), valine (V), phenylalanine (F), histidine (H) and proline (P), in the peptide sequences of SPH and MPH are believed to contribute to high antioxidant activity. Hence, SPH and MPH from patin have the potential as a natural functional ingredient in food and pharmaceutical industry.

Keywords: patin (Pangasius sutchi), protein hydrolysates, antioxidative peptides, mass spectrometry

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Authors: Muhammad Muzamil Khan, Asadullah Madni, Nina Filipczek, Jiayi Pan, Nayab Tahir, Hassan Shah, Vladimir Torchilin

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Lipid-polymer hybrid nanoparticles (LPHNP) are delivery systems for controlled drug delivery at tumor sites. The superior biocompatible properties of lipid and structural advantages of polymer can be obtained via this system for controlled drug delivery. In the present study, cisplatin-loaded lipid-chitosan hybrid nanoparticles were formulated by the single step ionic gelation method based on ionic interaction of positively charged chitosan and negatively charged lipid. Formulations with various chitosan to lipid ratio were investigated to obtain the optimal particle size, encapsulation efficiency, and controlled release pattern. Transmission electron microscope and dynamic light scattering analysis demonstrated a size range of 181-245 nm and a zeta potential range of 20-30 mV. Compatibility among the components and the stability of formulation were demonstrated with FTIR analysis and thermal studies, respectively. The therapeutic efficacy and cellular interaction of cisplatin-loaded LPHNP were investigated using in vitro cell-based assays in A2780/ADR ovarian carcinoma cell line. Additionally, the cisplatin loaded LPHNP exhibited a low toxicity profile in rats. The in-vivo pharmacokinetics study also proved a controlled delivery of cisplatin with enhanced mean residual time and half-life. Our studies suggested that the cisplatin-loaded LPHNP being a promising platform for controlled delivery of cisplatin in cancer therapy.

Keywords: cisplatin, lipid-polymer hybrid nanoparticle, chitosan, in vitro cell line study

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Authors: Renata Rank Miranda, Arandi Ginane Bezerra, Ciro Alberto Oliveira Ribeiro, Marco AntôNio Ferreira Randi, Carmen Lúcia Voigt, Lilian Skytte, Kaare Lund Rasmussen, Francisco Filipak Neto, Frank Kjeldsen

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Synergetic and antagonistic effects of drugs are well-known concerns in pharmacological assessments of dose and toxicity. Similar approach should be used in assessing cellular uptake and cytotoxicity of nanoparticles. Since nanoparticles are released into the aquatic environment they may interact with existing xenobiotics. Here we used biochemical assays and quantitative proteomics to assess the cytotoxicity of silver nanoparticles (AgNP) when human hepatoma HepG2 cells were co-exposed to 2 nm AgNP together with either Cd2+ or Hg2+ ions. Time-course experiments (2h, 4h, and 24h) were conducted to assess the first response to the exposure studies. The general trend was that a synergetic toxicological response was observed in cells exposed to both AgNP and Cd2+ or Hg2+, with AgNP and Cd2+ being more toxic. This was observed by a significant increase in the ROS and superoxide level of >35% in the case of AgNP+Cd2+ compared to the sum of responses of AgNP and Cd2+, individually. Metabolic activity and viability also dropped more for AgNP+Cd2+ (>10%) than for AgNP and Cd2+ combined. We used inductively coupled plasma mass spectrometry to investigate if AgNP facilitates larger influx of toxic metal ions into HepG2 cells. Only Hg2+ ions was found to be more efficiently engulfed as the concentration of Hg2+ was found 2.8 times larger compared to exposure experiments with only Hg2+. This effect was not observed for Cd2+. We now continue with deep proteomics studies to obtain wider details on the mechanism of the toxicity related to AgNP, Cd2+, and AgNP+Cd2+, respectively.

Keywords: nanotoxicology, silver nanoparticles, proteomics, human cell line

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Authors: Anish Shah, Steve A. Wakelin, Derrick Moot, Aurélie Laugraud, Hayley J. Ridgway

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A mixed pasture system of ryegrass-clover is used in New Zealand, where clovers are generally inoculated with commercially available strains of rhizobia. The community of rhizobia living in the soil and the way in which they interact with the plant are affected by different biotic and abiotic factors. In general, bacterial richness and diversity in soil varies by soil pH. pH also affects cell physiology and acts as a master variable that controls the wider soil physiochemical conditions such as P availability, Al release and micronutrient availability. As such, pH can have both primary and secondary effects on soil biology and processes. The aim of this work was to investigate the effect of soil pH on the genetic diversity and metabolic profile of Rhizobium leguminosarum strains nodulating clover. Soils were collected from 12 farms across New Zealand which had a pH(water) range of between 4.9 and 7.5, with four acidic (pH 4.9 – 5.5), four ‘neutral’ (5.8 – 6.1) and four alkaline (6.5 – 7.5) soils. Bacteria were recovered from nodules of Trifolium repens (white clover) and T. subterraneum (subterranean clover) grown in the soils. The strains were cultured and screened against a range of pH-amended media to demonstrate whether they were adapted to pH levels similar to their native soils. The strains which showed high relative growth at a given pH (~20% of those isolated) were selected for metabolic and taxonomic profiling. The Omnilog (Biolog Inc., Hayward, CA) phenotype array was used to perform assays on carbon (C) utilisation for selected strains. DNA was extracted from the strains which had differing C utilisation profiles and PCR products for both forward and reverse primers were sequenced for the following genes: 16S rRNA, recA, nodC, nodD and nifH (symbiotic).

