Molecular Weights of Polyethyleneimine-Dependent Physicochemical Tuning of Gold Nanoparticles and FRET-Based Turn-On Sensing of Polymyxin B.

Environmental monitoring and the detection of antibiotic contaminants require expensive and time-consuming techniques. To overcome these challenges, gold nanoparticle-mediated fluorometric "turn-on" detection of Polymyxin B (PMB) in an aqueous medium was undertaken. The molecular weight of polyethyleneimine (PEI)-dependent physicochemical tuning of gold nanoparticles (PEI@AuNPs) was achieved and employed for the same. The three variable molecular weights of branched polyethyleneimine (MW 750, 60, and 1.3 kDa) molecules controlled the nano-geometry of the gold nanoparticles along with enhanced stabilization at room temperature. The synthesized gold nanoparticles were characterized through various advanced techniques. The results revealed that polyethyleneimine-stabilized gold nanoparticles (PEI@AuNP-1-3) were 4.5, 7.0, and 52.5 nm in size with spherical shapes, and the zeta potential values were 29.9, 22.5, and 16.6 mV, respectively. Accordingly, the PEI@AuNPs probes demonstrated high sensitivity and selectivity, with a linear relationship curve over a concentration range of 1-6 µM for polymyxin B. The limit of detection (LOD) was calculated as 8.5 nM. This is the first unique report of gold nanoparticle nano-geometry-dependent FRET-based turn-on detection of PMB in an aqueous medium. We believe that this approach would offer a complementary strategy for the development of a highly sophisticated and advanced sensing system for PMB and act as a template for the development of new nanomaterial-based engineered sensors for rapid antibiotic detection in environmental as well as biological samples.

Aggregation-Resistant, Turn-On-Off Fluorometric Sensing of Glutathione and Nickel (II) Using Vancomycin-Conjugated Gold Nanoparticles.

Glutathione (GSH) and nickel (II) cation have an indispensable role in various physiological processes, including preventing the oxidative damage of cells and acting as a cofactor for lipid metabolic enzymes. An imbalance in the physiological level of these species may cause serious health complications. Therefore, sensitive and selective fluorescent probes for the detection of GSH and nickel (II) are of great interest for clinical as well as environmental monitoring. Herein, vancomycin-conjugated gold nanoparticles (PEI-AuNP@Van) were prepared and employed for the detection of GSH and nickel (II) based on a turn-on-off mechanism. The as-synthesized PEI-AuNP@Van was ~7.5 nm in size; it exhibited a spherical shape with face-centered cubic lattice symmetry. As compared to vancomycin unconjugated gold nanoparticles, GSH led to the turn-on state of PEI-AuNP@Van, while Ni2+ acted as a fluorescence quencher (turn-off) without the aggregation of nanoparticles. These phenomena strongly justify the active role of vancomycin conjugation for the detection of GSH and Ni2+. The turn-on-off kinetics was linearly proportional over the concentration range between 0.05-0.8 µM and 0.05-6.4 µM. The detection limits were 205.9 and 90.5 nM for GSH and Ni2+, respectively; these results are excellent in comparison to previous reports. This study demonstrates the active role of vancomycin conjugation for sensing of GSH and Ni2+ along with PEI-AuNP@Van as a promising nanoprobe.

Synthesis of vancomycin functionalized fluorescent gold nanoparticles and selective sensing of mercury (II).

Mercury ions (Hg2+) are widely found in the environment; it is considered a major pollutant. Therefore, the rapid and reliable detection of Hg2+ is of great technical interest. In this study, a highly fluorescent, sensitive, and selective fluorometric assay for detecting Hg2+ ions was developed using vancomycin functionalized and polyethyleneimine stabilized gold nanoparticles (PEI-f-AuNPs@Van). The as-made gold nanoparticles were highly fluorescent, with excitation and emission maxima occurring at 320 and 418 nm, respectively. The size of nanoparticles was ~7 nm; a zeta potential of ~38.8 mV was determined. The XRD analysis confirmed that the nanoparticles possessed crystalline structure with face centerd cubic symmetry. Using the PEI-f-AuNP@Van probe, the detection limit of Hg2+ ion was achieved up to 0.988 nM (within a linear range) by calculating the KSV. However, the detection limit in a natural environmental sample was shown to be 12.5 nM. Furthermore, the selectivity tests confirmed that the designed probe was highly selective to mercury (II) cations among tested other divalent cations. Owing to its sensitivity and selectivity, this approach for Hg2+ ions detection can be utilized for the analysis of real water samples.

A whole cell fluorescence quenching-based approach for the investigation of polyethyleneimine functionalized silver nanoparticles interaction with Candida albicans.

