Detection of recA gene in Acinetobacter baumannii

Submitted by
Malik Arslan Ali

REG NO: BSMIB02161012

In Partial Fulfillment for the Degree Bachelor of Science (BS) in Microbiology and
Biotechnology

INSTITUTE OF MOLECULAR BIOLOGY AND BIOTECHNOLOGY

THE UNIVERSITY OF LAHORE

2016-2020

i
 "Bismillah al rahman al rahim"

In the name of Allah the most gracious

the most merciful

ii
 AUTHOR’S DECLARATION

I, MALIK ARSLAN ALI hereby state that my (BS Microbiology and Biotechnology) thesis
titled “Detection of recA gene in Acinetobacter baumannin” is my own work and has not
been submitted previously by me for taking any degree from this university “The University
of Lahore” or anywhere else in the country/world.

At any time if my statement is found to be incorrect even after my graduation, the university
has the right to withdraw my (BS Microbiology and Biotechnology) degree.

Malik Arslan Ali

DATE

iii
 PLAGIARISM UNDERTAKING
I solemnly declare that research work presented in the thesis titled “Detection of recA gene in
Acinetobacter baumannii” is solely my research work with no significant contribution from any
other person. Small contribution/help wherever taken has been duly acknowledged and that
complete thesis has been written by me.

I understand the zero-tolerance policy of the HEC and the University “The University of
Lahore” towards plagiarism. Therefore, I as an Author of the above titled thesis declare that the
thesis is complete with no material omitted. Further, no portion of my thesis has been
plagiarized and any material used as reference is properly referred/cited.

I undertake that if I am found guilty of any kind of plagiarism in the above titled thesis even
after award of BS Microbiology and Biotechnology, the university reserves the right to
withdraw/revoke BS Microbiology and Biotechnology degree and that HEC and the
University has the right to publish my name on the HEC/University Website on which
names of students are placed who have submitted plagiarized thesis.
Official Name of the Student: Malik Arslan Ali
Registration No: BSMIB02161012 Date: / /2020
Signature: ________________________

This is to certify that research-work presented in the above mentioned thesis was conducted
under my supervision. I certify that the thesis is complete with no material omitted. Further, no
portion of the thesis has been plagiarized and any material used as reference is properly
referred/cited.

Official Name of the Supervisor: Ms.Nureen Zahra

Employee No: __________________ Date: / /2020
Signature: ________________________

iv
 CERTIFICATE OF APPROVAL

This is to certify that research work presented in the thesis, entitled “Detection of
recA gene in Acinetobacter baumannii” was conducted by Malik Arslan Ali under the
supervision of Ms.NUREEN ZAHRA

No part of this thesis has been submitted anywhere else for any other degree. This
thesis is submitted to the __________________________ in partial fulfillment of
requirements for the degree of BACHELOR OF SCIENCE in the field of
MICROBIOOGY AND BIOTECHNOLOGY, Institute of Molecular Biology and
Biotechnology, The University of Lahore.

Student Name: ___MALIK ARSLAN ALI______________ Signature:

Supervisor Name: __MS.NUREEN ZAHRA__ Signature:

Examination Committee:
a) Dr. Saba Shameem
Associate Professor
IMBB/CRIMM
The University of Lahore,
Lahore, Pakistan
Signature:
Date:

b) Ms. Naureen Zahra

Assistant Professor Signature:
IMBB/CRiMM Date:
The University of Lahore,
Lahore, Pakistan

Director IMBB/CRiMM: Dr. Arif Malik

Signature: Date: / /2020

v
 SUPERVISORY COMMITTEE

We, the Supervisory Committee, certify that the contents and form of thesis submitted by
MALIK ARSLAN ALI (REG. #BSMIB02161012) have been found satisfactory after
receiving all the evaluation reports (External/Internal) and recommended it for the award of the
degree of Bachelor of science (BS) in Microbiology and Biotechnology under title “Detection of
recA gene in Acinetobacter baumannii”.

SUPERVISOR:

Ms. Nureen Zahra

Assistant Professor
IMBB/CRIMM
The University of Lahore,
Lahore, Pakistan

vi
 ACKNOWLEDGEMENTS

I would pay my thanks to Ms.Nureen Zahra my research supervisor for extended support and
guidance at every step. I wish like to express my thanks to the all people who provided me
support in this endeavor.

