Fragment-based virtual screening identifies novel leads against Plasmepsin IX (PlmIX) of Plasmodium falciparum: Homology modeling, molecular docking, and simulation approaches.

Despite continuous efforts to develop safer and efficient medications, malaria remains a major threat posing great challenges for new drug discovery. The emerging drug resistance, increased toxicities, and impoverished pharmacokinetic profiles exhibited by conventional drugs have hindered the search for new entities. Plasmepsins, a group of Plasmodium-specific, aspartic acid protease enzymes, are involved in many key aspects of parasite biology, and this makes them interesting targets for antimalarial chemotherapy. Among different isoforms, PlmIX serves as an unexplored antimalarial drug target that plays a crucial role along with PlmV and X in the parasite's survival by digesting hemoglobin in the host's erythrocytes. In this study, fragment-based virtual screening was performed by modeling the three-dimensional structure of PlmIX and predicting its ligand-binding pocket by using the Sitemap tool. Screening identified the fragments with the XP docking score ≤ -3 kcal/mol from the OTAVA General Fragment Library (≈16,397 fragments), and the selected fragments were chosen for ligand breeding. The resulting ligands (≈69,858 ligands) were subsequently subjected to filtering based on the QikProp properties along with carcinogenicity testing performed using CarcinoPred-EL and then docked in the SP (≈14,078 ligands) as well as XP mode (≈3,104 ligands), and compared with that of control ligands 49C and I0L. The top-ranked ligands were taken further for the calculation of the free energy of binding using Prime MM-GBSA. Overall, a total of six complexes were taken further for MD simulation studies performed at 100 ns to attain a better understanding of the binding mechanisms, and compounds 3 and 4 were found to be the most efficient ones in silico. The analysis of compound 3 revealed that the carbonyl group present in position 1 on the isoindoline moiety (Arg554) was responsible for inhibitory activity against PlmIX. However, the analysis of compound 4 revealed that the amide linkage sandwiched between the phenyl ring and isoquinoline moiety (Lys555 and Ser226) as well as carbonyl oxygen of the carbamoyl group present at position 2 of the pyrazole ring (Gln222) were responsible for PlmIX inhibitory activity, owing to their crucial interactions with key amino acid residues.

Molecular epidemiology and molecular typing methods of Acinetobacter baumannii: An updated review.

The aim of this study was to go through the molecular methods used for typing of carbapenem-resistant Acientobacter baumannii (CRAB) isolates for investigating the molecular epidemiology all over the world. Multiple typing techniques are required to understand the source and nature of outbreaks caused by Acientobacter baumannii (A. baumannii) and acquired resistance to antimicrobials. Nowadays, there is gradual shift from traditional typing methods to modern molecular methods to study molecular epidemiology and infection control. Molecular typing of A. baumannii strains has been revolutionized significantly in the last 2 decades. A few sequencing-based techniques have been proven as a breakthrough and opened new prospects, which have not been achieved by the traditional methods. In this review, discussed different pre-existing and recently used typing methods to explore the molecular epidemiology of A. baumannii pertaining in context with human infections.

Preparation and Evaluation of Nanoemulsion of Citronella Essential Oil with Improved Antimicrobial and Anti-Cancer Properties.

The development of new pharmaceutical solutions for treating various diseases results from a growing understanding of the benefits of using essential oils. One of the most often used volatile materials among essential oils is the oil of the citronella plant, termed citronella essential oil (CITEO), which has potential for use in food and medicine. Its wide use is limited due to lipophilicity, high volatility and poor physicochemical stability. With this background, the present study aims to evaluate the properties of CITEO-nanoemulsion (CITEO-NE) by analyzing its antimicrobial activities against Staphylococcus aureus (S. aureus) and Candida albicans (C. albicans) and its anticancer activity against, human skin adenocarcinoma cell line (A431). The CITEO-NE was prepared and evaluated for the size range of 130 ± 5 nm, polydispersity index (PDI) of 0.127 and zeta potential -12.6 mV. The percentage % of entrapment efficiency (%EE) of nanoemulsions loaded with CIT was very high at the beginning of the study, at 95.5 ± 4.775%. The MIC was observed to be 500 µg/mL for CITEO and 250 µg/mL for CITEO-NE against S. aureus and 250 µg/mL for CITEO and 125 µg/mL for CITEO-NE against C. albicans. The time-kill assay also suggests the effectiveness of CITEO-NE against the test pathogens as a novel alternative therapy. The IC50 values of CITEO and CITEO-NE exhibited significant cytotoxic properties against the A431 cell line, with 41.20 µg/mL and 37.71 µg/mL, respectively. Hence, our findings revealed that encapsulation of CITEO increased the pharmacological properties.

