The effect of colloidal silver concentration and metallic copper against some hospital acquired ...

167 Pages·2014·5.72 MB·English

Republic of Iraq
Ministry of Higher Education
And Scientific Research
Baghdad University
College of Science

The effect of colloidal silver concentration and metallic
copper against some hospital acquired infections isolates

Thesis
Submitted to the College of Science/Baghdad University in
Partial Fulfillment of the Requirements for the Degree of
Master of Science in Biotechnology

By

Alyaa Mohameed Hadi

B.Sc., Biotechnology, College of Science, Baghdad University
(2007)

Supervised By

Professor Dr. Alice K. Melconian

2013 1435
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(۱۹ ﺔﻳﻵﺍ) ﻞﻤﻨﻟﺍ ﺓﺭﻮﺳ
Supervisor Certification
We certify that the preparation of this thesis was
made by Alyaa M. Hadi under our supervision at
Baghdad University/ College of Science, as a partial
fulfillment of the requirements for the Master Degree
of Science in Biotechnology.

Signature
Supervisor: Dr. Alice K. Melconian
Title: Professor
Date:

In view of the available recommendation, I forward
this thesis debate by the Examining Committee.

Signature
Name: Dr. Abdul Kareem Al-Kazaz
Head of Biotechnology Department
Date:
Committee Certification
We, the examining committee, certify that we have read
this thesis and examined the student in its contents and that,
according to our opinion, is accepted as a thesis for the
degree of Master of science in Biotechnology.
Signature Signature
Supervisor Name: Dr. Name: Dr.
Title: Title:
Date: Date:

Signature Signature
Name: Dr. Name: Dr.
Title: Title:
Date: Date:

:
Dedication

To my grandfather ... who taught me that even the largest task can be
accomplished if it is done one step at a time .

To my father .... , who gave me a solid foundation in my life .

To my mother .... , who is my rock of support and mentor par excellence

To my husband.... , who is my light in darkness , my comfort and laugh
in sadness .

To my brothers.... , who always supported me with the expression
of encouragement .

To my son.... , my soul , my angel , who always makes me ambitious
to do my best in this life .
Acknowledgments

In the name of ALLAH , most gracious , most merciful
First I would like to express my profound gratitude to my supervisor Professor
Dr.Alice K. Melconian for her guidance , scientific support and useful
suggestions to solve my problems during the period of this study .
I would like to thank the Dean of the college of Science Baghdad university
and the Dean of the Medical Technical Institute ,to the Head and chancery of
Biotechnology department for providing me with this opportunity to
complete my M.Sc study.Grateful acknowledgment is also expressed to the
Bacteriology Culture Laboratory of AL-Yarmouk Teaching Hospital and
Baghdad Teaching Hospital for helping me get bacterial samples .
Many thanks go to Dr.Hanan H. Ghulam for providing me with copper ,
aluminum and stainless steel materials , without forgetting the great help of
my brother Zaid who provided me with different concentrations of colloidal
silver solution from the U.K. I also extend my thanks to AL-Nooman
Hospital for making it possible for me to do my VITEK tests in their
laboratories . Special thanks to Mr. Ammar Khalid who helped me through
the period of research and provided me with all important information about
VITEK 2 compact system manuals. My acknowledgment is also expressed to
M.Sc. student Iman Hatif and Dr. Israa A. Zedan and all the staff of
bacteriology laboratory in the Biotechnology Department. I also thank my
friends and colleagues during my study for being good groups . My sincere
gratitude and love go to my mother , my father , my brothers , my husband
and my son , God bless them all , for supporting me through my life .

Alyaa M. Hadi
August- 2013
Summary I
________________________________________________________________

summary

This study is an attempt to prove that metallic copper and colloidal silver
solutions have antimicrobial activities against different bacterial species which
are considered as a major cause of nosocomial infections . One hundred and
twenty samples were collected from different clinical sources ( urine , blood ,
pus , sputum , stool and wound swabs ) during the period from October /2012
to December /2012 from different hospitals in Baghdad including AL-Yarmouk
teaching hospital and Baghdad teaching hospital . According to the collected
samples , bacterial isolates were distributed as ( 22 ) Escherichia coli isolates
( 18.3%) , ( 12 ) Pseudomonas aeruginosa isolates( 10%) and ( 14 )
Staphylococcus aureus isolates ( 11.6% ). All these isolates were identified
according to morphological and biochemical characteristics by using
conventional methods and VITEK2 compact system for confirmation . The
susceptibility test of the bacterial isolates was determined using (17)
antimicrobials for E.coli isolates , ( 15 ) antimicrobials for P.aeruginosa
isolates and ( 19 ) antimicrobials for S.aureus isolates . All these antimicrobials
used were detected by using VITEK2 compact system . The results showed
variation in resistance to the different antimicrobials tested on bacterial
isolates . From each bacterial species three bacterial isolates with the highest
resistance level were selected for this study . Since the bacterial species E.coli ,
P.aeruginosa and S.aureus are considered both as a major cause of
nosocomial infections and as the most common species persisted in the clinical
samples , these therefore were chosen to(1) evaluate the effectiveness of the
metallic copper to reduce the viability of the selected bacterial isolates
compared with the effect of both aluminum and stainless steel . (2) to evaluate
both the susceptibility of the bacterial isolates to different concentrations of
colloidal silver solutions using the agar-well diffusion method and study its
effect on the viability reduction of bacterial isolates by using plate-viable count.
An inoculum of ( 107 CFU/ml ) from each of the selected bacterial isolates was
inoculated onto coupons ( 1cm x 1 cm ) of copper , aluminum and stainless steel
Summary II
________________________________________________________________