Malik Arslan Ali

REG.#BSMIB02161012

vii
 CERTIFICATION LETTER

Name of Student: Malik Arslan Ali

Registration No.: Bsmib02161012

Thesis Title: Detection of recA gene in Acinetobacter baumannii

Name of Supervisor: Ms. Nureen Zahra

Department: INSTITUTE OF MOLECULAR BIOLOGY AND

BIOTECHNOLOGY

Overall Similarity Index: ____________________________

Highest Similarity Index to One Single Source: ____________________________

Checked By:

____________________________________
Librarian
Pharmacy Library
The University of Lahore

CC: Director QEC

DIRECTOR ORIC
Respective Head of Department
Office Record

viii
 DEDICATION
THIS RESEARCH STUDY IS DEDICATED TO MY
PARENTS, MY INSPIRATION AND MY SPIRIT WHO
OFFERED ME UNCONDITIONAL LOVE AND
SUPPORT ME ALL THE TIME. WITHOUT WHOM THE
PRESENT PROJECT WOULD HAVE BEEN A MERE
DREAM. WHERE EVER I STAND TODAY IS ALL
BECAUSE OF THEIR GUIDANCE, SUPPORT, WISDOM
AND LOVE”

ix
Abbreviations Full forms

ASR Antimicrobial Sensitivity Test

ICU Intensive Care Unit

MDR Multi Drug Resistant

OXA Oxacillinase

LIST OF ABBREVIATIONS

x
 LIST OF SYMBOLS

SYMBOLS NAME
% Percentage
No Number
C Degree Celsius
M Molar
Ml Milliliter

TABLE OF CONTENTS

xi
"Bismillah al rahman al rahim" II

SUPERVISORY COMMITTEE VI
ACKNOWLEDGEMENT VII
DEDICATION IX
LIST OF ABBREVIATIONS X
LIST OF SYMBOLS XI

CHAPTERS DESCRIPTION PAGE #

CHAPTER ONE INTRODUCTION 01

CHAPTER TWO REVIEW OF LITERATURE 06

CHAPTER THREE MATERIALS AND METHODS 10

CHAPTER FOUR RESULTS 12

CHAPTER FIVE DISCUSSION 14

CHAPTER SIX CONCLUSION 17

CHAPTER SEVEN LITERATURE CITED 19

LIST OF TABLES

S.# DESCRIPTION PAGE. #

1 Sequence of primer used 11

xii
 LIST OF FIGURES

S.# DESCRIPTION PAGE. #

1 PCR results of rec A sequence 13

xiii
 ABSTRACT
Acinetobacter baumannii has become now an important pathogen due to its ability to attain
multidrug resistance property. Nosocomial infections and outbreaks are now common due to this
pathogen. The aim of this study to identify the recA gene which helps to recognize the
A.baumannii species as well help in detection of resistant gene. In tertiary care hospitals of
Pakistan, collected isolates were identified by using standard microbiological technique. Among
these identified samples, this study is aim to detect the specie of Acinetobacter as well the
resistant gene recA by using the specific primer. These results showed that 22(73%) out of
30(100%) isolate were specifically A.baumannii. In order to overcome this threat, it is necessary
to devise some means to counter the antiobiotic resistance and find its mode of transmission of
this pathogen.

Keywords: recA, MDR, Acinetobacter baumanni, PCR

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CHAPTER ONE INTRODUCTION
Arslan, 2020

INTRODUCTION

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CHAPTER ONE INTRODUCTION
Arslan, 2020

Acinetobacter baumannii is basically a non-motile, pleomorphic, aerobic, gram-negative

bacillus, an opportunistic bacterial pathogen mostly related with the hospital infections.
Acinetobacter genus throughout the history of battle between mankind and disease has acquired
an important position. In current time this genus has acquired the central position due to the
multidrug resistant property. Nowadays, multidrug resistant property is becoming a problem,
more like a threat for the mankind (Aoife et al., 2012).

1.1 Historical Background

As far evolutionary history of bacteria is concern it is same for all bacteria. The first authentic
evolutionary concept was explained by the Charles Darwin On the Origin of Species in London
in year 1859. Now 160 years later the basic foundations are same, but genetics of these
foundations is now at whole different level. At the time of discover of Charles Darwin there was
no concept of genetics until the arrival of mendelian genetics (Eugene and Yuri., 2012).