Molecular typing methods & resistance mechanisms of MDR Klebsiella pneumoniae.

The emergence and transmission of carbapenem-resistant Klebsiella pneumoniae (CRKP) have been recognized as a major public health concern. Here, we investigated the molecular epidemiology and its correlation with the mechanisms of resistance in CRKP isolates by compiling studies on the molecular epidemiology of CRKP strains worldwide. CRKP is increasing worldwide, with poorly characterized epidemiology in many parts of the world. Biofilm formation, high efflux pump gene expression, elevated rates of resistance, and the presence of different virulence factors in various clones of K. pneumoniae strains are important health concerns in clinical settings. A wide range of techniques has been implemented to study the global epidemiology of CRKP, such as conjugation assays, 16S-23S rDNA, string tests, capsular genotyping, multilocus sequence typing, whole-genome sequencing-based surveys, sequence-based PCR, and pulsed-field gel electrophoresis. There is an urgent need to conduct global epidemiological studies on multidrug-resistant infections of K. pneumoniae across all healthcare institutions worldwide to develop infection prevention and control strategies. In this review, we discuss different typing methods and resistance mechanisms to explore the epidemiology of K. pneumoniae pertaining to human infections.

MALDI-TOF MS for Rapid Analysis of Bacterial Pathogens Causing Urinary Tract Infections in the Riyadh Region.

The successful treatment of bacterial disease is relied on selecting a suitable drug based on the type of bacteria and antimicrobial susceptibility testing. The study's objective was to identify bacterial isolates from urine samples of patients from the community, followed by antimicrobial susceptibility testing of the isolated bacteria. A total of seventy urine samples were received in the clinical microbiology laboratory; out of which 18 culture-positive cultures and by direct identification using MALDI-TOF MS (Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry) were identified. Of 18 identified bacteria, 17 (94%) were pathogenic. The culture demonstrated that the major species detected in urine samples were Escherichia coli, Klebsiella pneumoniae, Enterococcus faecalis, and Aeromonas caviae. E. coli (72.2%) was the most common bacterium retrieved from urine samples followed by K. pneumoniae (16.6%). Interestingly, all the isolates, except Enterococcus faecalis, were resistant to erythromycin. The isolates 8 of 13 (61.5%) were resistant to both of the cotrimoxazole and tetracycline. We performed MLST (Multi-locus Sequence Typing) typing of 13 E. coli isolates to study their genetic relatedness and diversity. MLST typing of E. coli showed a total of nine different STs (Sequence Types), which showed the diversity among them. ST 129 was the most common ST found in three E. coli isolates. In our study, two isolates with ST 1126 and ST 1432 represented the global clonal complex 155. MALDI-TOF MS provided dependable results for identifying the bacteria up to species level from urine samples by indirect culture methods. Such local surveillances are highly recommended for empirical therapy awareness and determining isolates' level of resistance.

Differential Gene Expression of Efflux Pumps and Porins in Clinical Isolates of MDR Acinetobacter baumannii.