The antimicrobial activity of metal nanoparticles can be considered a two-step process. In the first step, nanoparticles interact with the cell surface; the second step involves the implementation of the microbicidal processes. Silver nanoparticles have been widely explored for their antimicrobial activity against many pathogens. The interaction dynamics of functionalized silver nanoparticles at the biological interface must be better understood to develop surface-tuned biocompatible nanomaterial-containing formulations with selective antimicrobial activity. Herein, this study used the intrinsic fluorescence of whole C. albicans cells as a molecular probe to understand the cell surface interaction dynamics of polyethyleneimine-functionalized silver nanoparticles and antifungal mechanism of the same. The results demonstrated that synthesized PEI-f-Ag-NPs were ~ 5.6 ± 1.2 nm in size and exhibited a crystalline structure. Furthermore, the recorded zeta potential (+18.2 mV) was associated with the stability of NPS and shown a strong electrostatic interaction tendency between the negatively charged cell surface. Thus, rapid killing kinetics was observed, with a remarkably low MIC value of 5 µg/mL. PEI-f-Ag-NPs quenched the intrinsic fluorescence of C. albicans cells with increasing incubation time and concentration and have shown saturation effect within 120 min. The calculated binding constant (Kb = 1 × 105 M-1, n = 1.01) indicated strong binding tendency of PEI-f-Ag-NPs with C. albicans surface. It should also be noted that the silver nanoparticles interacted more selectively with the tyrosine-rich proteins in the fungal cell. However, calcofluor white fluorescence quenching showed non-specific binding on the cell surface. Thus, the antifungal mechanisms of PEI-f-Ag-NPs were observed as reactive oxygen species (ROS) overproduction and cell wall pit formation. This study demonstrated the utility of fluorescence spectroscopy for qualitative analysis of polyethyleneimine-functionalized silver nanoparticle interaction/binding with C. albicans cell surface biomolecules. Although, a quantitative approach is needed to better understand the interaction dynamics in order to formulate selective surface tuned nanoparticle for selective antifungal activity.

Rhizopus homothallicus rhino-orbital-cerebral mucormycosis: Six cases from a tertiary care Centre, North India.

OBJECTIVE: To report six cases of Rhizopus homothallicus rhino-orbital-cerebral mucormycosis in North India between April 2021 and July 2021. CASE DETAILS: All six patients had diabetes, concomitant SARS-CoV-2 infection, a history of oxygen requirement and steroid intake. Among these six cases 4 were female. All patients presented with sinus pain and peri-orbital swelling. COVID-19-associated mucormycosis (CAM) was diagnosed based on microbiological examination of the biopsied tissue, and its staging was determined radiologically by CT and MRI. Three patients were in stage III-C, the others were in stage II-C, II-D and IV-A. A multidisciplinary team treated the patients with extensive surgical debridement of the affected tissue, correction of predisposing comorbidities and administration of an antifungal agents. Patients were followed up for 6 months with routine direct nasal endoscopy to check the sinonasal cavity for any recurrence. All the six patients survived at 6 months of follow-up. CONCLUSION: A timely initiated multidisciplinary team-based approach can reduce the mortality in rhino-orbital-cerebral mucormycosis cases caused by R. homothallicus.

COVID-19-associated Mucormycosis: A clinico-epidemiological study.

BACKGROUND: There was an unprecedented increase in COVID-19-associated-Mucormycosis (CAM) cases during the second pandemic wave in India. METHODS: This observational study was done to know the epidemiological profile of CAM cases andincluded all patients admitted with mucormycosis between May 2021 and July 2021. RESULTS: Out of the enrolled 208 CAM cases (either SARS-CoV-2 RT-PCR or serology positive), 204, three and one had rhino-orbital-cerebral, pulmonary and gastrointestinal mucormycosis, respectively. 95.7 % of the patients had diabetes, out of which 42.3 % were recently diagnosed. Mean HbA1c was 10.16 ± 2.56 %. 82.5 % of the patients were unvaccinated. During their COVID-19 illness, 86.5 % were prescribed antibiotics, 84.6 % zinc preparations, 76.4 % ivermectin, and 64.9 % steroids, while only 39.5 % required oxygen therapy. The frequency of blood groups A, B, O and AB in our CAM patients was 29.5 %, 18.9 %, 38.9 % &12.6 %, respectively. At three months follow up, 60 (28.8 %) patients died, four (1.9 %) stopped antifungal treatment, and 144(69.23 %) were on antifungal treatment. 55 % (n = 33) of deaths occurred within 15 days of admission. Mortality was significantly associated with higher age, RT-PCR positive for SARS-CoV-2, raised serum creatinine and alkaline phosphatase during treatment. At 6 months follow-up, eight more patients died, three due to chronic kidney disease, four patients who had stopped treatment and one patient who was on a ventilator due to COVID-19 associated pneumonia and the rest 140(67.3 %) survived. CONCLUSION: Uncontrolled hyperglycemia, SARS-CoV-2 infection, rampant use of antibiotics, zinc supplementation and steroids were some of the risk factors for mucormycosis. Despite the overwhelming number of patients with an uncommon disease like mucormycosis, the six months mortality was much lower than expected.