As far evolution of Acinetobacter is concern, it has gone through some large changes in last three
decades. Its resistance to all known antibiotics and association with hospitalized infection is one
of the major reasons it is becoming a global threat. In coming generation antibiotic resistance is
going to be a major issue and it is going to create some serious problems. The taxonomic
historical lineage of Acinetobacter started in 1911 when a Dutch researcher Beijerinck found
isolates of Micrococcus calco-aceticus. After his founding, later on similar organisms were
discovered which were classified into fifteen different genera which included Diplococcus
mucosus, Achromobacter mucosus and Alcaligenes haemolysans. The genus name Acinetobacter
is derived from word akinetos which means non-motile, it was proposed by Brisou and Prévot in
year 1954 to distinguish the non-motile microorganism from the motile microorganisms while
studying the genus Achromobacter. After some time detailed research was conducted which
concluded a single genus Acinetobacter for it and in the year 1971 the Taxonomy
of Moraxella and Allied Bacteria accepted it. (Anton Y et al., 2008).

1.2 Habitat

As far the habitat of A. baumannii is concern it is ubiquitous in nature. It is almost found

everywhere from hospital to the natural reservoirs, except extreme environments. It is mostly
associated with hospital acquired infections. In some cases it was found that may be

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Arslan, 2020

environmental dispersion of this pathogen is primarily resulted from hospital acquired infections
or hospitalized patients. Still there is a lot to discuss about its natural habitat because due to its
increase in multidrug resistant property in recent decade it is found that it is present worldwide
and mostly associated with hospital acquired infections (Luísa C et al., 2014).

1.3 Prevalence

Worldwide multidrug resistant bacteria are emerging as a great threat, in recent years in Latin
America there is remarkable increase in the percentage of multidrug resistant bacteria. As far
bacterial specie of P. aeruginosa and A. baumannii are concern they are showing multidrug
resistant property and remarkable growth at an unprecedented rate. In currently conducted
surveys in different areas of Latin America it was discovered that carbapenem resistance for P.
aeruginosa was 66% and for A. baumannii it was as high 90%. (Mauro J et al., 2014). Where in
Spain when the concluded researches of two decades it was found that carbapenem-resistant
Acinetobacter baumannii prevalence was around 44%. In current scenario at the global threat
this level of prevalence may look much better as compare to the countries where prevailing
percentage is higher than 50% but we cannot blink the fact that this pathogen as a threat and due
to its ubiquitous nature, it can multiply, become more prevalent in no time. It is necessary in
those areas where it is not prevailing as a threat must not be allowed to become one (Asensio et
al., 2008).

As far Latin America and Europe is concerned the prevalence of multidrug resistant
Acinetobacter baumannii prevalence is mostly around 40% to 60% in European countries where
as compared to it in United States from the researches and surveys conducted between 2009 to
2013 it was found that almost 80% infections were caused by A. baumannii. (Bin C et al., 2017)

When we talk about carbapenem resistant A. baumannii, it is basically result of metallo-β-

lactamases and carbapenem-hydrolysing class Dβ-lactamases which restrict and limit the
therapeutic treatments alternatives in tackling the A. baumannii infections. Where in South
Africa the prevalence of Oxacillinase (OXA)-51 gene was 89% among the isolated samples
where OXA-23 had prevalence of 53%. As compared to the Latin American countries this figure
is quite high, and while counting the vulnerability of African countries in future this can create a

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Arslan, 2020

disaster, it is a possible that in near future such bacterias will create such outbreaks which we
haven’t seen since the Black Plague (M.M. Kock et al., 2015).

In China Surveillance of Antimicrobial Resistance Program conducted showed some large

variations and increments of infections rate in last decade. In researches conducted between 2004
to 2014 it was found that prevalence of the imipenem-resistant A. baumannii has elevated from
the 13% to 70% where extensively drug-resistant A. baumannii infections rate has increased
from 11% to 60%. Such elevations in multidrug resistant property is same around the world. But
china decade long program has shown the prospects in long run that how a pathogen can adapt
and attain this property at what rate in how much time. Recently this pathogen has shown
resistant to all known et al., 2017).