Keywords: bacterial diversity, clover, metabolic and taxonomic profiling, pH adaptation, rhizobia

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Authors: Morteza Elsa, Amirhossein Moghanian

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The major aim of this study was to evaluate the effect of CaO content on in vitro hydroxyapatite formation, MC3T3 cells cytotoxicity and proliferation as well as antibacterial efficiency of sol-gel derived SiO₂–CaO–P₂O₅ ternary system. For this purpose, first two grades of bioactive glass (BG); BG-58s (mol%: 60%SiO₂–36%CaO–4%P₂O₅) and BG-68s (mol%: 70%SiO₂–26%CaO–4%P₂O₅)) were synthesized by sol-gel method. Second, the effect of CaO content in their composition on in vitro bioactivity was investigated by soaking the BG-58s and BG-68s powders in simulated body fluid (SBF) for time periods up to 14 days and followed by characterization inductively coupled plasma atomic emission spectrometry (ICP-AES), Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), and scanning electron microscopy (SEM) techniques. Additionally, live/dead staining, 3-(4,5dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), and alkaline phosphatase (ALP) activity assays were conducted respectively, as qualitatively and quantitatively assess for cell viability, proliferation and differentiations of MC3T3 cells in presence of 58s and 68s BGs. Results showed that BG-58s with higher CaO content showed higher in vitro bioactivity with respect to BG-68s. Moreover, the dissolution rate was inversely proportional to oxygen density of the BG. Live/dead assay revealed that both 58s and 68s increased the mean number live cells which were in good accordance with MTT assay. Furthermore, BG-58s showed more potential antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) bacteria. Taken together, BG-58s with enhanced MC3T3 cells proliferation and ALP activity, acceptable bioactivity and significant high antibacterial effect against MRSA bacteria is suggested as a suitable candidate in order to further functionalizing for delivery of therapeutic ions and growth factors in bone tissue engineering.

Keywords: antibacterial, bioactive glass, hydroxyapatite, proliferation, sol-gel processes

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Authors: Suhair Saleh, Ahmad Aljaberi

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Cationic lipid-mediated delivery of nucleic acids represents an exciting approach for developing therapeutically realistic gene medicines. Elucidation of the molecular and formulation requirements for efficient lipofection is a prerequisite to enhance the biological activity of such delivery systems. To this end, the in vitro lipofection activity of the ionizable asymmetric 1,2-dialkoylamidopropane-based derivatives bearing single primary amine group as the cationic head group was evaluated. The electrostatic interactions of these cationic lipids with plasmid DNA in physiologically relevant medium were investigated by means of gel electrophoresis retardation and Eth-Br quenching assays. The effect of the presence of the helper lipid on these interactions was evaluated. The physicochemical properties of these lipids in terms of bilayer fluidity and extent of ionization were investigated using fluorescence anisotropy and surface potential techniques, respectively. The results showed that only the active lipid, 1,2lmp[5], existed in a liquid crystalline state at physiological temperature. Moreover, the extent of ionization of this lipid in assemblies was significantly higher that it's saturated analogues. Inclusion of the helper lipid DOPE improved the encapsulation and association between 1,2lmp[5] and plasmid DNA, which was reflected by the significant boost of lipofection activity of the 1,2lmp[5]/DOPE formulation as compared to the lipid alone. In conclusion, membrane fluidity and sufficient protonation of ionizable cationic lipid are required for efficient association and encapsulation of plasmid DNA and promoting improved in vitro lipofection activity.

Keywords: cationic lipids, gene delivery, lipofection, membrane fluidity, helper lipids, surface potential

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Authors: Pui Hong Chung, Wing Chi Ho, Jun Li, Cyrus Leung, Ek Yeoh

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Background: Cigarette smoking is associated with poor tuberculosis (TB) outcomes in terms of progression of active TB, relapse of TB and TB-related mortality, but the association with latent tuberculosis infection (LTBI) is unclear. The systematic review conducted aimed at studying the association between active smoking and LTBI, and likelihood of dose-response relationship. Methods: Two independent reviewers searched three electronic databases comprising PudMed, Medline by EBSCOHOST, ExcerptaMedica Database (EMBASE), from inception up to 31st Dec 2015 for studies reporting data on current smoking and the LTBI with tuberculin skin test (TST) or interferon-γ release assays (IGRAs) results, comparing the odds ratios (ORs) of outcome measure of TST or IGRAs among current smokers with 95% confidence intervals (CI). Results: Seven studies were identified, including six cross-sectional studies and one longitudinal cohort study. The outcome measures from three studies were in TST, three studies in IGRAs and one for both tests. For TST, OR ranging from 1.39 to 3.40 (95% CI) with all studies shown positive association between cigarette smoking and LTBI. For IGRAs, OR ranging from 0.47 to 1.89 (95% CI) with one study shown the negative association that might be related to impaired interferon-gamma production in immunosuppressive persons. One identified study demonstrated positive dose-response relationship in TST result. Conclusions: Cigarette smoking is likely to be a risk factor of LTBI. There is the important implication for TB and tobacco control program to halt TB by empowering public health policy. Further study is also needed to provide more evidence of the dose-response model/relationship.