and incubated at room temperature for various time periods ranging from 30
to180 minutes. Viability was determined using the plate -viable count method.
Copper results revealed a complete kill for tested bacterial isolates after 120
minutes of exposure at room temperature indicating a high significant rate of
viability reduction(P<0.0001) . In contrast , viable bacterial colonies were
detected on aluminum and stainless steel coupons after 180 minutes of exposure
. The results showed that the tested isolates were sensitive to colloidal silver
solutions at both concentrations. The killing rate studies for various time
periods ranging from 30 to 180 minutes at room temperature using plate viable
count showed that colloidal silver was highly bactericidal against the tested
bacterial isolates at both concentrations . Complete killing was achieved by 10
ppm concentration after( 180 ) minutes of exposure , which indicates high
significant (P<0.0001) in viability reduction .
Introduction_____________________________________________1___

Introduction

Nosocomial infections cause a substantial burden for health and economics
worldwide (Von Eiff et al.,2001; Weist et al.,2002). The most common
nosocomial infections agents include Staphylococcus aureus , Escherichia coli
and Pseudomonas aeruginosa , with additional concern is the emergence and
dissemination of organisms with increased resistance to antimicrobial agents
(Aavitsland et al.,1992; Lumio,1999). Escherichia coli is an opportunistic
pathogen that belongs to a large bacterial family Enterobactereace which are
Gram negative bacilli ( rod-shaped), facultative anaerobic , which is normally
found as harmless commensalism in human intestine tract . E. coli cause
infections in immunocomprimised patients like intestinal infections ,diarrhea
and extra intestinal infections such as meningitis , septicemia and urinary tract
infections (UTI) (Croxen and Finlay , 2010). P. aeruginosa is a Gram
negative , rod shaped , strictly aerobec pathogen (Todar ,2004). It is an
opportunistic pathogen that causes severe infections in individuals who are
immunocompromised , such as urinary tract infections , respiratory tract
infections , skin infections , especially in patients in the Intensive Care Unit
(ICU) ( Pollack,2000 ; Stover et al., 2000). The infections caused by
Pseudomonas aeruginosa are considered to be dangerous as these bacteria are
inherently resistant to many antimicrobials and have the ability to acquire
resistance to other effective antimicrobials that are commonly used in hospitals
( Gamal et al .,2007 ). Staphylococcus aureus is a Gram positive cocci ,
facultative anaerobic and an opportunistic pathogen that colonizes the skin and
mucosal surfaces of the human host. It causes wide range of infections
ranging ,in severity, from uncomplicated cellulitis , sepsis , impetigo ,to life
threatening infections such as endocarditis , osteomylitis , necrotizing
pneumonia ( Boucher et al.,2010). One of the most important S.aureus species
that cause considerable morbidity and mortality in hospitals is methicillin-
resistant S.aureus (MRSA). Owing to expression of an additional penicillin
Introduction_____________________________________________2___

binding protein, PBP2a, with decreased affinity to β-lactam agents, MRSA is
resistant to all beta-lactam antibiotics, including cephalosporin and
staphylococcal penicillins (Berger-Bachi,1999). Many nosocomial infections
are caused by these three bacterial species because of a prolonged hospital stay
and exposure to contaminated medical devices (Safdar and Maki,2002). Many
studies shows that copper surfaces may help diminish surface-related hygiene
problems . Dry copper surfaces proved to have a great killing efficiency
against a wide range of microbes (Casey et al.,2010 ; Espirito Santo et al.,2010
).The copper free ions accumulate inside the cell and prevent cell reproduction
by damaging the proteins and cell membranes . This will cause cell death
within hours (Gant et al.,2007) . Another antimicrobial agent is the colloidal
silver solution which consists of ultra fine small silver particles that remain
suspended in water. Colloidal silver is different from other conventional
antibiotics , microorganisms cannot build resistance to the action of silver
(Warren ,2003 ). The silver ions act as a catalyst that effectively disable the
metabolism enzymes to take their role ( Hall et al., 1999 ; Iroha et al.,2008 ).
The silver antimicrobial properties have been found extensively useful in
treating some serious infections caused by different bacterial species
( Wickless and Schwader , 2004). Consequently , the aim of the present study
was to succeed in isolation and identification of the methicillin-resistant
S. aureus , E. coli and P. aeruginosa and study the effect of the metallic
copper in comparison with aluminum and stainless steel and the effect of
different concentrations of colloidal silver solution on these isolates.
Several steps were carried out to achieve the aims of this study . They are
outlined as following :

1-Isolation and identification of methicillin-resistant Staphylococcus aureus ,

Escherichia coli and Pseudomonas aeruginosa from different clinical
sources .

2- Determination of the sensitivity of the identified isolates to different
antibiotics to determine the minimum inhibition concentration (MIC) for