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Microbial prevalence and antimicrobial susceptibility of wound isolates from tertiary institution
CHAPTER ONE
INTRODUCTION
1.0 Background of the study
A wound is a type of injury which happens relatively quickly in which
skin is torn, cut, or punctured (an open wound), or where blunt force trauma
causes a contusion (a closed wound). In pathology, it specifically refers to a
sharp injury which damages the dermis of the skin. Intact skin is the perfect
defence to bacterial invasion, but damage to the skin allows bacteria, fungi and
yeasts to enter (Young, 2012). More than 200 different species of bacteria
normally live on the skin1 and an open wound provides a moist, warm and
nutritious environment perfect for microbial colonisation and proliferation.
Bacteria colonise all chronic wounds and low levels of bacteria can benefit the
wound by increasing the amount of neutrophils, monocytes and macrophages
in the wound, thus improving levels of prostaglandin E2 and the formation of
collagen (Edwards, & Harding, 2004). When one or more microorganisms
multiply in the wound, local and systemic responses occur in the host, which
can lead to infection and a subsequent delay in healing (Angelet al.,2011).
Maintaining the bacteria at a level at which the host is in control is an
important part of avoiding wound infection (Cutting, 2010).
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Regardless of large amounts of bacteria, many wounds continue to heal
well. The ability of the patients immune system to deal with bacteria (host
response) and the type and amount of bacteria involved determines whether
clinical problems will occur (Young, 2012). Chronic wounds are open for
extended periods of time and the patients usually have underlying disease
processes, which leads to heavy colonisation with bacteria and/or fungi (Young,
2012). When chronic wounds are poorly perfused they are more susceptible to
infection, as blood delivers oxygen, nutrients and immune cells, thus providing
little opportunity for microorganisms to colonise and proliferate (Bowler et al.,
2001). Devitalised tissue, combined with fluid and nutrients from wound
exudate provide an ideal setting for bacterial proliferation (Cutting 2010). The
host response can often be improved by correcting or improving the underlying
diseases (Young, 2012).
1.2.0Aim and Objectives
Aim
This study is aimed at determining the Microbial prevalence and
antimicrobial susceptibility of wound isolates from tertiary institution.
The specific objectives were to:
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Microbial prevalence and antimicrobial susceptibility of wound isolates from tertiary institution
1.Isolate, characterize and identify the morphological biochemical test of
wound collected from tertiary institution
2.To identify types of Microbial prevalence over wound
3.To determine antimicrobial susceptibility of wound isolates from tertiary
institution
CHAPTER TWO
LITERATURE REVIEW
Overview of Wounds
A wound is a breakdown in the protective function of the skin; the loss of
continuity of epithelium, with or without loss of underlying connective tissue
(Bowler et al., 2001). Wounds can be accidental, pathological or post-operative.
An infection of this breach in continuity constitutes wound infection. Wound
infection is thus the presence of pus in a lesion as well as the general or local
features of sepsis such as pyrexia, pain and induration. Infection is believed to
occur when virulence factors expressed by one or more microorganisms in a
wound out-compete the host natural immune system (Bowler et al., 2001).
Wound infection is important in the morbidity and mortality of patients
irrespective of the cause of the wound. It is also important because it can delay
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healing and cause wound breakdown (Alexander, 1994). This is also associated
with longer hospital stay and increased cost of healthcare (Sule et al., 2002).
Wound infections are also significant in that they are the most common
nosocomial infection (Dionigi et al., 2001).
Studies on wound infection have largely focused on surgical site infections
(Sands et al., 1996; Garner, 1986 and Gaynes etal., 2001). This might be
because other types of wound infection are not problematic in the developed
world where most of these studies have been done. However, in developing and
resource-poor countries, other types of wound infection in addition to surgical
site infection are still important causes of morbidity and mortality (Mehta et al.,
2007; Anguzu & Olila, 2007; Fadeyi et al., 2008). Where studies have been
done on wound infections generally, regional and local variations have been
observed in terms of the causative micro-organisms (Sule et al., 2002; Wariso &
Nwachukwu, 2003; Egbe et al., 2011). This means that physicians need to
know the prevalent organisms and the resistance patterns existing in their
localities.
Classification
According to level of contamination a wound can be classified as
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Microbial prevalence and antimicrobial susceptibility of wound isolates from tertiary institution
Clean wound, a wound made under sterile conditions where there are no
organisms present in the wound and the wound is likely to heal without
complications (Kidd et al., 2000).
Contaminated wound, where the wound is as a result of accidental
injury where there are pathogenic organisms and foreign bodies in the
wound.
Infected wound, where the wound has pathogenic organisms present and
multiplying showing clinical signs of infection, where it looks yellow,
oozing pus, having pain and redness (Kidd et al., 2000).
Colonized wound, where the wound is a chronic one and there are a
number of organisms present and very difficult to heal as in a bedsore
(Kidd et al., 2000).