Keywords: latent tuberculosis infection, systematic review, active smoking, model

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Authors: Omar Ali, Adesh Ramsubhag, Jayaraj Jayaraman

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Seaweed extracts have been reported as plant biostimulants which could be a safer, organic alternative to harsh pesticides. The incentive to use seaweed-based biostimulants is becoming paramount in sustainable agriculture. The current study, therefore, screened a commercial extract of A. nodosum in tomatoes, cultivated in Trinidad to showcase the multiple beneficial effects. Foliar treatment with an A. nodosum commercial extract led to significant increases in fruit yield and a significant reduction of incidence of bacterial spots and early blight diseases under both greenhouse and field conditions. Investigations were carried out to reveal the possible mechanisms of action of this biostimulant through defense enzyme assays and transcriptome profiling via RNA sequencing of tomato. Studies into disease control mechanisms by A. nodosum showed that the extract stimulated the activity of enzymes such as peroxidase, phenylalanine ammonia-lyase, chitinase, polyphenol oxidase, and β-1,3-glucanase. Additionally, the transcriptome survey revealed the upregulation and enrichment of genes responsible for the biosynthesis of growth hormones, defense enzymes, PR proteins and defense-related secondary metabolites, as well as genes involved in the nutrient mobilization, photosynthesis and primary and secondary metabolic pathways. The results of the transcriptome study also demonstrated the cross-talks between growth and defense responses, confirming the bioelicitor and biostimulant value of seaweed extracts in plants. These effects could potentially implicate the benefits of seaweed extract and validate its usage in sustainable crop production.

Keywords: A. nodosum, biostimulants, elicitor, enzymes, growth responses, seaweeds, tomato, transcriptome analysis

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Authors: Thi Huong Vu, Vijay Jayasena, Zhongxiang Fang, Gary Dykes

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Carob kibble has recently received attention due to the presence of high level of polyphenol antioxidants. The capacity of microorganisms to improve antioxidant activities and total phenolics in carob kibble was investigated in the study. Two types of microorganisms including lactic acid bacteria Lactobacillus plantarum (L. plantarum) and yeast Saccharomyces cerevisiae (S. cerevisiae) were used in single and in their combination as starters. The total phenolic content was determined by the Folin–Ciocalteu method. Antioxidant activities were assessed scavenging capacity using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid (ABTS). The study found that S. cerevisiae alone considerably improved 55% total phenolics content at 15 h, while L. plantarum caused in a loss of 20% through the process. Antioxidant capacity of the yeast-fermented samples significantly increased by 43 % and 10 % in ABTS and DPPH assays, respectively. However, reduction of 13 % and 32 % inhibition were recorded in the carob treated with L. plantarum. In the combination of S. cerevisiae and L. plantarum (1:1), both total phenolic content and antioxidant activity of carob kibble were a similar trend as these of S. cerevisiae single, but a lower improvement. The antioxidant power of the extracts was linearly correlated to their total phenolic contents (R=0.75). The results suggested that S. cerevisiae alone was the better for enhancement of both total phenolic content and antioxidant activity in carob kibble using submerged fermentation. The efficiency of fermentation reached the highest at 15h. Thus submerged fermentation with S. cerevisiae offers a tool with simple and cost effective to further increase the bioactive potential of carob kibble, which is in use for food, cosmetic and pharmaceutical industries.

Keywords: antioxidant activity, carob kibble, lactobacillus plantarum, saccharomyces cerevisiae, total phenolics

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Authors: Eunsong Lee, Gae Baik Kim, Young Pil Kim

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Bisphenol A (BPA) is one of the endocrine disruptors (EDCs), which have been suspected to be associated with reproductive dysfunction and physiological abnormality in human. Since the BPA has been widely used to make plastics and epoxy resins, the leach of BPA from the lining of plastic products has been of major concern, due to its environmental or human exposure issues. The simple detection of BPA based on the self-assembly of aptamer-mediated gold nanoparticles (AuNPs) has been reported elsewhere, yet the detection sensitivity still remains challenging. Here we demonstrate an improved AuNP-based sensor of BPA by using fluorescence-combined AuNP colorimetry in order to overcome the drawback of traditional AuNP sensors. While the anti-BPA aptamer (full length or truncated ssDNA) triggered the self-assembly of unmodified AuNP (citrate-stabilized AuNP) in the presence of BPA at high salt concentrations, no fluorescence signal was observed by the subsequent addition of SYBR Green, due to a small amount of free anti-BPA aptamer. In contrast, the absence of BPA did not cause the self-assembly of AuNPs (no color change by salt-bridged surface stabilization) and high fluorescence signal by SYBP Green, which was due to a large amount of free anti-BPA aptamer. As a result, the quantitative analysis of BPA was achieved using the combination of absorption of AuNP with fluorescence intensity of SYBR green as a function of BPA concentration, which represented more improved detection sensitivity (as low as 1 ppb) than did in the AuNP colorimetric analysis. This method also enabled to detect high BPA in water-soluble extracts from thermal papers with high specificity against BPS and BPF. We suggest that this approach will be alternative for traditional AuNP colorimetric assays in the field of aptamer-based molecular diagnosis.