In the current scenario in Pakistan the prevalence of multidrug resistant A. baumannii has shown
the results by 100%. The antibiotic susceptibility test depicted that minocycline and tigecycline
drug showed most effective results against A. baumannii.( Shahzeera B et al., 2013). Where in
2014 research and sample collection showed some different results, among the isolated samples
from all over the Pakistan 66% were resistant to carbapenem resistant (Badrul et al., 2014).

1.4 Multidrug Resistant

A. baumannii is almost responsible for about 9% to 10% infections caused by gram-negative

bacteria. It is emerging as a global threat due to its ability to develop antibiotic resistance, which
is mostly result from adaptation of gene clusters of transposons and or integrons (Hervé Richet
and Pierre Edouard Fournier., 2006)

In last decade in America A. baumannii has shown particular increase in multidrug resistant
property. In a research conducted in central Ohio it was found that among the clinical isolates
there were different distinguish pulsotypes. It was found that among the clinical isolates
blaOXA-23 was 13% and IS Aba1 linked blaOXA-66 was 79.5%, which basically showed the
dissemination of imipenem drug resistant (Vijaya B et al., 2009). But in recent times some
advance researches are ongoing on multidrug resistant property and in isolates collected from
Nashville General Hospital when it was conformed that they are depicting high drug resistivity,

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CHAPTER ONE INTRODUCTION
Arslan, 2020

their molecular diagnosis was done on which it was found that superoxide dismutase B protein
played an important role in antibiotic resistance. Proteomic analysis was done and by the help of
multidimensional protein identification technology it was found that during high antibiotic
resistance activity there was forty-fold increase in protein differentiation activity (Steven D et
al., 2019). In research conducted from year 2006 to 2016 it was found that Mortality was 81.6%
in majority of the patients. High non-susceptibility rates were around 100 to 10.7% for the
evaluated antimicrobials which includes colistin, tigecycline and polymyxin B, where most
isolates were classified as extensively drug-resistant ,around 78%. Carbapenemase producing
isolates were about 92% among the isolates. Carbapenem-resistant isolates depicted that the
carbapenem-hydrolyzing class D β-lactamase were either blaOXA-23-like which were around
97.9% or blaOXA-143-like which were 2.1% (Priscila R et al., 2019). In 2018 a research was
conducted to find out the characterization of the carbapenemase genes which were responsible
for carbapenem resistivity, it was found that in 90% cases OXA-23 gene was responsible, where
for NDM it was 66.7% and GES was 50% (Raghdaa A et al., 2018). In 2011 a study was
conducted on recA gene and it was found that it was responsible for the SOS response, it was
responsible for the DNA damage and in this way, it generates a mutant recA. Later on, it was
found this gene was responsible for the survival in harsh and stressful conditions like in absence
of this gene there was decrease in survival ability of this pathogen. Other than this it was also
found that it was related to the multidrug resistant property in mutant form (Jesús A et al., 2011)

Worldwide antibiotic resistance property of this pathogen is becoming a threat and

Acinetobacter baumannii is basically a pathogen which is ubiquitous in nature, it is the main
reason it is emerging as global pathogen. In Pakistan, multidrug resistant property is also
becoming an imminent threat. In current scenario there is no demographic data and genotyping
of multidrug resistant property of this pathogen. In Present study of genotyping pattern of this
gene on samples collected from surrounding area we have 150 samples of pus and blood to
evaluate and analyze the Acinetobacter baumannii. Later, it was identified that among them 30
were the isolates of Acinetobacter baumannii, so in present study we will be analyzing them
genotypically and developing the results on basis of our demographic data. Our main target is
mutant form recA gene which is responsible for antibiotic resistance.

1.5 Main Objective

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CHAPTER ONE INTRODUCTION
Arslan, 2020

Our main objective is the identification Acinetobacter baumannii recA gene for multidrug
resistant property among the collected isolates.

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CHAPTER TWO REVIEW OF LITERATURE
Arslan, 2020

REVIEW OF LITERATURE

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CHAPTER TWO REVIEW OF LITERATURE
Arslan, 2020

In research conducted on samples from Nashville General Hospital in United states it was found
that almost 72% of the samples of A. baumannii were multidrug resistant. When it was
conformed that they are depicting high drug resistivity, their molecular diagnosis was done on
which it was found that superoxide dismutase B protein played an important role in antibiotic
resistance. Proteomic analysis was done and by the help of multidimensional protein
identification technology it was found that during high antibiotic resistance activity there was
forty-fold increase in protein differentiation activity (Steven D et al., 2019).

