Rapid identification of bacteria by the pattern of redox reactions rate using 2',7'-dichlorodihydrofluorescein diacetate.

Bacterial infection is a life-threatening situation, and its rapid diagnosis is essential for treatment. Apart from medical applications, rapid identification of bacteria is vital in the food industry or the public health system. There are various bacterial identification techniques, including molecular-based methods, immunological approaches, and biosensor-based procedures. The most commonly used methods are culture-based methods, which are time-consuming. The objective of this study is to find a fingerprint of bacteria to identify them. Three strains of bacteria were selected, and seven different concentrations of each bacterium were prepared. The bacteria were then treated with two different molar concentrations of the fluorescent fluorophore, dichlorodihydrofluorescein diacetate for 30 minutes. Then, using the fluorescence mode of a multimode reader, the fluorescence emission of each bacterium is scanned twice during 60 minutes. Plotting the difference between two scans versus the bacteria concentration results in a unique fluorescence pattern for each bacterium. Observation of the redox state of bacteria, during 90 minutes, results in a fluorescence pattern that is clearly a fingerprint of different bacteria. This pattern is independent of fluorophore concentration. Mean Squares Errors (MSE) between the fluorescence patterns of similar bacteria is less than that of different bacteria, which shows the method can properly identify the bacteria. In this study, a new label-free method is developed to detect and identify different species of bacteria by measuring the redox activity and using the fluorescence fluorophore, dichlorodihydrofluorescein diacetate. This robust and low-cost method can properly identify the bacteria, uses only one excitation and emission wavelength, and can be simply implemented with current multimode plate readers.

Crosslinked hydrogel loaded with chitosan-supported iron oxide and silver nanoparticles as burn wound dressing.

Burns can result in infection, disability, psychosocial and economic issues. Advanced wound dressings like hydrogel absorb exudate and maintain moisture. Considering the antimicrobial properties of silver nanoparticles and iron oxide nanoparticles, the efficiency of cross-linked hydrogel loaded with chitosan-supported iron oxide and silver nanoparticles for burn wounds repair was investigated in animal model. Cellulose hydrogel dressing made from carboxymethylcellulose and hydroxyethylcellulose crosslinked with different concentrations of citric acid (10, 15, 20, and 30%) was produced. The physicochemical characteristics of the synthetized hydrogels including Fourier-Transform Infrared spectroscopy, Thermal behavior, Swelling properties, and Scanning Electron Microscope (SEM) were evaluated. The silver nanoparticles and iron nanoparticles were produced and the characteristics, cytotoxicity, antimicrobial activities and their synergistic effect were investigated. After adding nanoparticles to hydrogels, the effects of the prepared wound dressings were investigated in a 14-day animal model of burn wound. The results showed that the mixture comprising 12.5 ppm AgNps, and IONPs at a concentration ≤100 ppm was non-cytotoxic. Moreover, the formulations with 20% CA had a swelling ratio of almost 250, 340, and 500 g/g at pHs of 5, 6.2, and 7.4 after one hour, which are lower than those of formulations with 5 and 10% CA. The total mass loss (59.31%) and the exothermic degradation happened in the range of 273-335 °C and its Tm was observed at 318.52 °C for hydrogels with 20% CA. Thus, the dressing comprising 20% CA which was loaded with 12.5 ppm silver nanoparticles (AgNPs) and 100 ppm iron oxide nanoparticles (IONPs) indicated better physicochemical, microbial and non-cytotoxic characteristics, and accelerated the process of wound healing after 14 days. It was concluded that the crosslinked hydrogel loaded with 12.5 ppm AgNPs and 100 ppm IONPs possesses great wound healing activity and could be regarded as an effective topical burn wound healing treatment.

From plants to antimicrobials: Natural products against bacterial membranes.