Size and Zeta Potential Clicked Germination Attenuation and Anti-Sporangiospores Activity of PEI-Functionalized Silver Nanoparticles against COVID-19 Associated Mucorales (Rhizopus arrhizus).

The SARS-CoV-2 infections in Indian people have been associated with a mucormycotic fungal infection caused by the filamentous fungi Rhizopus arrhizus. The sporangiospores of R. arrhizus are omnipresent in the environment and cause infection through inhalation or ingestion of contaminated air and foods. Therefore, the anti-sporangiospore activity of polyethyleneimine functionalized silver nanoparticles (PEI-f-Ag-NPs) with variable size and surface charge as a function of the molecular weight of PEI was explored. The results showed that both PEI-f-AgNP-1 and PEI-f-AgNP-2, potentially, attenuated the germination and reduced the viability of sporangiospores. Furthermore, the results showed that the minimum inhibitory concentration (MIC) values of both PEI-f-AgNP-1 and PEI-f-AgNP-2 (1.65 and 6.50 µg/mL, respectively) were dependent on the nanoparticle size and surface ζ potentials. Similarly, the sporangiospore germination inhibition at MIC values was recorded, showing 97.33% and 94% germination inhibition, respectively, by PEI-f-AgNP-1 and 2 within 24 h, respectively. The confocal laser scanning microscopy, SEM-EDS, and confocal Raman spectroscopy investigation of PEI-f-Ag-NPs treated sporangiospores confirmed size and surface charge-dependent killing dynamics in sporangiospores. To the best of our knowledge, this is the first investigation of the polyethyleneimine functionalized silver nanoparticle-mediated size and surface charge-dependent anti-sporangiospore activity against R. arrhizus, along with a possible antifungal mechanism.

Amine-Functionalized Silver Nanoparticles: A Potential Antiviral-Coating Material with Trap and Kill Efficiency to Combat Viral Dissemination (COVID-19).

The outbreak of COVID-19 has drastically affected the daily lifestyles of people globally where specific Coronavirus-2 transmits primarily by respiratory droplets. Structurally, the SARS-CoV-2 virus is made up of four types of proteins in which S-protein is indispensable among them, as it causes rapid replication in the host body. Therefore, the glycine and alanine composed of HR1 of S-protein is the ideal target for antiviral action. Different forms of surface-active PPEs can efficiently prevent this transmission in this circumstance. However, the virus can survive on the conventional PPEs for a long time. Hence, the nanotechnological approaches based on engineered nanomaterials coating on medical equipments can potentially prevent the dissemination of infections in public. Silver nanoparticles with tuneable physicochemical properties and versatile chemical functionalization provide an excellent platform to combat the disease. The coating of amine-functionalized silver nanoparticle (especially amine linked to aliphatic chain and trialkoxysilane) in its nanostructured form enables cloths trap and kill efficient. PPEs are a primary and reliable preventive measure, although they are not 100% effective against viral infections. So, developing and commercializing surface-active PPEs with trap and kill efficacy is highly needed to cope with current and future viral infections. This review article discusses the COVID-19 morphology, antiviral mechanism of Ag-NPs against SARS-CoV-2 virus, surface factors that influence viral persistence on fomites, the necessity of antiviral PPEs, and the potential application of amine-functionalized silver nanoparticles as a coating material for the development of trap and kill-efficient face masks and PPE kits.

Galactose-Clicked Curcumin-Mediated Reversal of Meropenem Resistance among Klebsiella pneumoniae by Targeting Its Carbapenemases and the AcrAB-TolC Efflux System.

