title: Intracellular, biofilm-inhibitory and membrane-damaging activities of nimbolide isolated from Azadirachta indica A. Juss (Meliaceae) against meticillin-resistant Staphylococcus aureus.
abstract: Staphylococcus aureus is a leading aetiologic agent of nosocomial- and community-acquired infectious diseases worldwide. The public health concern regarding staphylococcal infections is inflated by the increasing occurrence of multidrug-resistant strains, e.g. multidrug- and meticillin-resistant S.aureus (MDR MRSA). This study was designed to evaluate the intracellular killing, membrane-damaging and biofilm-inhibitory activities of nimbolide isolated from Azadirachta indica against MDR MRSA. In vitro antibacterial activity of nimbolide was determined by performing MIC, minimal bactericidal concentration (MBC) and time-kill kinetic studies. Bacterial membrane-damaging activity was determined by membrane perturbation and scanning electron microscopy (SEM) examination. Biofilm-inhibitory activities were determined by SEM. Cellular drug accumulation and assessments of intracellular activities were performed using Vero cell culture. SEM revealed that nimbolide caused significant membrane damage and lysis of the S. aureus cells. The biofilm structure was disrupted, and the biofilm formation was greatly reduced in the presence of nimbolide as examined by SEM. The level of accumulation of nimbolide in Vero cells incubated for 24 h is relatively higher than that of ciprofloxacin and nalidixic acid (Cc/Ce for nimbolide > ciprofloxacin and nalidixic acid). The viable number of intracellular S. aureus was decreased [reduction of ~2 log10 c.f.u. (mg Vero cell protein)-1] in a time-dependent manner in the presence of nimbolide (4× MBC) that was comparable to that of tetracycline and nalidixic acid. The significant intracellular, biofilm-inhibitory and bacterial membrane-damaging activities of nimbolide demonstrated here suggested that it has potential as an effective antibacterial agent for the treatment of severe infections caused by MDR MRSA.
authors.affiliation: Department of Biochemistry, University of Calcutta, 35 Ballygunge, Circular Road, Kolkata 700019, India.
authors.forename: Prodipta
authors.initials: P
authors.lastname: Sarkar
authors.affiliation: Department of Biochemistry, University of Calcutta, 35 Ballygunge, Circular Road, Kolkata 700019, India.
authors.forename: Saurabh
authors.initials: S
authors.lastname: Acharyya
authors.affiliation: Department of Biochemistry, University of Calcutta, 35 Ballygunge, Circular Road, Kolkata 700019, India.
authors.forename: Anirban
authors.initials: A
authors.lastname: Banerjee
authors.affiliation: Department of Chemistry, University of Calcutta, 92 Acharya Prafulla Chandra Road, Kolkata 700009, India.
authors.forename: Amarendra
authors.initials: A
authors.lastname: Patra
authors.affiliation: Department of Biochemistry, University of Calcutta, 35 Ballygunge, Circular Road, Kolkata 700019, India.
authors.forename: Karthika
authors.initials: K
authors.lastname: Thankamani
authors.affiliation: National Institute of Cholera and Enteric Diseases, P-33 C.I.T. Road, Scheme-XM, Beliaghata, Kolkata 700010, India.
authors.forename: Hemanta
authors.initials: H
authors.lastname: Koley
authors.affiliation: Department of Biochemistry, University of Calcutta, 35 Ballygunge, Circular Road, Kolkata 700019, India.
authors.forename: Prasanta K
authors.initials: PK
authors.lastname: Bag
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D031601:Azadirachta / Q000737:chemistry*; D018441:Biofilms / Q000187:drug effects*; D002462:Cell Membrane / Q000187:drug effects*; D002522:Chlorocebus aethiops; D006801:Humans; D036701:Limonins / Q000494:pharmacology*; D055624:Methicillin-Resistant Staphylococcus aureus / Q000187:drug effects* / Q000502:physiology; D010936:Plant Extracts / Q000494:pharmacology*; D013203:Staphylococcal Infections / Q000382:microbiology*; D014709:Vero Cells
chemical_list: D000900:Anti-Bacterial Agents; D036701:Limonins; D010936:Plant Extracts; C042198:nimbolide