Bacterial membrane barrier provides a cytoplasmic environment for organelles of bacteria. The membrane is composed of lipid compounds containing phosphatide protein and a minimal amount of sugars, and is responsible for intercellular transfers of chemicals. Several antimicrobials have been found that affect bacterial cytoplasmic membranes. These compounds generally disrupt the organization of the membrane or perforate it. By destroying the membrane, the drugs can permeate and replace the effective macromolecules necessary for cell life. Furthermore, they can disrupt electrical gradients of the cells through impairment of the membrane integrity. In recent years, considering the spread of microbial resistance and the side effects of antibiotics, natural antimicrobial compounds have been studied by researchers extensively. These molecules are the best alternative for controlling bacterial infections and reducing drug resistance due to the lack of severe side effects, low cost of production, and biocompatibility. Better understanding of the natural compounds' mechanisms against bacteria provides improved strategies for antimicrobial therapies. In this review, natural products with antibacterial activities focusing on membrane damaging mechanisms were described. However, further high-quality research studies are needed to confirm the clinical efficacy of these natural products.

New Insight into Vitamins E and K1 as Anti-Quorum-Sensing Agents against Pseudomonas aeruginosa.

Today, antivirulence compounds that attenuate bacterial pathogenicity and have no interference with bacterial viability or growth are introduced as the next generation of antibacterial agents. However, the development of such compounds that can be used by humans is restricted by various factors, including the need for extensive economic investments, the inability of many molecules to penetrate the membrane of Gram-negative bacteria, and unfavorable pharmacological properties and cytotoxicity. Here, we take a new and different look into two frequent supplements, vitamin E and K1, as anti-quorum-sensing agents against Pseudomonas aeruginosa, a pathogen that is hazardous to human life and responsible for several diseases. Both vitamins showed significant anti-biofilm activity (62% and 40.3% reduction by vitamin E and K1, respectively), and the expression of virulence factors, including pyocyanin, pyoverdine, and protease, was significantly inhibited, especially in the presence of vitamin E. Cotreatment of constructed biofilms with these vitamins plus tobramycin significantly reduced the number of bacterial cells sheltered inside the impermeable matrix (71.6% and 69% by a combination of tobramycin and vitamin E or K1, respectively). The in silico studies, besides the similarities of chemical structures, reinforce the possibility that both vitamins act through inhibition of the PqsR protein. This is the first report of the antivirulence and antipathogenic activity of vitamin E and K1 against P. aeruginosa and confirms their potential for further research against other multidrug-resistant bacteria.

Anti-quorum sensing potential of ketoprofen and its derivatives against Pseudomonas aeruginosa: insights to in silico and in vitro studies.

Antibiotics are usually used for the treatment of bacterial infections, but multidrug-resistant strains are a phenomenon that has been growing at an increasing rate worldwide. Thus, there is an increasing need for novel strategies for combatting infectious diseases. Many pathogenic bacteria apply quorum sensing (QS) to regulate their pathogenicity and virulence factors production. This circuit makes the QS system an attractive target for antibacterial therapy. In the present study, an important member of non-steroidal anti-inflammatory drugs (NSAIDs), by reducing the biofilm and producing QS-regulated virulence factors, ketoprofen and its synthetic derivatives were screened against the Pseudomonas aeruginosa PAO1. All compounds showed anti-biofilm activity (16-79%) and most of them presented anti-virulence activity. In the co-treatment of ketoprofen, G20, G21, or G77 with tobramycin, biofilm is significantly reduced (potentiated to > 50%) in the number of cells protected inside the impermeable matrix. The in silico studies in addition to the similarities between the chemical structures of PqsR natural ligands and ketoprofen derivatives reinforce the possibility that the mechanism of action is through PqsR inhibition. Based on the results, the anti-pathogenic effect was more appreciable in ketoprofen, G77, and G20.

Antimicrobial and cytotoxic activity of extracts from Salvia tebesana Bunge and Salvia sclareopsis Bornm cultivated in Iran.