Abrasion
In dermatology, an abrasion is a wound caused by superficial damage to
the skin, no deeper than the epidermis. It is less severe than a laceration, and
bleeding, if present, is minimal. Mild abrasions, also known as grazes or
scrapes, do not scar or bleed, but deep abrasions may lead to the formation of
scar tissue. A more traumatic abrasion that removes all layers of skin is called
an avulsion (Kidd et al., 2000).
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Abrasion injuries most commonly occur when exposed skin comes into
moving contact with a rough surface, causing a grinding or rubbing away of
the upper layers of the epidermis (Kidd et al., 2000).
The abrasion should be cleaned and any debris removed. A topical
antibiotic (such as Neosporin or bacitracin) should be applied to prevent
infection and to keep the wound moist (Kidd et al., 2000). Dressing the wound
is optional but helps to keep the wound from drying out which interferes with
healing. If the abrasion is painful, a topical analgesic (such as lidocaine or
benzocaine) can be applied, but for large abrasions. A systemic analgesic may
be necessary. Avoid exposing abraded skin to the sun as permanent
hyperpigmentation can develop (Kidd et al., 2000).
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Fig 2.1: Abrasion on the palm of a right hand, shortly after falling
Sources: Kidd et al., 2000
Chronic wound
A chronic wound is awoundthat does not heal in an orderly set of stages and
in a predictable amount of time the way most wounds do; wounds that do not
heal within three months are often considered chronic (Robert, 2005). Chronic
wounds seem to be detained in one or more of thephases of wound healing.
For example, chronic wounds often remain in theinflammatorystage for too
long (Robert, 2005; Taylor et al., 2005). In acute wounds, there is a precise
balance between production and degradation ofmoleculessuch ascollagen; in
chronic wounds this balance is lost and degradation plays too large a role
(Edwards et al., 2004; Schnfelder et al., 2005).
Chronic wounds may never heal or may take years to do so. These wounds
cause patients severe emotional and physicalstressand create a significant
financial burden on patients and the whole healthcare system (Augustin &
Maier, 2003).
Acute and chronic wounds are at opposite ends of a spectrum of wound healing
types that progress toward being healed at different rates (Kathleen, 2005).
Signs and Symptoms
https://en.wikipedia.org/wiki/Woundhttps://en.wikipedia.org/wiki/Wound_healinghttps://en.wikipedia.org/wiki/Inflammationhttps://en.wikipedia.org/wiki/Moleculehttps://en.wikipedia.org/wiki/Collagenhttps://en.wikipedia.org/wiki/Stress_(medicine)https://en.wikipedia.org/wiki/Wound_healinghttps://en.wikipedia.org/wiki/Inflammationhttps://en.wikipedia.org/wiki/Moleculehttps://en.wikipedia.org/wiki/Collagenhttps://en.wikipedia.org/wiki/Stress_(medicine)https://en.wikipedia.org/wiki/Wound7/25/2019 Ese Project
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Chronic wound patients often report pain as dominant in their lives (Krasner,
1998). It is recommended that healthcare providers handle the pain related to
chronic wounds as one of the main priorities in chronic wound management
(together with addressing the cause). Six out of ten venous leg ulcer patients
experience pain with their ulcer (Hofman, 1997), and similar trends are
observed for other chronic wounds.
Persistent pain (at night, at rest, and with activity) is the main problem for
patients with chronic ulcers (Catherine, 2006). Frustrations regarding
ineffective analgesics and plans of care that they were unable to adhere to were
also identified.
Cause
In addition to poor circulation, neuropathy, and difficulty moving, factors that
contribute to chronic wounds include systemic illnesses, age, and repeated
trauma. Comorbid ailments that may contribute to the formation of chronic
wounds include vasculitis (an inflammation of blood vessels), immune
suppression, pyoderma gangrenosum, and diseases that cause ischemia
(Robert, 2005). Immune suppression can be caused by illnesses or medical
drugs used over a long period, for example steroids (Robert, 2005). Emotional
stress can also negatively affect the healing of a wound, possibly by raising
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blood pressure and levels of cortisol, which lowers immunity (Augustin & Maier,
2003). What appears to be a chronic wound may also be a malignancy; for
example, cancerous tissue can grow until blood cannot reach the cells and the
tissue becomes an ulcer (Hofman et al., 1997). Cancer, especially squamous
cell carcinoma, may also form as the result of chronic wounds, probably due to
repetitive tissue damage that stimulates rapid cell proliferation (Hofman et al.,
1997). Another factor that may contribute to chronic wounds is old age
(Thomas, 2004). The skin of older people is more easily damaged, and older
cells do not proliferate as fast and may not have an adequate response to stress
in terms of gene upregulation of stress-related proteins (Thomas, 2004). In
older cells, stress response genes are overexpressed when the cell is not
stressed, but when it is, the expression of these proteins is not upregulated by
as much as in younger cells (Thomas, 2004). Comorbid factors that can lead to
ischemia are especially likely to contribute to chronic wounds. Such factors
include chronic fibrosis, edema, sickle cell disease, and peripheral artery
disease such as by atherosclerosis (Robert, 2005). Repeated physical trauma
plays a role in chronic wound formation by continually initiating the
inflammatory cascade. The trauma may occur by accident, for example when a
leg is repeatedly bumped against a wheelchair rest, or it may be due to
intentional acts. Heroin users who lose venous access may resort to 'skin
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popping', or injecting the drug subcutaneously, which is highly damaging to
tissue and frequently leads to chronic ulcers (Williams & Southern, 2005).