In research conducted from year 2006 to 2016 in Brazil it was found that Mortality was 81.6% in
majority of the patients due to A. baumannii. The samples collected from hospital in Londrina,
Brazil it was found that high non-susceptibility rates were around 100% to 10.7% for the
evaluated antimicrobials which includes colistin, tigecycline and polymyxin B, where most
isolates were classified as extensively drug-resistant ,around 78%. Carbapenemase producing
isolates were about 92% among the isolates. Carbapenem-resistant isolates depicted that the
carbapenem-hydrolyzing class D β-lactamase were either blaOXA-23-like which were around
97.9% or blaOXA-143-like which were 2.1% (Priscila R et al., 2019).

In wastewater treatment plants in Zagreb, Croatia it was found that in these plants A. baumannii
found, when they were isolated and tested for multidrug resistant property it was found that in
these wastewater treatment plants about 85% of the isolates were carbapenem resistant. As far
ubiquitous nature of this pathogen is concern it is not astonishing that it is found in wastewater
treatment. Where with carbapenem resistant isolates presence of OXA-23 gene was also
observed (Paul G et al., 2018)

In 2017 a research was conducted on A. baumannii due to its increasing antibiotic resistance. The
antibacterial effect of herbal oil which was extracted from 10 plants were tested which included
ginger, plai, clove and cinnamon. It was found later on that herbal oil extracted from cinnamon
exhibited potent antibacterial activity where the case with other plants extracted oil doesn’t
produce any viable results (Amornrat I et al., 2017)

In current researches and ongoing clinical trials, it is getting clear that issue of multidrug
resistant property is becoming a very big problem. But now new methods are being employed to
tackle the multidrug resistant property, in case of A. baumannii infections a new method is

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CHAPTER TWO REVIEW OF LITERATURE
Arslan, 2020

developed in which the antibiotcs are combined with the silver nanoparticles to treat the
infection. As far carbapenem resistant isolates are concern it is found that silver nano-particles
have the ability to completely restrict the antibiotic resistance ability of this pathogen when they
are used in combination with antibiotics (Guoqing W et al., 2016)

In China samples collected from the Central Hospital of Zhumadian it was found that four
groups of blaOXA-24, blaOXA-23, blaOXA-58 and blaOXA-51 were related to the OXA gene.
They were all present in multidrug resistant A. baumannii. From the PCR it was found that only
two of them OXA-51 and OXA-23 were responsible for the multidrug resistant property
(Chenggong Hou1 and Feng Yang., 2015)

A. baumannii multidrug resistivity is becoming a global problem and due to its association with
nosocomial infections it is also a common pathogen with hospital acquired infections. Among
the patients treated at Clinic of Anesthesiology and Reanimation at the University Clinical
Centre Tuzla, Bosnia it was found that among the registered patients’ nosocomial infections was
reported in 12% and about 51% of these infections were result of A. baumannii (Custovic et al.,
2014)

Bacterial persistence is a process by which bacteria survive the high concentration of antibiotics.
It is basically reported in may bacterias but as far A. baumannii is concern there was no
evaluation. Bacterial persistence result from the formation of the persistor cells. When research
was conducted on this pathogen it was found that cultures treated with the Polymyxin-B showed
the persister cells formation about 0.0007% to 10.1% (Valdir C et al., 2013)

With increase in infections of multidrug resistant A. baumannii it is a necessity to have such

methods by which we can identify quickly such pathogens. Mass spectrometry profile created by
the matrix-assisted laser desorption ionization-time of flight is a method which is developed to
detect the carbapenem resistant A. baumannii with specificity (Marie K et al., 2012)

With rise of multidrug resistant property, variations in genes are also occurring at a very fast rate
in a patient which was transfer from Egypt to Germany it was found that infection was result of
A. baumannii. When molecular characterization was done it was found that there was a variant
form of NDM-1 which was responsible for the carbapenem resistance, the variant was named
NDM-1 (Martin K et al., 2011).

