Haris M. Khan

Beta-lactams and Beta-lactamase-inhibitors in current- or potential-clinical practice: A comprehensive update

The use of successive generations of β-lactams has selected successive generations of β-lactamases including CTX-M ESBLs, AmpC β-lactamases, and KPC carbapenamases in Enterobacteriaceae. Moreover, this cephalosporin resistance, along with rising resistance to fluoroquinolones, is now driving the use of carbapenems and unfortunately the carbapenem resistance has emerged markedly, especially in Acinetobacter spp. due to OXA- and metallo-carbapenemases. The industry responded to the challenge of rising resistance and recently developed some novel β-lactams such as ceftobiprole, ceftaroline etc. and many β-lactam compounds, including β-lactamase-inhibitors, such as BMS-247243, S-3578, RWJ-54428, CS-023, SMP-601, NXL 104, BAL 30376, LK 157, and so on are under trials. This review provides the comprehensive accounts of the developments in penicillins, cephalosporins, carbapenems, and β-lactamase-inhibitors, and the insight about medicinal chemistry, mechanism(s) of action and resistance, potential strategies to overcome resistance due to β-lactamases, and also the recent advancements in the development of newer β-lactam compounds; some of which are still under trials and yet to be classified. This review will fill the gap since previously published reviews and will serve as a comprehensive update on the current topic.

Anti-biofilm efficacy of silver nanoparticles against MRSA and MRSE isolated from wounds in a tertiary care hospital.

Purpose: Different approaches have been used for preventing biofilm-related infections in health care settings. Many of these methods have their own de-merits, which include chemical-based complications; emergent antibiotic resistant strains, etc. The formation of biofilm is the hallmark characteristic of Staphylococcus aureus and S. epidermidis infection, which consists of multiple layers of bacteria encased within an exopolysachharide glycocalyx. Nanotechnology may provide the answer to penetrate such biofilms and reduce biofilm formation. Therefore, the aim of present study was to demonstrate the biofilm formation by methicillin resistance S. aureus (MRSA) and methicillin resistance S. epidermidis (MRSE) isolated from wounds by direct visualisation applying tissue culture plate, tube and Congo Red Agar methods. Materials and Methods: The anti-biofilm activity of AgNPs was investigated by Congo Red, scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) techniques. Results: The minimum inhibitory concentration (MIC) was found to be in the range of 11.25-45 μg/ml. The AgNPs coated surfaces effectively restricted biofilm formation of the tested bacteria. Double fluorescent staining (propidium iodide staining to detect bacterial cells and fluorescein isothiocyanate concanavalin A (Con A-FITC) staining to detect the exopolysachharides matrix) technique using CLSM provides the visual evidence that AgNPs arrested the bacterial growth and prevent the glycocalyx formation. In our study, we could demonstrate the complete anti-biofilm activity AgNPs at a concentration as low as 50 μg/ml. Conclusions: Our findings suggested that AgNPs can be exploited towards the development of potential anti-bacterial coatings for various biomedical and environmental applications. In the near future, the AgNPs may play major role in the coating of medical devices and treatment of infections caused due to highly antibiotic resistant biofilm.

Interaction of Al2O3 nanoparticles with Escherichia coli and their cell envelope biomolecules

The aim of this study is to investigate the antibacterial activity of aluminium oxide nanoparticles (Al2O3 NPs) against multidrug‐resistant clinical isolates of Escherichia coli and their interaction with cell envelope biomolecules.

Interaction of silver nanoparticles with Escherichia coli and their cell envelope biomolecules

The antibacterial effect of AgNPs was investigated by determining MIC/MBC and growth kinetics assay. The lowest MIC/MBC was found to be in the range of 11.25–22.5 µg ml−1. The growth kinetics curve shows that 25 µg ml−1 AgNPs strongly inhibits the bacterial growth. Confocal laser scanning electron microscopy (CLSM) shows that as the concentration of NPs increases, reduction in the number of cells was observed and at 50 µg ml−1 of NPs, 100% death was noticed. Scanning electron microscopy (SEM) shows cells were severely damaged with pits, multiple depressions, and indentation on cell surface and original rod shape has swollen into bigger size. High resolution‐transmission electron microscopic (HR‐TEM) micrograph shows that cells were severely ruptured. The damaged cells showed either localized or complete separation of the cell membrane. The NPs that anchor onto cell surface and penetrating the cells may cause membrane damage, which could result in cell lysis. The interaction of AgNPs to membrane biomolecules; lipopolysaccharide (LPS) and L‐α‐phosphatidyl‐ethanolamine (PE) were investigated by attenuated total reflectance–fourier transform infrared (ATR–FTIR) spectroscopy. LPS and PE showed IR spectral changes after AgNPs exposure. The O‐antigen part of LPS was responsible for interaction of NPs through hydrogen bonding. The phosphodiester bond of PE was broken by AgNPs, forming phosphate monoesters and resulting in the highly disordered alkyl chain. The AgNPs‐induced structural changes in phospholipid may lead to the loss of amphiphilic properties, destruction of the membrane and cell leaking. The biomolecular changes in bacterial cell envelope revealed by ATR–FTIR provide a deeper understanding of cytotoxicity of AgNPs.