Background: Acinetobacter baumannii is an opportunistic pathogen associated with healthcare infections and high mortality rates in intensive care units all over the globe. Porins and efflux pumps over-expression have been reported as contributing factors in escalating drug resistance and rendering treatment ineffective. In this study, we investigated the mechanisms of multidrug resistance (MDR) in A. baumannii clinical isolates. Methods: A total of 30 A. baumannii isolates were included in the present study from Nehru Hospital (PGIMER-Chandigarh) located in North India. Kirby Bauer disk diffusion assay and MIC were performed to determine the antimicrobial susceptibility pattern. Screening of beta-lactamases was performed using PCR. Relative gene expression of four RND, one MATE efflux pump, and two outer membrane proteins were determined using RT-PCR. Molecular typing of 22 isolates was carried out using MLST Oxford scheme. Results: CarO porin genes showed over-expression in 63% isolates followed by adeGandabeM efflux pump downregulation/underexpression (<0.5 fold), suggesting the carbapenem-susceptible phenotypic nature of the isolates. High prevalence of VIM-2, NDM-1, and OXA-23 genes was observed in A. baumannii isolates. Interestingly, NDM-1 and OXA-58 were traced in 10 and3 A. baumannii isolates respectively; 13 of 22 (59%) isolates showed novel Sequence Types (STs) in the Multi-Locus Sequence Typing (MLST) analysis. ST 1087 was most commonly found ST among all others (16 STs). Conclusions: This study indicated a possible role of carO porin genes and adeG (RND) andabeM (MATE) efflux pumps in carbapenem susceptibility of A. baumannii. New STs were also reported in the majority of the isolates.

Virulence Potential and Treatment Options of Multidrug-Resistant (MDR) Acinetobacter baumannii.

Acinetobacter baumannii is an opportunistic pathogen which is undoubtedly known for a high rate of morbidity and mortality in hospital-acquired infections. A. baumannii causes life-threatening infections, including; ventilator-associated pneumonia (VAP), meningitis, bacteremia, and wound and urinary tract infections (UTI). In 2017, the World Health Organization listed A. baumannii as a priority-1 pathogen. The prevalence of A. baumannii infections and outbreaks emphasizes the direct need for the use of effective therapeutic agents for treating such infections. Available antimicrobials, such as; carbapenems, tigecycline, and colistins have insufficient effectiveness due to the appearance of multidrug-resistant strains, accentuating the need for alternative and novel therapeutic remedies. To understand and overcome this menace, the knowledge of recent discoveries on the virulence factors of A. baumannii is needed. Herein, we summarized the role of various virulence factors, including; outer membrane proteins, efflux pumps, biofilm, penicillin-binding proteins, and siderophores/iron acquisition systems. We reviewed the recent scientific literature on different A. baumannii virulence factors and the effective antimicrobial agents for the treatment and management of bacterial infections.

Antibiofilm Activities of Biogenic Silver Nanoparticles Against Candida albicans.

Biofilms are microbial colonies that are encased in an organic polymeric matrix and are resistant to antimicrobial treatments. Biofilms can adhere to both biotic and abiotic surfaces, allowing them to colonize medical equipment such as urinary and intravenous catheters, mechanical heart valves, endotracheal tubes, and prosthetic joints. Candida albicans biofilm is the major etiological cause of the pathogenesis of candidiasis in which its unobstructed growth occurs in the oral cavity; trachea, and catheters that progress to systemic infections in the worst scenarios. There is an urgent need to discover novel biofilm preventive and curative agents. In the present investigation, an effort is made to observe the role of cyanobacteria-derived AgNPs as a new antibiofilm agent with special reference to candidiasis. AgNPs synthesized through the green route using Anabaena variabilis cell extract were characterized by UV-visible spectroscopy. The nanoparticles were spherical in shape with 11-15 nm size and were monodispersed. The minimum inhibitory concentration (MIC) of AgNPs was obtained at 12.5 µg/mL against C. albicans. AgNPs 25 µg/mL showed 79% fungal cell membrane permeability and 22.2% ROS production. AgNPs (25 µg/mL) also facilitated 62.5% of biofilm inhibition and degradation. Therefore, AgNPs could be considered as a promising antifungal agent to control biofilm produced by C. albicans.

Unravelling the interaction of glipizide with human serum albumin using various spectroscopic techniques and molecular dynamics studies.