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CHAPTER TWO REVIEW OF LITERATURE
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Despite the increment in multidrug resistant property of the A. baumannii in some areas third
degree of antibiotics are stiff effective. In Iran between year 2005 and 2006, 88 isolates were
separated and when antibiotic resistance was done it was found that 90% were showing
resistance to the ceftriaxone and piperacillin, 84% to the ceftazidime, 85% to the amikacin and
90% to the ciprofloxacin. As compared to them Imipenem was most effective drug with the
resistance among patients of 4.5%, from this it was found that imipenem was still in efficient
usage against these infections (Mohammad R et al., 2010)

Antibiotic resistance of A. baumannii is becoming a global threat and researchers are trying to
find out causes of this issue and ways to tackle this issue, where in some areas different
associations are studied between antibiotic resistance and other properties. In this context it was
found that biofilm forming ability of this pathogen might be related to the antibiotic resistance.
Where biofilm formation and regulation itself depend on various extracellular and intracellular
components which includes transcriptional regulatory system and bacterial cell density (Jennifer
A. Gaddy and Luis A. Actis., 2009)

In current control A. baumannii it was found that it is developing antibiotic resistance at an

alarming rate. This pathogen is responsible for majority of nosocomial infections acquired from
hospitals. As far carbapenem is concern in majority of areas in context of modern research it is
found that it is getting resistant against it where use of sulbactam is also becoming ineffective to
some extent due to antibiotic resistance ability. Where polymyxins are also being used for
antimicrobial activity. Where now methods and techniques being devised for this purpose
(Drosos and Matthew., 2008)

A. baumannii infections in Burn Intensive Care Unit have high mortality rate due to infections
caused by this pathogen. In current researches the result showed that 87% isolates were resistant
to the imipenem where as compared to it no pathogen showed resistant to the colistin. In 70% of
the cases cure was achieved where mortality rate was 30% in these cases. Burn intensive Care
Unit cases showed high infection rate because of the A. baumannii (Vincent T et al., 2007)

A. baumannii the clear ability to develop the extraordinary ability the antibiotic resistance. When
its strain where compared with a refrence strain it was found that multidrug resistant strain
showed the diminished levels of penicillin binding protein expression, where it also showed that

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it have higher level of the biofilm formation with the accumulation of the outer membrane
protein have a role in bacterial adhesion (Axel S et al., 2006)

In Kosin University Gospel Hospital in Busan, Korea there was outbreak of the A. baumannii
which involved 36 cases, later on when molecular characterization and sequencing was done it
was found that multidrug resistant phenotype remained unchanged over the long period of time.
When molecular characterization was done it was found that epidemic isolates were carrying
blaOXA-23, which was responsible for the resistant against the penicillins, carbapenems,
cephamycins and aminoglycosides. (Byung-Chan et al., 2005)

In India crude oil degrading bacterias were discovered, out of them 25 were strains of A.
baumannii, all of these strains were capable of degradation of hydrocarbons of crude oil. There
were total 8 phenovars considering their substrate usage profiles. When these variants were
analysed in regard to their ability to degrade the crude oil it was discovered that all strains have
different extent of degradation. In future these strains can play an important role in situ
bioremediation (Priyangshu M et al., 2004)

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CHAPTER THREE MATERIALS & METHODS
Arslan, 2020

MATERIALS AND METHODS

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CHAPTER THREE MATERIALS & METHODS
Arslan, 2020

Phenotypically detected sample were used in this study. Genotypic resistance pattern was
observed in 30 samples of phenotypically detected samples of A. baumannii for recA.

3.1 Molecular identification of resistant gene of A. baumannii

3.2 Extraction of DNA and gel electropherosis

Multidrug resistant strain of A. baumannii were molecularly identified afterwards their DNA
extracted by usage of WizPr e gDNA mini Kit and results were later confirmed by gel
electrophoresis.

3.3 Polymerase chain reaction

The amplification of resistant gene was done by usage of specific primer and standard protocol.
After this the results were observed on 1.5% agarose gel in TAE buffer with 100bp ladder.