Children who are repeatedly seen for a wound that does not heal are sometimes
found to be victims of a parent with Munchausen syndrome by proxy, a disease
in which the abuser may repeatedly inflict harm on the child in order to receive
attention (Vennemann et al., 2006).
Pathophysiology
Chronic wounds may affect only the epidermis and dermis, or they may affect
tissues all the way to the fascia (Crovetti et al., 2004). They may be formed
originally by the same things that cause acute ones, such as surgery or
accidental trauma, or they may form as the result of systemic infection,
vascular, immune, or nerve insufficiency, or comorbidities such as neoplasias
or metabolic disorders (Crovetti et al., 2004). The reason a wound becomes
chronic is that the bodys ability to deal with the damage is overwhelmed by
factors such as repeated trauma, continued pressure, ischemia, or illness
(Crovetti et al., 2004). Though much progress has been accomplished in the
study of chronic wounds lately, advances in the study of their healing have
lagged behind expectations. This is partly because animal studies are difficult
because animals do not get chronic wounds, since they usually have loose skin
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that quickly contracts, and they normally do not get old enough or have
contributing diseases such as neuropathy or chronic debilitating illnesses
(Thomas, 2004). Nonetheless, current researchers now understand some of the
major factors that lead to chronic wounds, among which are ischemia,
reperfusion injury, and bacterial colonization (Thomas, 2004).
Ischemia
Ischemia is an important factor in the formation and persistence of wounds,
especially when it occurs repetitively (as it usually does) or when combined
with a patients old age (Thomas, 2004). Ischemia causes tissue to become
inflamed and cells to release factors that attract neutrophils such as
interleukins, chemokines, leukotrienes, and complement factors (Thomas,
2004). While they fight pathogens, neutrophils also release inflammatory
cytokines and enzymes that damage cells (Thomas, 2004; Robert, 2005). One of
their important jobs is to produce Reactive Oxygen Species (ROS) to kill
bacteria, for which they use an enzyme called myeloperoxidase (Thomas, 2004).
The enzymes and ROS produced by neutrophils and other leukocytes damage
cells and prevent cell proliferation and wound closure by damaging DNA, lipids,
proteins (Alleva et al., 2005), the extracellular matrix (ECM), and cytokines that
speed healing (Thomas, 2004). Neutrophils remain in chronic wounds for longer
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than they do in acute wounds, and contribute to the fact that chronic wounds
have higher levels of inflammatory cytokines and ROS (Taylor et al., 2005).
Since wound fluid from chronic wounds has an excess of proteases and ROS,
the fluid itself can inhibit healing by inhibiting cell growth and breaking down
growth factors and proteins in the ECM. This impaired healing response is
considered uncoordinated (Krishnaswamy et al., 2014). However, soluble
mediators of the immune system (growth factors), cell-based therapies and
therapeutic chemicals can propagate coordinated healing (Lasagni et al., 2010).
It has been suggested that the three fundamental factors underlying chronic
wound pathogenesis are cellular and systemic changes of aging, repeated bouts
of ischemia-reperfusion injury, and bacterial colonization with resulting
inflammatory host response (Mustoe, 2004).
Bacterial Colonization
Since more oxygen in the wound environment allows white blood cells to
produce ROS to kill bacteria, patients with inadequate tissue oxygenation, for
example those who suffered hypothermia during surgery, are at higher risk for
infection (Thomas, 2004). The hosts immune response to the presence of
bacteria prolongs inflammation, delays healing, and damages tissue (Thomas,
2004). Infection can lead not only to chronic wounds but also to gangrene, loss
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of the infected limb, and death of the patient. More recently, an interplay
between bacterial colonization and increases in reactive oxygen species leading
to formation and production of biofilms has been shown to the generate chronic
wounds (Dhall, 2014). Like ischemia, bacterial colonization and infection
damage tissue by causing a greater number of neutrophils to enter the wound
site (Robert, 2005). In patients with chronic wounds, bacteria with resistances
to antibiotics may have time to develop (Halcon & Milkus, 2004). In addition,
patients that carry drug resistant bacterial strains such as methicillin-
resistant Staphylococcus aureus (MRSA) have more chronic wounds (Halcon &
Milkus, 2004).
Treatment
Though treatment of the different chronic wound types varies slightly,
appropriate treatment seeks to address the problems at the root of chronic
wounds, including ischemia, bacterial load, and imbalance of proteases
(Thomas, 2004). Various methods exist to ameliorate these problems, including
antibiotic and antibacterial use, debridement, irrigation, vacuum-assisted
closure, warming, oxygenation, moist wound healing, removing mechanical
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stress, and adding cells or other materials to secrete or enhance levels of
healing factors (Velander et al., 2004).