Glipizide is known to stimulate insulin secretion by ß-cells of the pancreas. It is a second-generation sulfonylurea drug used in the management of type 2 diabetes. The shorter biological half-life makes it a suitable candidate to be designed as a controlled release formulation. Human serum albumin (HSA), a major plasma protein plays a crucial role in the transportation of drugs, hormones, fatty acids, and many other molecules and determines their physiological fate and biodistribution. In this study, the interaction of glipizide with HSA was investigated under physiological conditions using multi-spectroscopic techniques corroborated with molecular docking and dynamics approach. It was found that glipizide integrates to HSA with a binding constant in the order of 105 M-1. The mode of fluorescence quenching by glipizide is static in nature with one binding site. Glipizide preferentially interacts with sub-domain IIA of HSA and their complexion is thermodynamically favorable. This interaction results in the loss of α-helical content of HSA. The energy transfer efficiency from HSA to glipizide was found to be 26.72%. In silico molecular docking and simulation studies ratified in vitro findings and revealed that hydrogen bonds and hydrophobic interactions are accountable for glipizide-HSA complex formation.Communicated by Ramaswamy H. Sarma.

Comparative Analysis of Bacteriophytochrome Agp2 and Its Engineered Photoactivatable NIR Fluorescent Proteins PAiRFP1 and PAiRFP2.

Two photoactivatable near infrared fluorescent proteins (NIR FPs) named "PAiRFP1" and "PAiRFP2" are formed by directed molecular evolution from Agp2, a bathy bacteriophytochrome of Agrobacterium tumefaciens C58. There are 15 and 24 amino acid substitutions in the structure of PAiRFP1 and PAiRFP2, respectively. A comprehensive molecular exploration of these bacteriophytochrome photoreceptors (BphPs) are required to understand the structure dynamics. In this study, the NIR fluorescence emission spectra for PAiRFP1 were recorded upon repeated excitation and the fluorescence intensity of PAiRFP1 tends to increase as the irradiation time was prolonged. We also predicted that mutations Q168L, V244F, and A480V in Agp2 will enhance the molecular stability and flexibility. During molecular dynamics (MD) simulations, the average root mean square deviations of Agp2, PAiRFP1, and PAiRFP2 were found to be 0.40, 0.49, and 0.48 nm, respectively. The structure of PAiRFP1 and PAiRFP2 were more deviated than Agp2 from its native conformation and the hydrophobic regions that were buried in PAiRFP1 and PAiRFP2 core exposed to solvent molecules. The eigenvalues and the trace of covariance matrix were found to be high for PAiRFP1 (597.90 nm2) and PAiRFP2 (726.74 nm2) when compared with Agp2 (535.79 nm2). It was also found that PAiRFP1 has more sharp Gibbs free energy global minima than Agp2 and PAiRFP2. This comparative analysis will help to gain deeper understanding on the structural changes during the evolution of photoactivatable NIR FPs. Further work can be carried out by combining PCR-based directed mutagenesis and spectroscopic methods to provide strategies for the rational designing of these PAiRFPs.

Identifying novel sphingosine kinase 1 inhibitors as therapeutics against breast cancer.

Sphingosine kinase 1 (SphK1) is a promising therapeutic target against several diseases including mammary cancer. The aim of present work is to identify a potent lead compound against breast cancer using ligand-based virtual screening, molecular docking, MD simulations, and the MMPBSA calculations. The LBVS in molecular and virtual libraries yielded 20,800 hits, which were reduced to 621 by several parameters of drug-likeness, lead-likeness, and PAINS. Furthermore, 55 compounds were selected by ADMET descriptors carried forward for molecular interaction studies with SphK1. The binding energy (ΔG) of three screened compounds namely ZINC06823429 (-11.36 kcal/mol), ZINC95421501 (-11.29 kcal/mol), and ZINC95421070 (-11.26 kcal/mol) exhibited stronger than standard drug PF-543 (-9.9 kcal/mol). Finally, it was observed that the ZINC06823429 binds tightly to catalytic site of SphK1 and remain stable during MD simulations. This study provides a significant understanding of SphK1 inhibitors that can be used in the development of potential therapeutics against breast cancer.

Impact of p53 arg72pro SNP on Breast Cancer Risk in North Indian Population.