Table 3.1 Sequence of primer used

Primer Sequence 5’-3’ Amplicon Reference

Name size
recA-F GATCGGATTGGAGAACCAGA 387 Aranda, J., Bardina, C., Beceiro,
recA-R ATTTCTGACCGCATTTCCAT A., Rumbo, S., Cabral, M. P.,
Barbé, J., & Bou, G. (2011).
Acinetobacter baumannii RecA
protein in repair of DNA damage,
antimicrobial resistance, general
stress response, and virulence.
Journal of bacteriology, 193(15),
3740–3747.
https://doi.org/10.1128/JB.00389-
11

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RESULTS
CHAPTER FIVE DISCUSSION
ARSLAN, 2020

In our study and research on phenotypically identified samples A. baumannii of genotypic

resistance pattern of A. baumannii of recA gene were observed in 73% (22 samples out of 30) of
the samples.

CHAPTER FOUR RESULTS

First name, 2018

Fig 4.4 PCR results of recA sequence (100bp ladder. 1.5% gel)

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CHAPTER FIVE DISCUSSION
ARSLAN, 2020

DISCUSSION

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CHAPTER FIVE DISCUSSION
ARSLAN, 2020

Jesús A et al., 2011 study conducted on recA gene showed that it was responsible for the SOS
response, it was responsible for the DNA damage and in this way, it generates a mutant recA.
Later on, it was found this gene was responsible for the survival in harsh and stressful conditions
like in absence of this gene there was decrease in survival ability of this pathogen. Other than
this it was also found that it was related to the multidrug resistant property in mutant form where
as compared to it in our results showed the increased prevalence of the A. baumannii as depicted
by our results. From analytical point of view there is a possibility that this gene not only poses a
threat due to its multidrug resistant ability but also increasing the chances of survival of pathogen
in stressful conditions. In comparison to the present work recA gene is responsible for its
resistance because its act as housekeeping gene and in phenotypically resistance isolates
indicated its genotypic resistance as well that is 73% .

Shahzeera et al., 2013 results showed that the prevalence of multidrug resistant A. baumannii
were 100% among their samples which were metallo β-lactamase and carbapenemase producers,
AmpC prevalence was 41 %where as compared to them our researched showed that recA was
more than in half of our collected samples from tertiary care hospitals Lahore. We can’t neglect
the possibility that these samples can be metallo β-lactamase and carbapenemase producers.
While in comparison to its present work indicates that recA gene because of being a
housekeeping gene show resistant property where phenotypically identified isolates showed
genotypic presence of recA gene in 73% of the isolates.

Badrul et al., 2014 results showed that threre were 65% of its isolates were resistant to
carbapenems due to the presence of New Delhi metallo-β-lactamase-1 and oxacillinase where in
our results presence of recA was more than 50%. In comparision to these results it is clear that in
recent time the multidrug resistance of this pathogen is rising as depicted by our research. As
compared to the multidrug resistance A. baumannii in Pakistan this pathogen has shown
remarkable growth in hospital acquired infections. There is quite a possibility that our collected
isolates also possess the New Delhi metallo-β-lactamase-1 and oxacillinase. Due to the
ubiquitous nature of this pathogen is found everywhere and this is one of the main reasons for its
association in all countries and all environments. A. baumannii is ubiquitous in nature because of
its infections are widespread it is also show MDR property. In comparison to this current work
depicted that recA gene is a major culprit for MDR property because of being a part of

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ARSLAN, 2020

housekeeping genes it is becoming a major threat as indicated by presence of its in 73% of

isolates.

Recently Abbas et al., 2018 researched showed that there was a relationship between reduced
expression of the recA gene, with increased expression of the lpsB, dnaK, and blsA are related to
the increased colistin resistance where as compared to our study shows that recA gene was
present in more than half of the collected samples so there is a possibility that due to the increase
presence of recA gene there will be less expression of expression of the lpsB, dnaK, and blsA are
related to the increased colistin resistance. As compared to it in current work study shows it is
evident that the prevalence of multidrug resistant A. baumannii is increasing as depicted presence
of recA gene in 73% of identified isolates.

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CHAPTER SIX
CONCLUSION
ARSLAN,2020

CONCLUSION
CHAPTER SIX
CONCLUSION
ARSLAN,2020
This study showed the increment in rate of nosocomial outbreaks in Pakistan. In this study we
did the molecular characterization of A.baumannii and identified recA gene which is responsible
for the multidrug resistant property. Our results showed that among the isolated samples acquired
from tertiary care hospitals 73% isolates have recA gene. In order to overcome this widespread
problem, we need to take proper measures to overcome the threat of multidrug resistant property
of this pathogen.
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LITERATURE CITED

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