Preventing and Treating Infection
To lower the bacterial count in wounds, therapists may use topical antibiotics,
which kill bacteria and can also help by keeping the wound environment moist
(Brem, 2004; Patel, 2000), which is important for speeding the healing of
chronic wounds (Taylor et al., 2005; Thomas et al., 2005). Some researchers
have experimented with the use of tea tree oil, an antibacterial agent which
also has anti-inflammatory effects (Halcon & Milkus, 2004). Disinfectants are
contraindicated because they damage tissues and delay wound contraction
(Patel et al., 2000). Further, they are rendered ineffective by organic matter in
wounds like blood and exudate and are thus not useful in open wounds.[32]
A greater amount of exudate and necrotic tissue in a wound increases
likelihood of infection by serving as a medium for bacterial growth away from
the hosts defenses (Thomas, 2004). Since bacteria thrive on dead tissue,
wounds are often surgically debrided to remove the devitalized tissue (Patel et
al., 2000). Debridement and drainage of wound fluid are an especially
important part of the treatment for diabetic ulcers, which may create the need
for amputation if infection gets out of control. Mechanical removal of bacteria
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and devitalized tissue is also the idea behind wound irrigation, which is
accomplished using pulsed lavage (Thomas, 2004). Removing necrotic or
devitalzed tissue is also the aim of maggot therapy, the intentional introduction
by a health care practitioner of live, disinfected maggots into non-healing
wounds. Maggots dissolve only necrotic, infected tissue; disinfect the wound by
killing bacteria; and stimulate wound healing. Maggot therapy has been shown
to accelerate debridement of necrotic wounds and reduce the bacterial load of
the wound, leading to earlier healing, reduced wound odor and less pain. The
combination and interactions of these actions make maggots an extremely
potent tool in chronic wound care.
Negative pressure wound therapy (NPWT) is a treatment that improves ischemic
tissues and removes wound fluid used by bacteria (Thomas, 2004; Kathleen,
2005). This therapy, also known as vacuum-assisted closure, reduces swelling
in tissues, which brings more blood and nutrients to the area, as does the
negative pressure itself (Kathleen, 2005). The treatment also decompresses
tissues and alters the shape of cells, causes them to express different mRNAs
and to proliferate and produce ECM molecules (Kathleen, 2005; Robert, 2005).
Treating Trauma and Painful Wounds
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Persistent chronic pain associated with non-healing wounds is caused by
tissue (nociceptive) or nerve (neuropathic) damage and is influenced by
dressing changes and chronic inflammation. Chronic wounds take a long time
to heal and patients can suffer from chronic wounds for many years.[33]
Chronic wound healing may be compromised by coexisting underlying
conditions, such as venous valve backflow, peripheral vascular disease,
uncontrolled edema and diabetes mellitus. If wound pain is not assessed and
documented it may be ignored and/or not addressed properly. It is important to
remember that increased wound pain may be an indicator of wound
complications that need treatment, and therefore practitioners must constantly
reassess the wound as well as the associated pain.
Optimal management of wounds requires holistic assessment.
Documentation of the patients pain experience is critical and may range from
the use of a patient diary, (which should be patient driven), to recording pain
entirely by the healthcare professional or caregiver (Osterbrink, 2003). Effective
communication between the patient and the healthcare team is fundamental to
this holistic approach. The more frequently healthcare professionals measure
pain, the greater the likelihood of introducing or changing pain management
practices.
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At present there are few local options for the treatment of persistent pain,
whilst managing the exudate levels present in many chronic wounds. Important
properties of such local options are that they provide an optimal wound healing
environment, while providing a constant local low dose release of ibuprofen
during war time.
If local treatment does not provide adequate pain reduction, it may be
necessary for patients with chronic painful wounds to be prescribed additional
systemic treatment for the physical component of their pain. Clinicians should
consult with their prescribing colleagues referring to the WHO pain relief
ladder of systemic treatment options for guidance. For every pharmacological
intervention there are possible benefits and adverse events that the prescribing
clinician will need to consider in conjunction with the wound care treatment
team.
Diagnosis of Wounds
Diagnostic and therapeutic devices have gone through a great development
since the invention of Ignc Semmelweis published in 1846. Paradoxically
opportunities of healthcare represent the most important factor of recent
nosocomial infections (Katalin, 2010).
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Diagnosis of wound infection can be a daunting task in resource-poor settings.
There is often a lack of adequate diagnostic equipment or requisite personnel
(Hart and Kariuki, 1998). Thus, a diagnostic dilemma confronts physicians in
the absence of local epidemiological data on wound infections which could aid
empiric treatment. This dilemma coupled with the fact that there are no
established evidence-based clinical practice guidelines for wound infections,
makes management of wound infections difficult in resource-poor settings like
the Niger Delta region of Nigeria.