Salvia, a member of the Lamiaceae family, represents more than 58 species in Iran. In the present study, antibacterial and cytotoxic activity of extracts obtained from the roots of Salvia tebesana and Salvia sclareopsis were investigated. The antibacterial activity of the extracts was investigated against 4 bacterial strains and yeast using serial dilution method. The petroleum ether and CH2Cl2 extracts of S. tebesana showed a good activity against Gram-positive bacteria particularly Bacillus cereus (MIC 1.25 mg/mL) while Gram-negative bacteria and yeast were resistant to the extracts. Also, the cytotoxic effects of the extracts on A2780 (ovarian), MCF-7 (breast) and DU 145 (prostate) cancer cell lines were examined using AlamarBlue® assay. The petroleum ether and CH2Cl2 extracts of S. tebesana were found to be cytotoxic against the tested cell lines, with IC50 values less than 50 µg/mL. The petroleum ether extract also showed a potent anti-proliferative activity against DU 145 cells with the lowest IC50 value (6.25 µg/mL).

Selection of specific aptamer against enrofloxacin and fabrication of graphene oxide based label-free fluorescent assay.

Specific ssDNA aptamers for the antibiotic enrofloxacin (ENR) were isolated from an enriched nucleotide library by SELEX (Systematic Evolution of Ligands by EXponential enrichment) method with high binding affinity. After seven rounds, five aptamers were selected and identified. Apt58 with highest affinity and sensitivity (Kd = 14.19 nM) was employed to develop a label-free fluorescent biosensing approach based on aptamer, graphene oxide (GO) and native fluorescence of ENR for determination of ENR residue in raw milk samples. Under optimized experimental conditions, the linear range was from 5 nM to 250 nM and LOD was calculated to be 3.7 nM, and the recovery rate was between 94.1% and 108.5%. The integration of aptamer and GO in this bioassay provides a promising way for rapid, sensitive and cost-effective detection of ENR in real samples like raw milk.

Identification and biological activity of the volatile compounds of Glycyrrhiza triphylla Fisch. & C.A.Mey.

Chemical composition and biological (antimicrobial, antioxidant and cytotoxic) activities of essential oils (EO) obtained from the aerial parts of Glycyrrhiza triphylla Fisch. & C.A.Mey (G. triphylla) were evaluated in the present study. The EO was isolated and analyzed using gas chromatography-mass spectrometry (GC-MS). Fifty-five compounds representing 99.3% of the total oil composition were identified. Major components of the oil were ß-caryophyllene (25.4%), limonene (16.7%), ß-myrcene (16.0%) and α-humulene (4.4%). The oil composition was dominated by the presence of sesquiterpene hydrocarbons comprising 43.6% of the total oil. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the oil were determined against eight bacterial strains and one fungus. The EO showed a good antibacterial activity against both Gram-positive and Gram-negative bacteria. The most susceptible strain was Micrococcus luteus (MIC = 2.7 µg/mL, MBC = 43.6 µg/mL). The antioxidant potential of the EO was examined using DPPH and ß-carotene/linoleic acid (BCB) assays. The oil was considerably active in the DPPH assay (IC50 = 100.40 ± 0.03 µg/mL). Moreover, in vitro cytotoxic activity was assessed against six cancer cell lines using MTT assay. The EO showed no significant cytotoxic activity. In light of the present findings, G. triphylla oil may deserves to be further investigated for its potential therapeutic effects and also as a natural preservative in food industry.

Investigation of Pseudomonas aeruginosa quorum-sensing signaling system for identifying multiple inhibitors using molecular docking and structural analysis methodology.

Pseudomonas aeruginosa is an opportunistic human pathogen and a common Gram-negative bacterium in hospital-acquired infections. It causes death in many burn victims, cystic-fibrosis and neutropenic-cancer patients. It is known that P. aeruginosa biofilm maturation and production of cell-associated and extracellular virulence factors such as pyocyanin, elastase and rhamnolipids are under the control of a quorum-sensing (QS) system. Among several proteins involved in the Pseudomonas QS mechanism, LasR and PqsE play an important role in its cascade signaling system. They can cause increases in QS factors, biofilm maturation, and the production of virulence factors. Therefore, inhibition of these proteins can reduce the pathogenicity of P. aeruginosa. According to the structure of corresponding auto-inducers bound to these proteins, in silico calculations were performed with some non-steroidal anti-inflammatory drugs (NSAIDs) to estimate possible interactions and find the co-inhibitors of LasR and PqsE. The results showed that oxicams (Piroxicam and Meloxicam) can interact well with active sites of both proteins with the Ki of 119.43 nM and 4.0 µM for Meloxicam and 201.39 nM and 4.88 µM against LasR and PqsE, respectively. These findings suggested that Piroxicam and Meloxicam can be used as potential inhibitors for control of the P. aeruginosa QS signaling system and biofilm formation, and may be used in the design of multiple inhibitors.