Keywords: bisphenol A, colorimetric, fluoroscence, gold-aptamer nanobiosensor

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Authors: Ahmed M. Debela, Mayreli Ortiz , Ciara K. O´Sullivan

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The completion of the human genome Project (HGP) has paved the way for mapping the diversity in the overall genome sequence which helps to understand the genetic causes of inherited diseases and susceptibility to drugs or environmental toxins. Arrayed primer extension (APEX) is a microarray based minisequencing strategy for screening disease causing mutations. It is derived from Sanger DNA sequencing and uses fluorescently dideoxynucleotides (ddNTPs) for termination of a growing DNA strand from a primer with its 3´- end designed immediately upstream of a site where single nucleotide polymorphism (SNP) occurs. The use of DNA polymerase offers a very high accuracy and specificity to APEX which in turn happens to be a method of choice for multiplex SNP detection. Coupling the high specificity of this method with the high sensitivity, low cost and compatibility for miniaturization of electrochemical techniques would offer an excellent platform for detection of mutation as well as sequencing of DNA templates. We are developing an electrochemical APEX for the analysis of SNPs found in the MYH7 gene for group of cardiomyopathy patients. ddNTPs were labeled with four different redox active compounds with four distinct potentials. Thiolated oligonucleotide probes were immobilised on gold and glassy carbon substrates which are followed by hybridisation with complementary target DNA just adjacent to the base to be extended by polymerase. Electrochemical interrogation was performed after the incorporation of the redox labelled dedioxynucleotide. The work involved the synthesis and characterisation of the redox labelled ddNTPs, optimisation and characterisation of surface functionalisation strategies and the nucleotide incorporation assays.

Keywords: array based primer extension, labelled ddNTPs, electrochemical, mutations

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Authors: Walaa H. Salama, Azza M. Abdel-Aty, Afaf S. Fahmy

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Background: Egyptian Cobra; Naja haje (Elapidae) is one of most common snakes, widely distributed in Egypt and its envenomation causes multi-organ failure leading to rapid death. Thus, Different medicinal plants showed a protective effect against venom toxicity and may complement the conventional antivenom therapy. Aim: The present study was designed to assess both the antioxidant capacity of methanolic extract of rosemary leaves and evaluate the neutralizing ability of the extract against hepatotoxicity induced by Naja haje venom. Methods: The total phenolic and flavonoid contents and the antioxidant capacity of the methanolic rosemary extract were estimated by DPPH and ABTS Scavenging methods. In addition, the rosemary extract were assessed for anti-venom properties under in vitro and in vivo standard assays. Results: The rosemary extract had high total phenolic and flavonoid content as 12 ± 2 g of gallic acid equivalent per 100 gram of dry weight (g GAE/100g dw) and 5.5 ± 0.8 g of catechin equivalent per 100 grams of dry weight (g CE/100g dw), respectively. In addition, the rosemary extract showed high antioxidant capacity. Furthermore, The rosemary extract were inhibited in vitro the enzymatic activities of phospholipase A₂, L-amino acid oxidase, and hyaluronidase of the venom in a dose-dependent manner. Moreover, indirect hemolytic activity, hepatotoxicity induced by venom were completely neutralized as shown by histological studies. Conclusion: The phenolic compounds of rosemary extract with potential antioxidant activity may be considered as a promising candidate for future therapeutics in snakebite therapy.

Keywords: antioxidant activity, neutralization, phospholipase A₂ enzyme, snake venom

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Authors: Ozlem Bahadir Acikara, Mert Ilhan, Ekin Kurtul, Karel Smejkal, Esra Kupeli Akkol

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Present study is aimed to investigate in vitro inhibitory effects of the extracts prepared from the aerial parts of Podospermum canum (Asteraceae) on hyaluronidase, collagenase, and elastase enzymes using a bioassay-guided fractionation. Inhibitory effects of the extract, sub-extracts, fractions obtained by column chromatography, and isolated compounds on collagenase, elastase, and hyaluronidase were performed by using in vitro enzyme inhibitory assays based on spectrophotometric evaluation. The ethyl acetate and remaining water extracts prepared from the plant displayed significant inhibitory activities on collagenase and elastase, while petroleum ether and chloroform extracts did not show any inhibitory activity. Eleven known compounds: arbutin, 6'-O-caffeoylarbutin, cichoriin, 3,5-dicaffeoylquinic acid methyl ester, apigenin-7-O-β-glucoside, luteolin-7-O-β-glucoside, apigenin-7-O-β-rutinoside, isoorientin, orientin, vitexin, procatechuic acid, and compound 4-hydroxy-benzoic acid 4-(6-O-α-rhamnopyranosyl-β-glucopyranosyl) benzyl ester have been obtained from ethyl acetate sub-extract of the plant through bioassay-guided fractionation and isolation. Results of the present study have revealed that among the isolated compounds, apigenin-7-O-β-glucoside, luteolin-7-O-β-glucoside, apigenin-7-O-β-rutinoside and isoorientin showed potent enzyme inhibitory activities. However, methanolic extract of P. canum displayed a greater inhibitory activity than fractions and isolated compounds both on collagenase and elastase.