BACKGROUND: Genetic changes in p53 gene contribute to breast cancer susceptibility. OBJECTIVE AND METHODS: A case-control study and a meta-analysis were performed to investigate the role of p53 codon72 SNP with breast cancer susceptibility in Indian women. RESULTS: p53 heterozygous arginine variant was associated with decreased risk of breast cancer in total cohort. In meta-analysis, Allelic and GG vs. CC genetic comparison model were found to be associated with breast cancer risk. Moreover, recessive comparison model indicated a protective correlation with breast cancer occurrence. CONCLUSION: The findings of our case-control study and meta-analysis suggest a significant association between p53 Arg72Pro polymorphism and an increased risk of breast cancer in Indian population.

Study on the interaction of antidiabetic drug Pioglitazone with calf thymus DNA using spectroscopic techniques.

Pioglitazone is an antidiabetic drug used to treat type 2 diabetes mellitus. Interaction of Pioglitazone with calf thymus DNA was investigated using multispectroscopic techniques and molecular docking study. Quenching and binding constant was calculated at 3 different temperatures. The binding constant of Pioglitazone with calf thymus DNA was calculated to be 6.49 × 103 M-1 at 293 K. The quenching mechanism was found to be a static process, and thermodynamic parameters revealed van der Waals interactions and hydrogen bonds to be the major force working in Pioglitazone-DNA interaction. Pioglitazone follows the nonintercalative mode of binding and was involved in complex formation with DNA through minor groove binding and electrostatic interactions. Experiments like KI quenching studies, dye displacement assays, Circular Dichroism (CD) spectroscopy, DNA melting study, and viscosity measurements studies supported the nonintercalative mode of binding. This was further corroborated by molecular docking studies.

Structure prediction and functional analyses of a thermostable lipase obtained from Shewanella putrefaciens.

Previous experimental studies on thermostable lipase from Shewanella putrefaciens suggested the maximum activity at higher temperatures, but with little information on its conformational profile. In this study, the three-dimensional structure of lipase was predicted and a 60 ns molecular dynamics (MD) simulation was carried out at temperatures ranging from 300 to 400 K to better understand its thermostable nature at the molecular level. MD simulations were performed in order to predict the optimal activity of thermostable lipase. The results suggested strong conformational temperature dependence. The thermostable lipase maintained its bio-active conformation at 350 K during the 60 ns MD simulations.

Interaction of procarbazine with calf thymus DNA-a biophysical and molecular docking study.

Interaction of procarbazine (PCZ) with calf thymus DNA was studied using biophysical and molecular docking studies. Procarbazine was to interact with DNA with a binding constant of 6.52 × 103  M-1 as calculated using ultraviolet-visible spectroscopy. To find out the binding mode, molecular docking was performed that predicted PCZ to interact with DNA through groove binding mode with binding affinity of -6.7 kcal/mole. To confirm the groove binding nature, different experiments were performed. Dye displacement assays confirmed the non-intercalative binding mode. Procarbazine displaced Hoechst dye from the minor groove of DNA while it was unable to displace intercalating dyes. There was no increase in the viscosity of DNA solution in presence of PCZ. Also, negligible change in the secondary structure of DNA was observed in presence of PCZ as evident by circular dichroism spectra. Procarbazine caused decrease in the melting temperature of DNA possibly because of decrease in the stability of DNA caused by groove binding interaction of PCZ with DNA.

SCREENING OF PHOTOSYNTHETIC O2 -EVOLVING PROKARYOTES FOR AN INSULIN-LIKE ANTIGEN(1).

Diabetes mellitus (DM), a metabolic disorder, is becoming a major health problem worldwide. Insulin is the single hope for management of type 1 diabetes, but it is not always available or suitable. For finding additional bioresources, the present study was performed. ELISA-based preliminary screening of cyanobacterial biomass using antihuman insulin antibody have detected an insulin-like antigen in Spirulina platensis S-5, Spirulina NCCU-482, and Spirulina NCCU-483. Their similarity with insulin-like antigen was further confirmed by electrophoretic mobility using bovine insulin as marker.