The role of transient receptor potential (TRP) channels in phagocytosis: A comprehensive review.

When host cells are exposed to foreign particles, dead cells, or cell hazards, a sophisticated process called phagocytosis begins. During this process, macrophages, dendritic cells, and neutrophils engulf the target by expanding their membranes. Phagocytosis of apoptotic cells is called efferocytosis. This process is of significant importance as billions of cells are eliminated daily without provoking inflammation. Both phagocytosis and efferocytosis depend on Ca2+ signaling. A big family of Ca2+ permeable channels is transient receptor potentials (TRPs) divided into nine subfamilies. We aimed to review their roles in phagocytosis. The present review article shows that various TRP channels such as TRPV1, 2, 3, 4, TRPM2, 4, 7, 8, TRPML1, TRPA1, TRPC1, 3, 5, 6 have roles at various stages of phagocytosis. They are involved in the phagocytosis of amyloid ß, α-synuclein, myelin debris, bacteria, and apoptotic cells. In particular, TRPC3 and TRPM7 contribute to efferocytosis. These effects are mediated by changing Ca2+ signaling or targeting intracellular enzymes such as Akt. In addition, they contribute to the chemotaxis of phagocytic cells towards targets. Although a limited number of studies have assessed the role of TRP channels in phagocytosis and efferocytosis, their findings indicate that they have critical roles in these processes. In some cases, their ablation completely abolished the phagocytic function of the cells. As a result, TRP channels are potential targets for developing new therapeutics that modulate phagocytosis.

Natural-derived acetophenones: chemistry and pharmacological activities.

Acetophenones are naturally occurring phenolic compounds which have found in over 24 plant families and also fungi strains. They are exist in both free or glycosides form in nature. The biological activities of these compounds have been assayed and reported including cytotoxicity, antimicrobial, antimalarial, antioxidant and antityrosinase activities. Herein, we review the chemistry and biological activity of natural acetophenone derivatives that have been isolated and identified until January 2024. Taken together, it was reported 252 acetophenone derivatives in which the genera Melicope (69) and Acronychia (44) were the principal species as producers of acetophenones.

In vitro antimicrobial and antibiofilm screening of eighteen Iranian medicinal plants.

BACKGROUND: Natural products are one of the best candidates for controlling drug-resistant pathogens, the advantages of which include low production costs and low side effects. In this study, as potential antimicrobials, the anti-bacterial and antibiofilm activities of several Iranian native medicinal plants were screened. METHODS: The antibacterial/antifungal and anti-biofilm activities of 18 medicinal plants including Reseda lutea L., Nepeta sintenisii Bunge., Stachys turcomanica Trautv., Stachys lavandulifolia Vahl, Diarthron antoninae (Pobed.) Kit Tan., Ziziphora clinopodioides Lam., Euphorbia kopetdaghi Prokh, Euphorbia serpens Kunth., Hymenocrater calycinus Benth., Scutellaria pinnatifida A.Ham., Viola tricolor L., Hypericum helianthemoides (Spach) Boiss., Hypericum scabrum L., Convolvulus lineatus L., Scabiosa rotata M.Bieb Greuter & Burdet, Delphinium semibarbatum Bien. Ex Boiss., Glycyrrhiza triphylla Fisch. & C.A.Mey., and Ziziphus jujuba Mill., against two Gram-positive bacteria, Staphylococcus aureus, Bacillus cereus, as well as two Gram-negative bacteria, Pseudomonas aeruginosa, Escherichia coli; and Candida albicans as a fungal strain, were evaluated. The minimum inhibitory concentration (MIC) and minimum bactericidal/fungicidal concentration (MBC/MFC) values of the extracts against tested microorganisms were reported and we investigated their effect on the biofilm inhibition of Pseudomonas aeruginosa PAO1, Staphylococcus epidermis, Staphylococcus aureus and Streptococcus mutans. In addition, the effect of the extracts on the eradication of the biofilms of these bacteria was evaluated. RESULTS: In this study, H. scabrum was found to exhibit potentially significant activity against Gram-positive bacteria with the MIC range of 6.25-25 µg/mL. This extract also showed a significant effect on inhibiting the biofilm of S. aureus, S. mutans, and S. epidermidis and eradicating the biofilm of S. epidermidis DSMZ 3270. In addition, Hymenocrater calycinus root extract had moderate antibacterial activity against B. cereus with the MIC and MBC 62.5 µg/mL, respectively. CONCLUSIONS: The results of this study showed that the root extracts of two plants, Hypericum scabrum and Hymenocrater calycinus, had antimicrobial and anti-biofilm effects. Based on the observed anti-biofilm effects, these two plants may be considered in future studies to find responsible antimicrobial compounds.