Our study was designed to establish baseline indices of wound infection at the
tertiary institution, Oghara Teaching Hospital, Oghara, by looking at the
prevalent micro-organisms involved in wound infections, associated factors and
drug resistance patterns.
Pus
Suppuration, the formation of pus, is a common sequel of acute inflammation.
Pus consists of living, dead and disintegrated neutrophils, living and dead
microorganisms and the debris of tissue cells, all suspended in the
inflammatory exudates. An abscess is a localized or discrete focus of pus.
However, pus may occur diffusely in loose tissues or body cavities.
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Bacterial infection is the usual cause of suppuration and such bacteria are
said to be pyogenic (pus forming) and include Staphylococcus aureus,
Streptococcus pyogenes,Pseudomonas aeruginosa,Proteusspecies,Escherichia
coli,Klebsiella species,Clostridium perfringes, Bacteroides among others.
Pyogenic infections are either polymicrobial or monomicrobial and they maybe
endogenous or exogenous. Pyogenic infections occur in abscesses, chronic
wounds from diabetic patients, decubitus ulcer or bed sores, burns wound
infections, post-operative wound infections, cellulitis, bites, suppurative
lymphadenitis, exudates from body cavities and pyomyositis.
Various studies across the globe have been consistent enough to show a
predictable bacterial profile in pyogenic wound infections. This makes an
important observation for a clinician who intends to start empirical treatment
to his patients, while laboratory cultures reports are awaited.
A study on aerobic bacterial profile and antimicrobial susceptibility pattern of
pus isolates in a South Indian tertiary care hospital revealedStaphylococcus
aureus(24.29%) was the most common isolates, followed byPseudomonas
aeruginosa(21.49%),Escherichia coli(14.02%),Klebsiella pneumonia(12.15%),
Streptococcus pyogenes (11.23%),Staphylococcus epidermidis (9.35%) and
Proteusspecies (7.47%) (Raoet al.,2014). Another study on isolation of
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different types of bacteria from pus revealed alsoStaphylococcus aureusto be
the predominant microorganism (40%) followed byKlebsiellaspecies (33%),
Pseudomonasspecies (18%),
Escherichia coli(16%), and
Proteusspecies (7%)
(Verma 2012).
A study done in a University teaching hospital in Nigeria, revealed
Staphylococcus aureus(42.3%),Pseudomonas aeruginosa(32.9%),Escherichia
coli(12.8%) andProteus mirabilis(12.8%) are associated with surgical wound
infections (Nwachukwuet al.,2009). These findings agree with those reported
in Kenya on surgical site infections, thatStaphylococcus aureuswas the most
prevalent bacterial isolate (Dindaet al.,2013). These findings also agree with a
study done in Uganda that identifiedStaphylococcus aureusas the commonest
causative agent of septic post-operative wounds (Anguzuet al.,2007).
A study done on the bacteriology of surgical site infections in Karachi , revealed
the most common pathogen isolate wasStaphylococcus aureus (50.32%),
followed by Pseudomonas aeruginosa (16.33%),Escherichia coli(14.37%),
Klebsiella pneumonia (11.76%), Streptococcus pyogenes (1.30%), and
miscellaneous gram negative rods (5.88%) includingAcinetobacter baumannii,
Proteus mirabilisandCitrobacter diversus(Mahmood 2010). A cross-sectional
study designed to determine the distribution of the bacterial pathogens and
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their antimicrobial susceptibility from suspected cases of post-operative wound
infections, also revealedStaphylococcus aureus(63%) was the most frequently
isolated pathogenic bacteria, followed byEscherichia coli(12%),Pseudomonas
species (9.5%),Klebsiellaspecies (5%),Proteusspecies (3.5%) and coagulase
negativeStaphylococcusspecies (3.5%) (Shriyanet al.,2010).
Various studies across the globe have been consistent enough to show a
predictable bacterial profile in pyogenic wound infections. This makes an
important observation for a clinician who intends to start empirical treatment
to his patients, while laboratory cultures reports are awaited.
A study on aerobic bacterial profile and antimicrobial susceptibility pattern of
pus isolates in a South Indian tertiary care hospital revealedStaphylococcus
aureus(24.29%) was the most common isolates, followed byPseudomonas
aeruginosa(21.49%),Escherichia coli(14.02%),Klebsiella pneumonia(12.15%),
Streptococcus pyogenes (11.23%),Staphylococcus epidermidis (9.35%) and
Proteusspecies (7.47%) (Raoet al.,2014). Another study on isolation of
different types of bacteria from pus revealed alsoStaphylococcus aureusto be
the predominant microorganism (40%) followed byKlebsiellaspecies (33%),
Pseudomonasspecies (18%),Escherichia coli(16%), andProteusspecies (7%)
(Verma 2012).