Keywords: Asteraceae, collagenase, elastase, hyaluronidase, Podospermum canum

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Authors: Panagiotis T. Kanellos, Vaios T. Karathanos, Andriana C. Kaliora

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Introduction: The role of ghrelin and obestatin in appetite regulation has been investigated. However, data on ghrelin and obestatin changes after food ingestion are negligible. Objective: We aimed at assessing the appetite-regulating hormones, ghrelin, and obestatin, and furthermore calculate ghrelin/obestatin ratio in healthy normal-weight subjects after consumption of raisins. This survey is a comparative study of a glucose control with raisins containing fructose and glucose in similar concentrations as well as fibers. Methodology: Ten apparently healthy subjects who reported no history of glucose intolerance, diabetes, gastrointestinal disorders, or recent use of any antibiotics were enrolled in the study. The raisins used (Vitis vinifera) originate in Greece and are distributed worldwide as Corinthian raisins. In a randomized crossover design, all subjects after an overnight fast consumed, either 50g of glucose diluted in 240 mL of water (control) or 74 g of raisins (sugar content 50 g) with a 5-day interval between individual trials. Vein blood samples were collected at baseline and at 60, 120 and 180 min postprandially. In blood samples ghrelin and obestatin were measured applying specific enzyme linked immuno absorbent assays. Results: The subjects were of mean age 26.3 years, with BMI of 21.6 kg/m2, waist circumference of 77.7 cm, normal serum lipidemic parameters and normal HbA1c levels. Ghrelin levels were significantly lower after raisin consumption compared to glucose at 120 and at 180 min post-ingestion (p= 0.011 and p= 0.035, respectively). However, obestatin did not reach statistical significance between the two interventions. The ghrelin/obestatin ratio was found significantly lower (p=0.020) at 120 min after raisin ingestion compared to control. Conclusion: Two isocaloric foods containing equal amounts of sugars, however with a different composition, have different effects on appetite hormones ghrelin and obestatin in normal-weight healthy subjects.

Keywords: appetite, ghrelin, obestatin, raisins

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Authors: N. G. Vepkhvadze, T. Enukidze

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Anthrax is a fatal disease caused by strains of Bacillus anthracis, a spore-forming gram-positive bacillus that causes the disease anthrax in animals and humans. Anthrax is a zoonotic disease that is also well-recognized as a potential agent of bioterrorism. Infection in humans is extremely rare in the developed world and is generally due to contact with infected animals or contaminated animal products. Testing of this zoonotic disease began in 1907 in Georgia and is still being tested routinely to provide accurate information and efficient testing results at the State Laboratory of Agriculture of Georgia. Each clinical sample is analyzed by RT-PCR and bacteriology methods; this study used Real-Time PCR assays for the detection of B. anthracis that rely on plasmid-encoded targets with a chromosomal marker to correctly differentiate pathogenic strains from non-anthracis Bacillus species. During the period of 2015-2022, the State Laboratory of Agriculture (SLA) tested 250 clinical and environmental (soil) samples from several different regions in Georgia. In total, 61 out of the 250 samples were positive during this period. Based on the results, Anthrax cases are mostly present in Eastern Georgia, with a high density of the population of livestock, specifically in the regions of Kakheti and Kvemo Kartli. All laboratory activities are being performed in accordance with International Quality standards, adhering to biosafety and biosecurity rules by qualified and experienced personnel handling pathogenic agents. Laboratory testing plays the largest role in diagnosing animals with anthrax, which helps pertinent institutions to quickly confirm a diagnosis of anthrax and evaluate the epidemiological situation that generates important data for further responses.

Keywords: animal disease, baccilus anthracis, edp, laboratory molecular diagnostics

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Authors: Hicham Amar, Mustapha El Ghorfi, Yassine Taha, Abdellatif Elghali, Rachid Hakkou, Mostafa Benzaazoua

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Phosphate mine waste rock (PMWR) generated during ore extraction is continuously increasing, resulting in a significant environmental footprint. The main objectives of this study consist of i) elaboration of the sampling strategy of PMWR piles, ii) a mineralogical and chemical characterization of PMWR piles, and iii) 3D block model creation to evaluate the potential valorization of the existing PMWR. Destructive drilling using reverse circulation from 13 drills was used to collect samples for chemical (X-ray fluorescence analysis) and mineralogical assays. The 3D block model was created based on the data set, including chemical data of the realized drills using Datamine RM software. The optical microscopy observations showed that the sandy phosphate from drills in the PMWR piles is characterized by the abundance of carbonate fluorapatite with the presence of calcite, dolomite, and quartz. The mean grade of composite samples was around 19.5±2.7% for P₂O₅. The mean grade of P₂O₅ exhibited an increasing tendency by depth profile from bottom to top of PMWR piles. 3D block model generated with chemical data confirmed the tendency of the mean grades’ variation and may allow a potential selective extraction according to %P₂O₅. The 3D block model of P₂O₅ grade is an efficient sampling approach that confirmed the variation of P₂O₅ grade. This integrated approach for PMWR management will be a helpful tool for decision-making to recover the residual phosphate, adopting the circular economy and sustainability in the phosphate mining industry.