Green synthesis of silver nanoparticles and characterization of their inhibitory effects on AGEs formation using biophysical techniques.

Advanced glycation end-products (AGEs) resulting from non-enzymatic glycation are one of the major factors implicated in secondary complications of diabetes. Scientists are focusing on discovering new compounds that may be used as potential AGEs inhibitors without affecting the normal structure and function of biomolecules. A number of natural and synthetic compounds have been proposed as AGE inhibitors. In this study, we investigated the inhibitory effects of AgNPs (silver nanoparticles) in AGEs formation. AgNPs (~30.5 nm) synthesized from Aloe Vera leaf extract were characterized using UV-Vis spectroscopy, energy-dispersive X-ray spectroscopy (EDX), high resolution-transmission electron microscopy, X-ray diffraction and dynamic light scattering (DLS) techniques. The inhibitory effects of AgNPs on AGEs formation were evaluated by investigating the degree of reactivity of free amino groups (lysine and arginine residues), protein-bound carbonyl and carboxymethyl lysine (CML) content, and the effects on protein structure using various physicochemical techniques. The results showed that AgNPs significantly inhibit AGEs formation in a concentration dependent manner and that AgNPs have a positive effect on protein structure. These findings strongly suggest that AgNPs may play a therapeutic role in diabetes-related complications.

Gum arabic capped-silver nanoparticles inhibit biofilm formation by multi-drug resistant strains of Pseudomonas aeruginosa

Clinical isolates (n = 55) of Pseudomonas aeruginosa were screened for the extended spectrum β‐lactamases and metallo‐β‐lactamases activities and biofilm forming capability. The aim of the study was to demonstrate the antibiofilm efficacy of gum arabic capped‐silver nanoparticles (GA‐AgNPs) against the multi‐drug resistant (MDR) biofilm forming P. aeruginosa. The GA‐AgNPs were characterized by UV‐spectroscopy, X‐ray diffraction, and high resolution‐transmission electron microscopy analysis. The isolates were screened for their biofilm forming ability, using the Congo red agar, tube method and tissue culture plate assays. The biofilm forming ability was further validated and its inhibition by GA‐AgNPs was demonstrated by performing the scanning electron microscopy (SEM) and confocal laser scanning microscopy. SEM analysis of GA‐AgNPs treated bacteria revealed severely deformed and damaged cells. Double fluorescent staining with propidium iodide and concanavalin A‐fluorescein isothiocyanate concurrently detected the bacterial cells and exopolysaccharides (EPS) matrix. The CLSM results exhibited the GA‐AgNPs concentration dependent inhibition of bacterial growth and EPS matrix of the biofilm colonizers on the surface of plastic catheters. Treatment of catheters with GA‐AgNPs at 50 µg ml−1 has resulted in 95% inhibition of bacterial colonization. This study elucidated the significance of GA‐AgNPs, as the next generation antimicrobials, in protection against the biofilm mediated infections caused by MDR P. aeruginosa. It is suggested that application of GA‐AgNPs, as a surface coating material for dispensing antibacterial attributes to surgical implants and implements, could be a viable approach for controlling MDR pathogens after adequate validations in clinical settings.

blaCTX-M, blaTEM, and blaSHV in Enterobacteriaceae from North-Indian tertiary hospital: high occurrence of combination genes

OBJECTIVE To delineate the frequency of occurrence of bla(CTX-M), bla(TEM), and bla(SHV) in Enterobacteriaceae from North-Indian tertiary hospital. METHODS A random collection of a subset of 45 Escherichia coli (E. coli) and 28 Klebsiella pneumoniae (K. pneumoniae) that was resistant to a third generation cephalosporin and obtained during 2007-2008 was selected for detailed screening for bla(CTX-M), bla(TEM), and bla(SHV) by monoplex PCRs. The isolates demonstrating the presence of bla(CTX-M) alleles were characterized for the specific CTX-M-genogroup by using a multiplex PCR. RESULTS Resistance to cefoperazone, ceftazidime, ceftriaxone, cefotaxime, cefoxitin and piperacillin was 100% each in K. pneumoniae isolates, whereas these resistance-rates for E. coli isolates were 93.1%, 83.8%, 91.9%, 93.6%, 97.3% and 97.1%, respectively. Concomitant resistance to aminoglycosides, quinolones and aztreonam was also noticed. Presence of any of the bla genes (bla(CTX-M), bla(TEM), and bla(SHV)) was noticed in a total of 28 (38.4%) isolates of the 73 isolates studied. Many isolates demonstrated occurrence of these genes in various combinations. bla(CTX-M), bla(TEM), and bla(SHV) were noticed in 28.8%, 10.9% and 13.7% isolates, respectively. Multiplex PCR in bla(CTX-M) harboring isolates demonstrated the presence of CTX-M-Genogroup-1 and sequencing for the specific CTX-M-type revealed presence of CTX-M-15 type. RAPD typing showed wide diversity in isolates. CONCLUSIONS This is amongst the premier report describing the simultaneous occurrence of bla(TEM), bla(SHV), and bla(ampC) in Indian Enterobacteriaceae and that wider dissemination of these genes, as demonstrated by diversity of isolates, raises concern and emphasizes a need for extensive search for the presence of these gene pools in Indian subcontinent.