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A study done in a University teaching hospital in Nigeria, revealed
Staphylococcus aureus(42.3%),Pseudomonas aeruginosa(32.9%),Escherichia
coli(12.8%) andProteus mirabilis(12.8%) are associated with surgical wound
infections (Nwachukwuet al.,2009). These findings agree with those reported
in Kenya on surgical site infections, thatStaphylococcus aureuswas the most
prevalent bacterial isolate (Dindaet al.,2013). These findings also agree with a
study done in Uganda that identifiedStaphylococcus aureusas the commonest
causative agent of septic post-operative wounds (Anguzuet al.,2007).
A study done on the bacteriology of surgical site infections in Karachi , revealed
the most common pathogen isolate wasStaphylococcus aureus (50.32%),
followed by Pseudomonas aeruginosa (16.33%),Escherichia coli(14.37%),
Klebsiella pneumonia (11.76%), Streptococcus pyogenes (1.30%), and
miscellaneous gram negative rods (5.88%) includingAcinetobacter baumannii,
Proteus mirabilisandCitrobacter diversus(Mahmood 2010). A cross-sectional
study designed to determine the distribution of the bacterial pathogens and
their antimicrobial susceptibility from suspected cases of post-operative wound
infections, also revealedStaphylococcus aureus(63%) was the most frequently
isolated pathogenic bacteria, followed byEscherichia coli(12%),Pseudomonas
species (9.5%),Klebsiellaspecies (5%),Proteusspecies (3.5%) and coagulase
negativeStaphylococcusspecies (3.5%) (Shriyanet al.,2010).
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A study on microbiological profile of diabetic foot ulcers and its antibiotic
susceptibility pattern in a teaching hospital in Gujarat, revealed that
Pseudomonas aeruginosa(27%) was the most common isolate causing diabetic
foot infections followed byKlebsiellaspecies (22%),Escherichia coli(19%),
Staphylococcus aureus (17%), Proteus species (7%), Enterococci (3%),
Acinetobacter(2%), CoNS (2%) andProvidencia(1%) (Mehtaet al.,2014). The
predominance of gram negative bacilli in diabetic pus has also been reported in
another study (Sivakumariet al.,2009). However,Staphylococcalspecies was
the primary pathogen in most of wound infections of diabetic patients (Daniel
et al.,2013).
A study done in a tertiary hospital, Pakistan on burn wounds, revealed
Staphylococcus aureus(57.98%) to be the most causative organism in burn
wound infections followed byPseudomonas aeruginosa(19.33%),Klebsiella
pneumonia(8.4%),Proteusspecies (4.2%),Staphylococcus epidermidis(3.36%),
Escherichia coliandEnterobacter(2.52%) each,CitrobacterandSerratia(0.84%)
each (Ahmedet al.,2013). Though a study done in Ibadan, Nigeria on burn
wound infections revealedKlebsiellaspecies to be the most commonly isolated
pathogen, constituting 34.4%, closely followed byPseudomonas aeruginosa
(29.0%) andStaphylococcus aureus(26.8%) (Kehindeet al.,2004).
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In a two year period study done on bacterial profile of burn wounds infections
at a burn unit Nishtar hospital Multan, the frequency of gram negative
organisms was found to be high withPseudomonas aeruginosa(54.4%) being
the most common isolate, followed byStaphylococcus aureus(22%),Klebsiella
species (8.88%),Staphylococcus epidermidis (5.79%),Acinetobacterspecies
(4.63%),Proteusspecies (2.70%) andEscherichia coli(1.54%) (Shahzadet al.,
2012).
A three year review of bacteriological profile and antibiogram on burn wounds
isolates in Van,Turkey revealed the most frequent bacterial isolate was
Acinetobacter baumannii(23.6%), followed by coagulase negativeStaphylococci
(13.6%),Pseudomonas aeruginosa (12%),Staphylococcus aureus (11.2%),
Escherichia coli(10%),Enterococcusspecies (8.8%) andKlebsiella pneumonia
(7.2%) (Bayram et al.,2013). Even though gram negative bacteria are being
increased significantly but stillStaphylococcus aureusis being continued as a
major etiological agent of pyogenic infections.
Antimicrobial resistance
The prevalence of antimicrobial resistance varies greatly between and within
countries and different pathogens. Also antimicrobial resistance patterns of
bacteria isolates keep changing and evolving with time and place.
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Microbial prevalence and antimicrobial susceptibility of wound isolates from tertiary institution
Data from the past several years show an increasing resistance to ampicillin,
penicillin and amoxicillin which were considered first line drugs for treatment
of pyogenic infections ( Anguzuet al.,2007, Shriyanet al.,2010, Binduet al.,
2014).
A study on prevalence and antimicrobial susceptibility of bacteria isolated from
skin and wound infections revealed gram positive cocci were highly sensitive to
vancomycin, teicoplanin, linezolid and chloramphenicol and gram negative
bacilli showed high degree of sensitivity to imipenem, piperacillin/tazobactam
and aminoglycosides. The least sensitivity was exhibited for penicillin,
ampicillin, tetracycline, cotrimoxazole and cephalosporins (Kaupet al.,2014).