The potential therapeutic impact of a topical bacteriophage preparation in treating Pseudomonas aeruginosa-infected burn wounds in mice.

Aim: This study compared a topical formulation containing lytic phages with a routine antibiotic in the murine model of burn/Pseudomonas aeruginosa infected wound healing. Methods & Materials: Isolated and purified lytic bacteriophages from hospital sewage were added to the polyethylene glycol (PEG) based ointment. A second-degree burned wound on the back of twenty-four adult female mice was created. The wounds were infected subcutaneously with 100 µL of 1 × 102-3 CFU/mL P. aeruginosa. After 24 h, mice were randomly assigned to one of four groups: mice received a standard antibiotic (antibiotic-treated group), mice received an ointment without bacteriophage (PEG-based group), mice received a PEG-ointment with bacteriophage (bacteriophage-treated group), or mice received no treatment (untreated-control group). Every two days, the contraction of burned wounds, physical activity, and rectal body temperature were recorded. On day 10, mice were sacrificed, and the wounds were cut off and evaluated histopathologically. Results: In ointments containing PEG, bacteriophages were active and stable. The mice receiving bacteriophage and PEG-based ointment had substantially different wound contraction in primary wound healing (P = 0.001). When compared to the control group, the bacteriophage-treated group showed significant variations in wound contraction (P = 0.001). The wound contraction changed significantly between the antibiotic and PEG-based groups (P = 0.002). In all groups, physical activity in mice improved over time, with significant differences (P = 0.001). When the 8th day was compared to the days 2, 4, and 6, significant changes were found (P = 0.001, P = 0.02, and P = 0.02, respectively). Both the positive control and bacteriophage-treated groups showed perfect wound healing histopathologically. However, no significant variations in microscopic histopathological criteria were found between the groups. Conclusion: Formulated phage ointment could be a promising approach for treating infected burn wounds infected by P. aeruginosa in mice with no allergic reactions.

Anti-quorum sensing and antibiofilm activity of coumarin derivatives against Pseudomonas aeruginosa PAO1: Insights from in vitro and in silico studies.

Objectives: Biofilm-associated infections are challenging to manage or treat since the biofilm matrix is impenetrable to most antibiotics. Therefore, the best approach to deal with biofilm infections is to interrupt the construction during the initial levels. Biofilm formation has been regulated through the quorum sensing (QS) network, making it an attractive target for any antibacterial therapy. Materials and Methods: Here, some coumarin members, including umbelliprenin, 4-farnesyloxycoumarin, gummosin, samarcandin, farnesifrol A, B, C, and auraptan, have been assessed as QS inhibitors in silico and in vitro. Their potential inhibitory effects on biofilm formation and virulence factor production of Pseudomonas aeruginosa PAO1 were evaluated. Results: First, the interaction of these compounds was investigated against one of the major transcriptional regulator proteins, PqsR, using molecular docking and structural analysis methodology. After that, in vitro evaluations indicated that 4-farnesyloxycoumarin and farnesifrol B showed considerable reduction in biofilm formation (62% and 56%, respectively), virulence factor production, and synergistic effects with tobramycin. Moreover, 4-farnesyloxycoumarin significantly (99.5%) reduced PqsR gene expression. Conclusion: The biofilm formation test, virulence factors production assays, gene expression analysis, and molecular dynamic simulations data demonstrated that coumarin derivatives are a potential anti-QS family through PqsR inhibition.