Keywords: 3D modelling, reverse circulation drilling, circular economy, phosphate mine waste rock, sampling

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Authors: Zatil Athaillah, Anastasia Devi, Dian Muzdalifah, Wirasuwasti Nugrahani, Linar Udin

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During tempe fermentation, some chemical changes occurred and they contributed to sensory, appearance, and health benefits of soybeans. Many studies on health properties of tempe have specialized on their isoflavones. In this study, other components of tempe, particularly water soluble chemicals, was investigated for their biofunctionality. The study was focused on the ability to suppress MCF-7 breast cancer cell growth and antioxidant activity, as expressed by DPPH radical scavenging activity, total phenols and total flavonoids, of the water extracts. Fermentation time of tempe was extended up to 120 hr to increase the possibility to find the functional components. Extraction yield and soluble nitrogen content were also quantified as accompanying data. Our findings suggested that yield of water extraction of tempe increased as fermentation was extended up to 120 hr, except for a slight decrease at 72 hr. Water extracts of tempe showed inhibition of MCF-7 breast cancer cell growth, as shown by lower IC50 values when compared to control (unfermented soybeans). Among the varied fermentation timescales, 60-hr period showed the highest activity (IC50 of 8.7 ± 4.95 µg/ml). The anticancer activity of extracts obtained from different fermentation time was positively correlated with total soluble nitrogens, but less relevant with antioxidant data. During 48-72 hr fermentation, at which cancer suppression activity was significant, the antioxidant properties from the three assays were not higher than control. These findings indicated that water extracts of tempe from extended fermentation could inhibit breast cancer cell growth but further study to determine the mechanism and compounds that play important role in the activity should be conducted.

Keywords: tempe, anticancer, antioxidant, phenolic compounds

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Authors: William L. Whitehouse, Louisa H. Y. Lo, Andrew B. Kinghorn, Simon C. C. Shiu, Julian. A. Tanner

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C-reactive protein (CRP) is an acute-phase reactant and sensitive indicator for sepsis and other life-threatening pathologies, including systemic inflammatory response syndrome (SIRS). Currently, clinical turn-around times for established CRP detection methods take between 30 minutes to hours or even days from centralized laboratories. Here, we report the development of an electrochemical biosensor using redox probe-tagged DNA aptamers functionalized onto cheap, commercially available screen-printed electrodes. Binding-induced conformational switching of the CRP-targeting aptamer induces a specific and selective signal-ON event, which enables single-step and reagentless detection of CRP in as little as 1 minute. The aptasensor dynamic range spans 5-1000nM (R=0.97) or 5-500nM (R=0.99) in 50% diluted human serum, with a LOD of 3nM, corresponding to 2-orders of magnitude sensitivity under the clinically relevant cut-off for CRP. The sensor is stable for up to one week and can be reused numerous times, as judged from repeated real-time dosing and dose-response assays. By decoupling binding events from the signal induction mechanism, structure-switching electrochemical aptamer-based sensors (SS-EABs) provide considerable advantages over their adsorption-based counterparts. Our work expands on the retinue of such sensors reported in the literature and is the first instance of an SS-EAB for reagentless CRP detection. We hope this study can inspire further investigations into the suitability of SS-EABs for diagnostics, which will aid translational R&D toward fully realized devices aimed at point-of-care applications or for use more broadly by the public.

Keywords: structure-switching, C-reactive protein, electrochemical, biosensor, aptasensor.

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Authors: Daniel Assefa, Engeda Dessalegn, Chetan Chauhan

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Moringa stenopetala is a socioeconomic valued tree that is widely available and cultivated in the Southern part of Ethiopia. The leaves have been traditionally used as a food source with high nutritional and medicinal values. The present work was carried out to evaluate the effect of thermal treatment on the total phenolic content, antioxidant and alpha-amylase inhibition activities of aqueous leaf extracts during maceration and different decoction time interval (5, 10 and 15 min). The total phenolic content was determined by the Folin-ciocalteu methods whereas antioxidant activities were determined by 2,2-diphenyl-1-picryl-hydrazyl(DPPH) radical scavenging, reducing power and ferrous ion chelating assays and alpha-amylase inhibition activity was determined using 3,5-dinitrosalicylic acid method. Total phenolic content ranged from 34.35 to 39.47 mgGAE/g. Decoction for 10 min extract showed ferrous ion chelating (92.52), DPPH radical scavenging (91.52%), alpha-amylase inhibition (69.06%) and ferric reducing power (0.765), respectively. DPPH, reducing power and alpha-amylase inhibition activities showed positive linear correlation (R2=0.853, R2= 0.857 and R2=0.930), respectively with total phenolic content but ferrous ion chelating activity was found to be weakly correlated (R2=0.481). Based on the present investigation, it could be concluded that major loss of total phenolic content, antioxidant and alpha-amylase inhibition activities of the crude leaf extracts of Moringa stenopetala leaves were observed at decoction time for 15 min. Therefore, to maintain the total phenolic content, antioxidant, and alpha-amylase inhibition activities of leaves, cooking practice should be at the optimum decoction time (5-10 min).