Gram positive isolates in pus were most susceptible to vancomycin,
levofloxacin, oxacillicin and clindamycin whereas among the gram negative
isolates in pus, the most susceptible drugs were piperacillin/tazobactam,
levofloxacin, imipenem, aztreonam and amikacin (Raoet al.,2014).
Raoet al., 2013, reported that out of 144 aerobic isolates from pus samples in
post-operative wound infections 94.4% were sensitive to imipenem, 75.5% to
amikacin, 27% to ciprofloxacin, 22.2% to gentamicin, 21.5% to cotrimoxazole,
12.5% to cefotaxime, 9.7% to ceftazidime and 6.25% to amoxicillin/clavulanic
acid. All isolates were resistant to ampicillin. 33% ofStaphylococcus aureus
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were sensitive to methicillin and among the CoNS, 58.3% were sensitive
methicillin. All gram positive cocci isolated were sensitive to vancomycin and all
gram negative isolates were sensitive to imipenem (Raoet al., 2013).
S.aureusisolates showed the highest resistance to penicillin (100%), ampicillin
(95.5%), ceftriaxone (81.8%), vancomycin (65.2%) while the least resistance was
exhibited to amoxicillin/clavulanic acid (30.3%).Klebsiellaspp were resistant to
gentamicin (100%), chloramphenicol(87.5%), ceftriaxone (87.5%) and
ciprofloxacin (62.5%).E.colispp were resistant to ampicillin (100%), gentamicin
(46.7%), chloramphenicol(40%), ceftriaxone (40%) and ciprofloxacin (40%).
Proteus spp were resistant to ampicillin(100%), chloramphenicol(66.7%),
gentamicin (33.3%) and ceftriaxone (33.3%).Pseudomonasspp were resistant
to gentamicin (50%), chloramphenicol (100%), amoxicillin/clavulanic acid
(100%), ampicillin (100%) and ceftriaxone (100%). Allproteus and
pseudomonasisolates were susceptible to ciprofloxacin. Isolates of CoNS
showed 100% resistance to vancomycin, ceftriaxone, ampicillin and penicillin
but sensitive to chloramphenicol. Single and multiple antimicrobial resistances
were observed in 6.8% and 93.2% of the isolates, respectively. No bacterial
isolates was found to be sensitive to all antibiotics tested ( Dessalegnet al.,
2014). Aminoglycosides and quinolones were found to be the most susceptible
drugs in aerobic bacterial isolates from wound infections (Al-azawi, 2013,
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Anguzu et al.,2007). Sensitivity ofS.aureus isolates from burn wound
infections at a hospital in Ethiopia were 93.9% vancomycin, 90.9%
clindamycin, 86.4% kanamycin and 86.4% erythromycin. Resistance of
S.aureusisolates above 50% rates was observed in penicillin, methicillin,
polymyxin B and chloramphenicol 95.5%, 77.3%, 68.2% and 51.5%
respectively (Tigistet al., 2012).
Acinetobacterisolates showed almost complete resistance to cephalosporins
(cephalexin 98.7%, cefuroxime 98.2%, cefotaxime 93.2%, ceftriaxone 93.3%,
ceftazidime 87.5%, cefaclor 97.4%), piperacillin ( 94.7%), gentamicin (81.3%),
while lower rates of resistance were shown in amikacin 68.3% and ciprofloxacin
69.7%. The most effective antimicrobial drug was doxycycline with the lowest
resistance rate of 22.1% (Elmanama 2006).
Azithromycin , gatifloxacin, amikacin, ampi/subbuctam and ciprofloxacin were
found to be highly susceptible to gram negative organisms in pus while
amikacin, azithromycin, ciprofloxacin, clindamycin, cloxacillin,
chloramphenicol, moxifloxacin, linezolid and gatifloxacin were highly sensitive
for gram positive organisms in pus (Vermaet al.,2012, Verma 2012). However,
most of gram negative isolates in diabetic foot ulcers were resistant to
amikacin, piperacillin/tazobactam, gentamicin, ampicillin-sulbactam and
gatifloxacin. The gram negative bacilli were highly sensitive to imipenem and
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polymyxin. 69.4% of GNB were ESBL producer. Gram positive isolates were
found to be susceptible to vancomycin, linezolid, ampicillin/sulbactam,
tetracycline and neomycin. 60% ofStaphylococcus aureuswere methicillin
resistant and were sensitive to vancomycin and linezolid (Mehtaet al.,2014).
Gram negative organisms were highly resistant to ampicillin and ceftriaxone (
lactam antibiotics). Ciprofloxacin was highly active against all gram negative
organisms and also gram positive cocci (Nwachukwu et al.,2009). 100%
vancomycin resistanceStaphylococcus aureuswas isolated from wounds of
diabetic patients (Danielet al.,2013). In that studyStaphylococcus aureusonly
showed sensitivity to gentamycin and tetracycline.
CHAPTER THREE
MATERIALS AND METHODS
Sample size
A total of 80 wound swabs submitted at the general culture bench from in-
patients in different wards of the hospital, 36 of which were male and 34
female. Inclusion criterion was patients with purulent wounds.
