Naveen Chaudhary

Dept of medical microbiology

PGIMER,CHANDIAGRH

Multidrug resistance bacteria:-challenging to

modern medicine

Bacteria can be inactivated through various physical

and chemical means

Heat

Radiations

Phenol ,alcohol etc

Introduction Plasma :- one of the four fundamental states of

matter, the others being solid, liquid, and gas.

Atmospheric pressure non-thermal plasma (APP):- Efficiently and effectively control multidrug resistant microorganisms

APP :-Efficient source of a combination of electronically excited atoms, charged particles (electrons, ions),ozone (O3), UV photons and radicals

Two types of plasma systems were investigated:

Nano-second pulsed plasma (NPP) as liquid discharge plasma

Argon gas-feeding dielectric barrier discharge (Ar-DBD) as a form of surface plasma

Reactive oxygen species (ROS) :-are chemically reactive molecules containing oxygen

Peroxides

Superoxide

hydroxyl radical

singlet oxygen

Strains used S. aureus (wild type)

Penicillin-resistant S. aureus (PRSA)

Methicillin-resistant S. aureus (MRSA)

Gentamicin-resistant S. aureus (GRSA)

Nano-second pulsed plasma

Shockwave measurement

Argon gas-feeding dielectric barrier discharge

Measurement of the ROS: Bacterial samples in saline were exposed to the 4th discharge of NPP and to 5 min of DBD plasma

All exposed bacterial samples were washed with PBS, and 500 ml of 10 Mm H2DCFDA were added

The mean fluorescence intensity was determined at the corresponding excitation and emission wavelengths

Molecular dynamics (MD) simulations used to study the interaction of the reactive ( O, OH, O3 and H2O2) as well as non-reactive ( O2, H2O) plasma species with PG of S. aureus.

X-ray photoelectron spectroscopy

XPS used to investigate the variation in the functional groups (C-H/C-C, C-OH and C=O) of peptidoglycan (PG)

Morphology analysis by SEM

Quantitative real time PCR:-A quantitative real time polymerase chain reaction (Q-PCR) used for analysis of the level of mecA, mecI,

mecRI and femA gene in the MRSA as well as wild strains was performed after exposure to NPP and DBD plasma

Analysis of change in Temperature and PH

Inactivation of bacterial strains

Colonies and SEM analysis

Gene expression analyses after NPP and Ar-DBD treatment

Composition of bacteria with plasma treatment obtained using XPS

Average fraction of important bonds of the PG after

impact of various plasma species.

Examined the protein oxidation to understand the structural modification for both proteins after NPP treatment

Observed the oxidation in calf thymus DNA after the NPP treatment

Discussion β-lactamase activity and SEM analysis:-If compare

the β –lactamase activity, we can observe that both NPP and DBD resulted in a decrease in activity, which may be due to the inactivation of the β -lactamase enzyme as well as its related gene.

SEM images show that the NPP treatment crushed many bacterial spores due to shock waves while only the DBD treatment changed the morphology

pH role in bacterial inactivationThey observed that the pH in saline solution became

~4.2 after 5 min of DBD treatment and ~5.3 after 4th discharge of NPP treatment.

Found no significant antimicrobial effects for both bacteria lines in an acidic environment, which is also supported by previous research.

Role of combined effect of the RS, pH and shock

waves on the inactivation of bacteria:-The role of the

pH is only to increase the reactivity of the radicals, and

the pH has no direct effect on the bacterial inactivation in

this study.

The inactivation of S. aureus with both plasma Sources

occurs with almost the same radicals, although the

amount can vary from plasma source to plasma source.

The shock waves play an important role in the

inactivation, which creates a difference in the efficiency of

NPP and Ar-DBD treatments

CONCLUSION XPS and MD simulation results show that PG is

oxidized due to plasma action that may help in inactivation of S. aureus. Therefore, research on plasma-liquid interactions is a recommended field for further new advances in plasma medicine like wound healing, dermatology and dentistry