Synthesis and characterization of Schiff base octaazamacrocyclic complexes and their biological studies

A condensation reaction between 1,2-diphenylethane-1,2-dione dihydrazone (DPEDDH) and dimethyl or diethyloxalate in methanol resulted in a novel Schiff base octaazamacrocyclic ligand, (L): (6,7,14,15-tetraoxa-2,3,10,11-tetraphenyl-1,4,5,8,9,12,13,16-octaazacyclohexadecane-1,3,9,11-tetraene). Subsequently metal complexes of the type [MLX2] and [CuL]X2; (M=Mn(II), Co(II), Ni(II) and Zn(II); X=Cl or NO3) were synthesized by the reaction of the free macrocyclic ligand (L) with the corresponding metal salts in 1:1 molar ratio. These complexes were characterized on the basis of analytical data, molar conductivity and magnetic susceptibility measurements, ESI-mass, IR, NMR ((1)H and (13)C), EPR and electronic spectral studies. The thermal stability of the complexes was also studied by TGA and DTA analyses. These studies show that all the complexes have octahedral arrangement around the metal ions except copper complexes which are square planar. The ligand and its complexes were screened for their antibacterial activity in vitro against Gram-positive (Staphylococcus aureus) and Gram-negative (Escherichia coli) bacteria and were also studied for their anticancer activity against the human cancer cells lines: HeLa (Human cervical carcinoma), MCF7 (Human breast adenocarcinoma) and Hep3B (Human Hepatocellular carcinoma). The recorded IC50 values for the tested compounds show moderate to good cytotoxicity against these cancer cell lines. The copper complex, [CuL]Cl2, showed excellent antimicrobial activity against tested microorganisms which is almost equivalent to the standard drug ciprofloxacin.

Protective effect of L. donovani antigens using glucan as an adjuvant.

Golden hamsters were immunized with various antigen fractions of Leishmania donovani promastigotes. Beta 1,3-glucan was used as an adjuvant in these vaccination experiments. The results indicate that immunization of animals with the microsomal fraction (subcellular fraction III) in combination with glucan confers considerable immune protection against L. donovani infection. The immune protection was confirmed by correspondingly lower parasite burden in the livers and spleens of test animals compared to controls. Additionally, the vaccinated animals showed positive skin test responsiveness after challenge, along with increased antibody titres. Immunization of animals with whole and particulate antigen fractions was also found to afford a high degree of resistance. The other subcellular and soluble antigen fractions conferred very little protection. In these experiments, glucan was found to be a potent adjuvant when injected, intraperitoneally, with Leishmania antigens. Similar doses of parasite extracts given without an adjuvant were able to confer only very little or no protection.

Increasing antimicrobial resistance among uropathogens: Is fosfomycin the answer?

Introduction: Urinary tract infection (UTI) is one of the most common infectious diseases in clinical practice. The choice of antibiotics for the treatment of UTI is limited by the rising rates of antibiotic resistance. There is an urgent need to discover new effective treatment solutions. Fosfomycin may be an interesting alternative to the currently used treatments of UTIs. Materials and Methods: The study was conducted over 6 months period (January to June 2013) in Department of Microbiology, JNMCH, AMU, Aligarh. A total of 1840 urine samples were submitted. Culture and sensitivity was done as per standard microbiological procedures. Methicillin-resistant Staphylococcus aureus (MRSA), high-level aminoglycoside resistance (HLAR), extended spectrum beta-lactamases (ESBL), AmpC and metallo-beta-lactamases (MBL) production was detected. Results: Culture was positive in 504 (27.4%) cases. Gram-negative etiology was identified in 390 (73%) cases. ESBL production was detected in 154 (37.1%) while 82 (21.6%) were Amp C. No, MBL was detected. Among Gram-positive bacteria, 68 (51.5%) were MRSA, while 4 (13.3%) were vancomycin resistant enterococci (VRE). HLAR was seen in 53.3% of enterococci. Fosfomycin was effective in 100% of MRSA, VRE, ESBL, HLAR, and overall, susceptibility to fosfomycin in AmpC producers was extremely high (99%). Norfloxacin and cotrimoxazole were not proved effective as only three isolates were sensitive to norfloxacin, while all Gram-negative isolates were resistant to cotrimoxazole. Pseudomonas species showed 65% and 75% susceptibility to colistin and polymixin B, respectively. Conclusion: Fosfomycin has emerged as a promising option, especially in cases involving multi-drug-resistant pathogens in which previous antibiotics have failed to cure the infection.