In over eighty years, despite successive antibiotics discoveries, the rapid advent of multidrug resistance among bacterial pathogens has jolted our misapprehension of success over them. Resistance is spreading faster than the discovery of new antibiotics/antimicrobials. Therefore, the search for better antimicrobials/additives becomes prudent. A water-soluble curcumin derivative (Curaq) was synthesised, employing a Cu (I) catalysed 1, 3-cyclo addition reaction; it has been evaluated as a potential treatment for multidrug-resistant isolates and as an antibiotic adjuvant for meropenem against hypervirulent multidrug-resistant Klebsiella pneumoniae isolates. We also investigated its solubility and effect over carbapenemase activity. Additionally, we investigated its impact on the AcrAB-TolC system. We found that Curaq inhibited bacterial growth at a minimal concentration of 16 µg/mL; at a 32 µg/mL concentration, it killed bacterial growth completely. Only nine (9.4%) Klebsiella isolates were sensitive to meropenem; however, after synergising with Curaq (8 µg/mL), 85 (88.54%) hvKP isolates became sensitive to the drug. The Curaq also inhibited the AcrAB-TolC efflux system at 1 µg/mL concentration by disrupting the membrane potential and causing depolarisation. The kinetic parameters obtained also indicated its promise as a carbapenemase inhibitor. These results suggest that Curaq can be an excellent drug candidate as a broad-spectrum antibacterial and anti-efflux agent.

Utility of nested polymerase chain reaction for fungus in detecting clinically suspected patients of invasive fungal infections and its clinical correlation and comparison with fungal culture.

AIMS AND OBJECTIVES: The aim and objective of this study is to detect invasive fungal infections (IFIs) early and with more sensitivity by the nested polymerase chain reaction (PCR) for fungus as compared to fungal culture in clinically suspected patients and also explore its correlation in reference to age, duration of symptoms, immunocompromised status, and other risk factors predisposing to IFIs. MATERIALS AND METHODS: In this cross-sectional study, 50 suspected patients admitted in medical acute care unit/intensive care unit (ACU/ICU) of Sir Sunderlal Hospital, Banaras Hindu University, Varanasi, India, comprised the study. All cases were selected based on the predefined inclusion and exclusion criteria. A detailed history, clinical examination, and all required investigations were done in all suspected patients. Blood samples were taken for nested-PCR for fungus and culture. Nested PCR was performed on extracted DNA form samples collected from all participants under the study. RESULTS: Our study comprised of 50 suspected immunocompromised patients of IFIs. Among the participants under the study, the most common risk factor was diabetes mellitus (28% cases). Nearly two-thirds (60%) of the cases were 50 years or more. Around 70% of the cases had a history of illness more than 2 weeks. Nested PCR for fungus came out to be positive in 21/50 patients (42%); however, fungal culture was positive in none. Among the admitted patient in ACU/ICU, 75% were neutropenic. CONCLUSIONS: IFIs are more common in immunocompromised individuals, patients with comorbidities, long history of symptoms, and elderly population. Nested PCR for fungus has a high sensitivity (as compared to the fungal culture), and also they are rapid in giving the results. Thus, nested PCR for fungus can be used in a cost-effective manner for the early and reliable diagnosis of clinically suspected IFIs.

Making of water soluble curcumin to potentiate conventional antimicrobials by inducing apoptosis-like phenomena among drug-resistant bacteria.

The upsurge of multidrug resistant bacterial infections with declining pipeline of newer antibiotics has made it imperative to develop newer molecules or tailor the existing molecules for more effective antimicrobial therapies. Since antiquity, the use of curcumin, in the form of Curcuma longa paste, to treat infectious lesions is unperturbed despite its grave limitations like instability and aqueous insolubility. Here, we utilized "click" chemistry to address both the issues along with improvisation of its antibacterial and antibiofilm profile. We show that soluble curcumin disrupts several bacterial cellular processes leading to the Fenton's chemistry mediated increased production of reactive oxygen species and increased membrane permeability of both Gram-positive and Gram-negative bacteria. We here report that its ability to induce oxidative stress can be harnessed to potentiate activities of ciprofloxacin, meropenem, and vancomycin. In addition, we demonstrated that the soluble curcumin reported herein even sensitizes resistant Gram-negative clinical isolates to the Gram-positive specific antibiotic vancomycin, thereby expanding the antibacterial spectrum of this drug. This work shows that the soluble curcumin can be used to enhance the action of existing antimicrobials against both Gram-positive and Gram-negative bacteria thus strengthening the antibiotic arsenal for fighting resistant bacterial infections for many years to come.

Molecular weight of polyethylenimine-dependent transfusion and selective antimicrobial activity of functional silver nanoparticles.