Keywords: alpha-amylase inhibition, antioxidant, Moringa stenopetala, total phenolic content

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Authors: Kyungae Jo, Eun Young Kim, Byungsoo Shin, Kwang Soon Shin, Hyung Joo Suh

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Gamma-aminobutyric acid (GABA) and 5-hydroxytryptophan (5-HTP) are amino acids of digested nutrients or food ingredients and these can possibly be utilized as non-pharmacologic treatment for sleep disorder. We previously investigated the GABA/5-HTP mixture is the principal concept of sleep-promoting and activity-repressing management in nervous system of D. melanogaster. Two experiments in this study were designed to evaluate sleep-promoting effect of GABA/5-HTP mixture, to clarify the possible ratio of sleep-promoting action in the Drosophila invertebrate model system. Behavioral assays were applied to investigate distance traveled, velocity, movement, mobility, turn angle, angular velocity and meander of two amino acids and GABA/5-HTP mixture with caffeine treated flies. In addition, differentially expressed gene (DEG) analyses from next generation sequencing (NGS) were applied to investigate the signaling pathway and functional interaction network of GABA/5-HTP mixture administration. GABA/5-HTP mixture resulted in significant differences between groups related to behavior (p < 0.01) and significantly induced locomotor activity in the awake model (p < 0.05). As a result of the sequencing, the molecular function of various genes has relationship with motor activity and biological regulation. These results showed that GABA/5-HTP mixture administration significantly involved the inhibition of motor behavior. In this regard, we successfully demonstrated that using a GABA/5-HTP mixture modulates locomotor activity to a greater extent than single administration of each amino acid, and that this modulation occurs via the neuronal system, neurotransmitter release cycle and transmission across chemical synapses.

Keywords: sleep, γ-aminobutyric acid, 5-hydroxytryptophan, Drosophila melanogaster

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Authors: Jae Bok Heo, Yun Mi Lee, Hee Rang Yun

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Several GTPases are required for ribosome biogenesis and assembly. We recently characterized rice (Oryza sativa) nuclear/nucleolar GTPase 2 (OsNug2), belonging to the YlqF/YawG family of GTPases, as playing a role in pre-60S ribosomal subunit maturation. To investigate the potential factors involved in regulating the function of OsNug2, yeast two-hybrid screens were carried out using OsNug2 as bait. Rice serine/threonine kinase 1 (OsSTK1) was identified as a potential interacting protein candidate. In vitro pull down and bimolecular fluorescence complementation assays confirmed the interaction between OsNug2 and OsSTK1, and like green fluorescent protein-tagged OsNug2, green fluorescent protein-tagged OsSTK1 was targeted to the nucleus of Arabidopsis protoplasts. OsSTK1 was not found to affect the GTP-binding activity of OsNug2; however, when recombinant OsSTK1 was included in OsNug2 assay reaction mixtures, OsSTK1 increased the GTPase activity of OsNug2. To test whether OsSTK1 phosphorylates OsNug2 in vitro, a kinase assay was performed. OsSTK1 was found to have weak autophosphorylation activity and strongly phosphorylated serine 209 of OsNug2. Yeast complementation testing resulted in a GAL::OsNug2(S209N) mutant-harboring yeast strain exhibiting a growth-defective phenotype on galactose medium at 39°C, divergent from that of a yeast strain harboring GAL::OsNug2. The intrinsic GTPase activity of mutant OsNug2(S209N) was found to be similar to that of OsNug2, was not fully enhanced upon weak binding of OsSTK1. Our findings reported here indicate that OsSTK1 functions as a positive regulator protein of OsNug2 by enhancing the GTPase activity of OsNug2, and that the phosphorylation of serine 209 of OsNug2 is essential for the complete function of OsNug2 in ribosome biogenesis.

Keywords: OsSTK1, OsNug2, GTPase activity, GTP binding activity, phosphorylation

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Authors: Arun Jyothi Bheemagani, Chakrapani Pullagummi, Anupalli Roja Rani

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Exploration of the chemical constituents of the plants and pharmacological screening may provide us the basis for the development of novel agents. Plants have provided us some of the very important life saving drugs used in the modern medicine. The aim of our work was to screen the phytochemical constituents and antimicrobial and antioxidant activities of methanol extract of leaves of Annona cherimola Mill plant from Tirumala forest, Tirupathi. It was originally called Chirimuya by the Inca people who lived where it was growing in the Andes of South America, is an edible fruit-bearing species of the genus Annona from the family Annonaceae. Annona cherimola Mill is a multipurpose tree with edible fruits and is one of the sources of the medicinal products. The antibacterial activity was measured by agar well diffusion method; the diameter of the zone of bacterial growth inhibition was measured after incubation of plates. The inhibitory effect was studied against the pathogenic bacteria (Klebsiella pneumonia, Bacillus subtilis, Staphylococcus aureus and Escherichia coli (E. coli). Antioxidant assays were also performed for the same extracts by spectrophotometric methods using known standard antioxidants as reference. The studied plant extracts were found to be very effective against the pathogenic microorganisms tested. The methanolic extract of Annona cherimola Mill from showed maximum activity against Escherichia coli and Staphylococcus aureus and the least concentration required showing the activity was 0.5mg/ml. Phytochemical screening of the plants revealed the presence of flavonoids, alkaloids, steroids and carbohydrates. Good presence of antioxidants was also found in the methanolic extracts.