Evaluating the Antivirulence Effects of New Thiazolidinedione Compounds Against Pseudomonas aeruginosa PAO1.

Pseudomonas aeruginosa is an opportunistic pathogen that causes several serious health problems and numerous forms of virulence. During the treatment of P. aeruginosa infections, the development of multidrug-resistant isolates creates significant clinical problems. Using antivirulence compounds to disrupt pathogenicity rather than killing the bacterium may be an interesting strategy to overcome this problem, because less harsh conditions will exist for the development of resistance. To reduce pathogenicity and biofilm formation, newly synthesized analogs of imidazolyl (8n) and previously synthesized analogs (8a-8m) with a similar backbone [the 5-(imidazolyl-methyl) thiazolidinediones] were tested against pyoverdine and pyocyanin production, protease activity, and biofilm formation. Compared to the positive control group, the best compounds reduced the production of pyoverdine (8n) by 89.57% and pyocyanin (8i) by 22.68%, and protease activity (8n) by 2.80% for PAO1 strain, at a concentration of 10 µM. Moreover, the biofilm formation assay showed a reduction of 87.94% (8i) for PAO1, as well as 30.53% (8d) and 44.65% (8m) for 1074 and 1707 strains, respectively. The compounds used in this study did not show any toxicity in the human dermal fibroblasts and 4T1 cells (viability higher than 90%). The in silico study of these compounds revealed that their antivirulence activity could be due to their interaction with the PqsR, PqsE, and LasR receptors.

A Review of Routine Laboratory Biomarkers for the Detection of Severe COVID-19 Disease.

As the COVID-19 pandemic continues, there is an urgent need to identify clinical and laboratory predictors of disease severity and prognosis. Once the coronavirus enters the cell, it triggers additional events via different signaling pathways. Cellular and molecular deregulation evoked by coronavirus infection can manifest as changes in laboratory findings. Understanding the relationship between laboratory biomarkers and COVID-19 outcomes would help in developing a risk-stratified approach to the treatment of patients with this disease. The purpose of this review is to investigate the role of hematological (white blood cell (WBC), lymphocyte, and neutrophil count, neutrophil-to-lymphocyte ratio (NLR), platelet, and red blood cell (RBC) count), inflammatory (C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), and lactate dehydrogenase (LDH)), and biochemical (Albumin, aspartate aminotransferase (AST) and alanine aminotransferase (ALT), blood urea nitrogen (BUN), creatinine, D-dimer, total Cholesterol, low-density lipoprotein (LDL), and high-density lipoprotein (HDL)) biomarkers in the pathogenesis of COVID-19 disease and how their levels vary according to disease severity.

Bacterial biofilms and their resistance mechanisms: a brief look at treatment with natural agents.

Biofilm is a complex community of microorganisms residing within a polysaccharide and/or protein matrix. Biofilm can be produced by several microorganisms, including various bacteria and fungi. Nowadays, the resistance of biofilm-growing cells to antimicrobials originated from the structural nature of biofilms, and phenotypic alteration of sessile cells is becoming a global issue. Bacterial biofilms are important in various aspects of human health, including chronic infections, dental plaque, and infection of indwelling medical devices such as catheters. They are also a major problem in other industries, including oil recovery, drinking water distribution, papermaking, metalworking, and food processing. Estimates indicate that more than 80% of infectious diseases are biofilm-derived. The aim of this study is to describe mechanisms of antibiotic resistance to provide a better perspective on how to manage it. Moreover, the current strategies for biofilm inhibition were described. Considering that plants are a valuable source of abundant natural chemicals to create prophylactic and therapeutic medicines against biofilm-based infections, significant natural compounds with anti-biofilm properties were highlighted. Finally, natural anti-biofilm compounds under clinical trial evaluation were summarized to provide a background for more extensive researches and assist in opening a new window to novel treatments.

Ultra selective and high-capacity dummy template molecular imprinted polymer to control quorum sensing and biofilm formation of Pseudomonas aeruginosa.