Synthetic cationic polymer-mediated synthesis of silver nanoparticles and selective antimicrobial activity of the same were demonstrated. Polyethyleneimine (PEI)-coated silver nanoparticles showed antimicrobial activity against Acinetobacter baumannii as a function of the polymeric molecular weight (MW) of PEI. Silver nanoparticles were coated with PEI of three different MWs: Ag-NP-1 with PEI exhibiting a MW of 750,000, Ag-NP-2 with PEI exhibiting a MW of 1300, and Ag-NP-3 with PEI exhibiting a MW of 60,000. These nanoparticles showed a particle size distribution of 4-20 nm. The nanoparticles exhibited potent antimicrobial activity against A. baumannii, with the minimum inhibitory concentration of Ag-NP-1, Ag-NP-2, and Ag-NP-3 on the order of 5, 10, and 5 µg/mL, respectively, and minimum bactericidal concentration of Ag-NP-1, Ag-NP-2, and Ag-NP-3 on the order of 10, 20, and 10 µg/mL, respectively. Fluorescence imaging of Ag-NPs revealed selective transfusion of Ag-NPs across the cell membrane as a function of the polymeric MW; differential interaction of the cytoplasmic proteins during antimicrobial activity was observed.

Extracellular hydrolytic enzyme activities of the different Candida spp. isolated from the blood of the Intensive Care Unit-admitted patients.

BACKGROUND: Candida spp. secretes various extracellular hydrolytic enzymes. These enzymes are the important virulence factor for the pathogenesis of Candida. We assessed four different enzymatic activities of Candida isolates obtained from bloodstream infections. MATERIALS AND METHODS: We isolated 79 strains of different Candida species from the blood of the Intensive Care Unit-admitted patients. Species were identified by conventional methods including culture characteristic, germ tube, sugar assimilation, and Dalmau's culture technique. Phospholipase, proteinase, hemolysin, and esterase enzymatic activities were determined by the Plate method. RESULTS: Non albicans candida were the most common isolates from the blood of the ICU admitted patient with a predominance of Candida tropicalis. Hemolytic activity was the most prominent enzyme activity followed by the proteinase activity. Candida albicans (89.86%) was the major proteinase producer, while 95.8% of C. tropicalis produced hemolysin. No esterase activity was shown by the Candida glabrata and Candida krusei. CONCLUSION: No significant difference was observed between the two most common causative agents of candidemia: C. albicans and C. tropicalis.

Quantum curcumin mediated inhibition of gingipains and mixed-biofilm of Porphyromonas gingivalis causing chronic periodontitis.

Periodontitis is a biofilm-associated irreversible inflammation of the periodontal tissues. Reports suggest the role of Porphyromonas gingivalis specific Arg- and Lys-specific proteinases in the orchestration of the initiation and progression of periodontal diseases. These proteinases are precisely termed as gingipains R and K. Curcumin is an active polyphenol that is extracted from the rhizomes of Curcuma longa. However, the molecule curcumin owing to its high hydropathy index and poor stability has not been able to justify its role as frontline drug modality in the treatment of infectious and non-infectious diseases as claimed by several investigators. In the present study, at first, we synthesized and characterized quantum curcumin, and investigated its biocompatibility. This was subsequently followed by the evaluation of the role of quantum curcumin as an antimicrobial, anti-gingipains and antibiofilm agent against Porphyromonas gingivalis and select reference strains. We have successfully synthesized the quantum curcumin utilizing a top-down approach with the average size of 3.5 nm. Apart from its potent antimicrobial as well as antibiofilm properties, it also significantly inhibited the gingipains in a dose-dependent manner. At the minimal concentration of 17.826 µM, inhibition up to 98.7% and 89.4% was noted for gingipain R and K respectively. The data was also supported by the in silico docking experiments which revealed high exothermic enthalpies (-7.01 and -7.02 cal mol-1). Besides, the inhibition constant was found to be 7.24 µM and 7.1 µM against gingipains R and K respectively. The results suggest that quantum curcumin is a potential drug candidate which needs further clinical validation.

Correction: Quantum curcumin mediated inhibition of gingipains and mixed-biofilm of Porphyromonas gingivalis causing chronic periodontitis.

[This corrects the article DOI: 10.1039/C8RA08435A.].

White grain mycetoma caused by Scedosporium apiospermum in North India: a case report.

Mycetoma is chronic granulomatous infection of skin and subcutaneous tissue caused by both bacteria and fungi. We report a case of mycetoma caused by Scedosporium apiospermum in the right foot of a 45-year-old farmer in north India. The patient had a history of trauma in the sole of the right foot followed by discharge of white granules along with proximal progression. Scedosporium apiospermum was identified based on colony characteristics and microscopic features on slide culture. Mycetoma is a progressive disease. Foot is commonly affected in persons who walk barefoot, especially in south India. Untreated mycetoma progress and involve the underlying fasciae and tissues along with bones often resulting in loss of limb. By prompt and reliable diagnosis with corresponding antimicrobial administration, we can prevent further progression and limb disability.