Keywords: annona cherimola, phytochemicals, antioxidant and antibacterial activity, methanol extract

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Authors: Maria L. G. Lourenco, Viviane Y. Hibaru, Keylla H. N. P. Pereira, Fabiana F. Souza, Joao C. P. Ferreira, Simone B. Chiacchio, Luiz H. A. Machado

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Neonatal conjunctivitis, or ophthalmia, is an infection of the conjunctiva or cornea before opening the eyelids. It is believed that immunodeficiency contributes to the development of the condition. This study aims at reporting a case of ophthalmia neonatorum in a dog, in addition to its diagnosis and treatment. A litter of five pug neonates was admitted to the Sao Paulo State University (UNESP) Veterinary Hospital, Botucatu, Sao Paulo, Brazil, with complaints of ocular secretion. The neonates were five days old. The clinical examination revealed that three newborns presented swelling in the ocular region and a purulent secretion in the medial corner of the eye that was exerting pressure on the ocular globes, which are compatible with the description of this disease. The diagnosis was made based on the clinical signs and bacterial culture of the secretion, which revealed the presence of bacteria belonging to the genus Staphylococcus sp. The laboratory assays did not reveal any alterations. The treatment was instituted gently, opening the eyelids early and cleaning the purulent ocular secretion with saline solution. An ophthalmic ointment with retinol, amino acids, methionine, and chloramphenicol (Epitezan®) was prescribed four times a day for seven days. Blood plasma (2 mL/100 g) was administered subcutaneously because bacterial infections in neonates may represent a failure in the transference of passive immunity. A more thorough cleaning of the environment was also recommended. Neonatal conjunctivitis has a simple diagnosis and treatment. If not treated early, it can evolve to adherence of the eyelids to the cornea, ulceration, and perforation of the cornea. Therefore, the prognosis is favorable as long as the condition is diagnosed early, and the treatment is instituted quickly.

Keywords: ophthalmia neonatorum, neonatal infection, puppy, newborn

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Authors: Abdelkarim Mahdhi, Fdhila Kais, Faouzi Lamari, Zeineb Hmila, Fathi Kamoun, Maria Ángeles Esteban, Amina Bakhrouf

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Aquaculture is the world’s fastest growing food-production sector. However, one of the most serious problems regarding the culture of marine fishes is the mortality associated with pathogenic bacteria that occurs in the critical phases of larval development. Conventional approaches, such as the use of antimicrobial drugs to control diseases, have had limited success in the prevention or cure of aquatic diseases. Promising alternatives to antibiotics are probiotics, which are food supplements consisting of live microorganisms that benefit the host organism. In the search for more effective and environmentally friendly treatments with probionts against pathogenic species in shrimp larval culture, the probiotic properties of Bacillus strains isolated from Artemia culture such as antibacterial activity, adhesion, pathogenicity, toxicity and the effect of marine stress on viability and survival were investigated, as well as the changes occurring in their properties. Analyses showed that these bacteria corresponded to the genus Bacillus sp. Antagonism and adherence assays revealed that these strains have an inhibitory effect against pathogenic bacteria in vitro and in vivo conditions and are fairly adherent. Challenge tests performed with Artemia larvae provided evidence that the tested Bacillus strains were neither pathogenic nor toxic to the host. The tested strains maintained their viability and their probiotic properties during the period of study. The results suggest that the tested strains have suffered changes allowing them to survive in seawater in the absence of nutrients and outside their natural host, identifying them as potential probiotic candidates for Artemia culture.

Keywords: bacillus, probiotic, cell viability, stress response

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Authors: Suhas Kadam, Vishal Chandanshive, Niraj Rane, Sanjay Govindwar

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Fimbristylis dichotoma, Ammannia baccifera, and their co-plantation consortium FA were found to degrade methyl orange, simulated dye mixture, and real textile effluent. Wild plants of Fimbristylis dichotoma and Ammannia baccifera with equal biomass showed 91 and 89% decolorization of methyl orange within 60 h at a concentration of 50 ppm, while 95% dye removal was achieved by consortium FA within 48 h. Floating phyto-beds with co-plantation (Fimbristylis dichotoma and Ammannia baccifera) for the treatment of real textile effluent in a constructed wetland was observed to be more efficient and achieved 79, 72, 77, 66 and 56% reductions in ADMI color value, chemical oxygen demand, biological oxygen demand, total dissolve solid and total suspended solid of textile effluent, respectively. High performance thin layer chromatography, gas chromatography-mass spectroscopy, Fourier transform infrared spectroscopy, Ultra violet-Visible spectroscopy and enzymatic assays confirmed the phytotransformation of parent dye in the new metabolites. T-RFLP analysis of rhizospheric bacteria of Fimbristylis dichotoma, Ammannia baccifera, and consortium FA revealed the presence of 88, 98 and 223 genera which could have been involved in dye removal. Toxicity evaluation of products formed after phytotransformation of methyl orange by consortium FA on bivalves Lamellidens marginalis revealed less damage in the gills architecture when analyzed histologically. Toxicity measurement by Random Amplification of Polymorphic DNA (RAPD) technique revealed normal banding pattern in treated methyl orange sample suggesting less toxic nature of phytotransformed dye products.

Keywords: constructed wetland, phyto-bed, textile effluent, phytoremediation

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