Here a highly selective molecular imprinting polymer was developed to attenuate biofilm formation of the multidrug-resistant pathogen Pseudomonas aeruginosa by disrupting the intermolecular signaling system. Firstly, a dummy template molecular imprinting polymer (MIP) was rationally designed through molecular modeling to capture 2-heptyl-3-hydroxy-4-quinolone (Pseudomonas quinolone signal). This multifunctional signaling molecule interferes with the pathogenicity of P. aeruginosa as an auto-inducer. Then, the synthesized MIP and the non-imprinted polymer (NIP) as reference polymer were evaluated for their binding capacity and biofilm inhibition. The results indicated a significant difference in biofilm inhibition (∼56%) between imprinted (∼67%) and non-imprinted (∼11%) polymer, which is an impressive level, especially for the treatment of various surfaces affected by P. aeruginosa. These results open a new window in the special biological application of MIPs as a promising candidate to reduce concerns in clinical or industrial issues by preventing microbial infections.

A fluorescent sensing strategy for ultrasensitive detection of oxytetracycline in milk based on aptamer-magnetic bead conjugate, complementary strand of aptamer and PicoGreen.

Misuse of antibiotics in animal husbandry and presence of their residues in animal foods is a serious crisis worldwide and thus, monitoring the level of them in food samples is vital for human health. Herein, a fluorescent aptasensor was developed for highly sensitive quantification of oxytetracycline (OTC) in food samples. This method is based on OTC aptamer conjugated to magnetic beads, functioned as recognition element, complementary strand of OTC aptamer, and PicoGreen (PG) as a sensitive double-stranded DNA (dsDNA) fluorescent dye. Formation of OTC aptamer-magnetic bead conjugate provides the opportunity of sample condensation and separation technology. Additionally, the presence of complementary strand leads to significant fluorescence signal alteration of aptasensor in the presence or absence of target and a noteworthy improvement of the aptasensor sensitivity. In the absence of target, complementary strand could bind to aptamer and form dsDNA on the surface of magnetic bead. As a consequence, adding PG to the sample leads to observation of high fluorescence signal from sample. In contrast, once OTC is added to the sample, it binds to OTC aptamer-magnetic bead complex and prevents hybridization of OTC aptamer and its complementary strand. Hence, after addition of PG to the sample, a weak fluorescence intensity is measured. Under optimized conditions, the linear ranges for OTC detection were 0.2-2 nM and 2-800 nM. The detection limit was calculated to be as low as 0.15 nM for the fabricated aptasensor. Besides the great sensitivity, proposed method demonstrated superior specificity towards OTC once it was used against several antibiotics. More significantly, the recovery rates of OTC in milk ranged from 96.46% to 101.5%, implying the great feasibility of designed sensor as well as its potential to be employed for analysis of OTC in real samples.

Point-of-care detection of Escherichia coli O157:H7 in water using AuNPs-based aptasensor.

OBJECTIVES: Access to safe drinking and irrigation water has always been one of the major human concerns worldwide. Thus, rapid, sensitive, and inexpensive approaches for pathogenic bacteria detection, such as Escherichia coli O157:H7 (EHEC) that can induce important infectious diseases, are highly on demand. MATERIALS AND METHODS: In this study, a sensitive aptamer-based AuNPs bioassay was developed that demonstrated its potential to detect EHEC. In the presence of the target bacterium, the specific adsorbed aptamer, leaves AuNPs surface and interacts with EHEC. The bare nanoparticles aggregate in the presence of NaCl and the color shifts from red to purple and blue depending on the bacterial concentration. RESULTS: The proposed aptasensor exhibited a good linear response over a wide concentration range of 876 to 107 CFU/ml and was closely correlated with the line equation of "y=0.0094x+0.0006" (R2= 0.9861). It also showed a low detection limit (LOD) of 263 CFU/ml (Signal/Noise=3). No response was recorded in the presence of other tested bacterial strains including Listeria monocytogenes and Salmonella typhi, indicating the high selectivity of the aptasensor. CONCLUSION: This biosensor may be used as a smart device to screen water reservoirs and prevents the outbreak of EHEC-related life-threatening contagious diseases.