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RESEARCH ETHICS

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Arfi Khan

on 9 July 2015

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Transcript of RESEARCH ETHICS

Evaluation of Synergistic Antibacterial Effects of Antibiotic Loaded Chitosan Nanoparticles on Methicillin-Resistant
Staphylococcus aureus
and
Escherichia coli

Supervised By: Dr. M. Arfat Yameen
Presented By: Sana Mushtaq
CIIT/FA13-R00-032/ATD

S. aureus

Attachment host cell Colonization
Infection
Pathogenecity is based on:
Exchange of Genetic Information
Virulence Factors
Cell Surface
Extracellular enzymes
Toxins
S. aureus
and MRSA are resistant to Beta-Lactam, Macrolides, Aminoglycosides, & quinolones.

Emerging vancomycin resistant
S. aureus
E. coli
Community associated & Nosocomial infections
Major cause of epidemic infant diarrhea is O:H serotypes of
E. coli
ESBL
E. coli
an emerging threat to patients
GIT commensal- found in every healthy individual
In US, 73480 patients are infected with
E. coli
per year

Cont....
Combating resistance
Designing new drug regimen

Adopting alternative drug delivery systems

New drug moiety

Nano-medicine
Nanotechnology/Nanomedicine
Aims & Objectives
To evaluate the enhanced antibacterial activity of antibiotic loaded chitosan nanoparticles against resistant strains of G +ive and G -ive bacteria
Isolation & identification of resistant strains of MRSA &
E. coli
Evaluation of antimicrobial susceptibility
Synthesis & characterization of blank & drug loaded nanoparticles
Evaluation of synergistic antibacterial acitivity
Biochemical & Molecular identification
Growth on selective/differential media
Gram +ive
Mannitol salt agar-
Sheep blood agar-
Gram -ive
MacConkey agar
Gram staining
Cont....
Cont....
DNA extraction & PCR
MRSA
nuc
A (270 bp) &
mec
A (533 bp) gene detection



E. coli
16S rRNA (1400 bp) &
mec
A (884 bp) gene detection
Antimicrobial susceptibility testing
Disc diffusion test
Kirby Bauer Method
Methods of CNPs preparation
Ionic gelation method
Chemical used
Chitosan- low molecular weight deacetylated chitin poly D- glucosamine (Sigma Aldrich)
Sodium tripolyphosphate 85% Techical grade (Sigma Aldrich)
Ceftriaxone sodium- 83.2% potency (Cirin phm.)
Cont....

SEM





TEM






Irregular and spherical nanoparticles




XRD
Antimicrobial susceptibility testing with blank and Ceftriaxone loaded CNPs
Antimicrobial susceptibility testing
Background
Evolution of mankind lead towards evaluation of diseases

S. aureus
- The Gram +ive
MRSA


E. coli
- The Gram -ive
ESBL
E. coli
S. aureus
causes blood, soft tissue, and skin infections.
MRSA causes nosocomial infections
One third of individuals are
S. aureus
carriers
80,000 patients per year are infected with MRSA

Cont ....
Three major classes
Intestinal strains
Commensal strains
Extra intestinal pathogenic strains
Colonization on host mucosa Evade host defense system Multiplication
Infection
Pathogenecity is based on
Precipitating factors
Toxins
Cont....
E. coli
& ESBL
E. coli
are resistant against beta lactams & qiunolones.

Emergence of rapid resistance against third and fourth generation cephalosporins & flouroquinolones .
(Collignon, 2009)
(Aries de Sousa & de Lencastre, 2004)
(Gordon & Lowy, 2008)
(Novick, 2000; Decker, 2008; Gordon & Lowy, 2008; Huang & Platt, 2003; Cookson et. al., 2007)
(Levine & Edelman, 1984; States et. al., 2005; Allocati et. al., 2013)
(Russo & Johnson, 2000; Salayers & Whitt, 2002; Levine & Edelman, 1984)
Multifunctional nanoparticles
Targeted drug delivery
Nanoparticles- Novel carrier system
Drug incorrporated synthetic & natural polymer nanopartcle
Chitosan- an example of natural polymer
Biodegradable
Biocompatible
Beta-(1-4)-linked D-glucosamine
Catalase & Coagulase tests
MRSA- Catalase & Coagulase +ive





TSI, Citrate, & Indole tests
E. coli
- TSI & Indole +ive & Citrate -ive
Synthesis of Chitosan nanoparticles (CNPs)
Different ratios of Chitosan:TPP:Ceftriaxone

Characterization of CNPs
Nanoparticle size analysis
Particle Size






PDI
blank- 0.31 & Cef-CNPs- 0.462



(Zeta ZF, Malrem UK)
Morphology of CNPs
MIC with ceftriaxone
Entrapment efficiency & Loading capacity
Questions & Answers
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FTIR
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Conclusion
In-vitro study revealed that chitosan blank nanoparticles possess their own antibacterial activity against resistant strains of both Gram positive and Gram negative bacteria
Loading of Ceftriaxone in CNPs gave the enhanced antibacterial acitivity.
Zeta potential
(Greenwood & Kendall, 1999; Hanaor et. al., 2012)
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Dr. Nisar ur Rehman
HOD/Chairman, Dept. of pharmacy, CIIT, Atd.
Dr. M. Arfat Yameen
Assistant prof., Dept. of pharmacy, CIIT, Atd.
Dr. Faiz Rabbani
Assistant prof., Dept. of Chemistry, CIIT, Atd.
Dr. M. Arfan
Assistant prof., School of Natural Schiences, NUST, Isd.

Future prospects

Stability studies

Evaluation of enhanced antibacterial effects of antibiotics against resistant microbes by In-vivo studies.

Targeted antibiotic loaded polymeric nanoparticles against resistant microbes using mAbs.

Toxicological studies of nanoparticles for their safe use.


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Acknowledgments
NIBGE, Faisalabad
NUST, Islamabad.
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