Clinical specimens
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Specimens were collected aseptically with sterile cotton wool swabs from post
operative wound infections. Pus samples /wound swabs were collected with
aseptic precautions and were transported to the laboratory without delay.
Blood agar, MacConkey agar and Nutrient agar were used for isolation and
study of cultural characters. The plates were incubated at 37C for 24 hours in
an incubator. Isolated colonies were subjected to Gram staining and
biochemical tests for identification. Biochemical tests are performed by API20E
and Vitek2 systems. Most resistant isolate is further identified by 16S rRNA
sequencing.
Culture of specimen
The specimens were inoculated on blood, chocolate and MacConkey agar plates
(Oxoid, Basingstoke, U.K). The plates were incubated aerobically at 37 0C for
24 to 48 hours. Pure colonies were kept in nutrient agar slants. The nutrient
agar slants were incubated at 37oC for 18 24 h before storage in the
refrigerator at 4C pending biochemical analysis.
Identification of bacterial pathogens
Pure cultures were characterized using morphological appearances on selective
and differential media. Motility test and biochemical tests such as catalase,
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coagulase, oxidase, Voges Proskauer, hydrogen sulphide production, urease,
methyl red, indole, citrate and sugar utilization tests were carried out according
to standard techniques.
Antibiotics
A total of eight (8) antibiotics, which represent the most commonly prescribed
antibiotics for treatment of wound infections in the study area, were used in
the study. Oxoid antibiotic discs used were amoxicillin (AMX, 10 g),
ceftriaxone (CRO, 30 g), ceftazidime (CAZ, 30 g), ciprofloxacin (CIP, 10 g)
and gentamicin (CN, 30 g).
Antibiotic susceptibility testing
Antimicrobial susceptibility test were carried out on isolated and identified
colonies of Gram-negative bacteria using commercially prepared antibiotic disk
(Span diagnostics) on Nutrient agar plates by the disk diffusion method,
according to the Central Laboratory Standards Institute (CLSI) guidelines.
Antibiotics used in our study were Gentamycin (GEN), Angmenycin (AUG),
Cefitriazonc (CTR), Erythromycin (ERY), Cefixime (CEF), Imepenem (IMP),
Meroparem (MEM) and Lavofloxacin (LEV).
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CHAPTER FOUR
RESULT PRESENTATION
During the study, 80 wounds swabs were collected and cultured on Mac-
conkey, Blood and Chocolate Agar, this was after they were incubated at 37o
for 24hrs, thus in the wound sample fifty (50) out of eighty (80) samples have
growth which amounted to 62.5% growth rate. The bacterial isolate recorded in
this study areKlebsiellapnemoniae, Psuedomonas aeruginosa,Escherichia
coli,Staphylococcusaureus,ProteusvulgarisandProteusMirabilisas shown in
table 4.1. Gram negative bacilli were responsible for 70% of wound
infections.Staphylococcus aureuswas the only gram positive organism
isolated.Staphylococcus aureuswas the most prevalent pathogen detected in
the swabs, whilePseudomonasaeruginosa,Proteusmirabilisandvulgariswas
the least detected isolate.
Based on cultural, morphological and biochemical characteristics of the
organisms isolated, a total of six (6) bacterial species isolates were identified in
the 50 wounds swabs samples studied.Escherichia coli(20%),ProteusMirabilis
(10%),ProteusVulgaris(10%),Pseudomonasaeruginosa(10%), ,Klebsiella
pneumonia(20%) andStaphylococcusaureus(30%). Thus, table 2 above shows
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Microbial prevalence and antimicrobial susceptibility of wound isolates from tertiary institution
that staphyloccous aureus is more prevalent in the studied wound swab and
this is closely followed byEscherichia coliandKlebsiellapneumonia.
The bacterial isolates exhibited a high resistance to the antibiotics tested,
with the organismsKlebsiellapneumoniaandStaphylococcusaureusresistant
to all of the tested antibiotics in the same vein majority of the organisms
isolated with the exception of two (2) strains ofProteusmirabiliswere all
resistant to cefixime (Table3).Pseudomonas aeruginosaexhibited a very high
resistance to the tested antibiotics with few sensitivity observed in meropenem,
imepenem, levofloxacin and cefixime, the lowest recorded resistance being
E.coliwas relatively susceptible to gentamycin, meropenem, imepenem,
levofloxacin, and Augmentin.
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3705182/table/T5/http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3705182/table/T5/7/25/2019 Ese Project
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Microbial prevalence and antimicrobial susceptibility of wound isolates from tertiary institution
4.1 Microscopic and Morphological characteristics of the isolates
Table 4.1.Identification chart for bacteria isolates
Cultural
xteristics
Morp. Gram
stain
Catalase Oxidase Citrate indole Motility H2S Urease Lactose Coaulase
test
!erobic
test
"athoen
identified
Creamy round
mucoid
irreular colony
#od $ % & % & $ & % % % + Kleb
Pneumon
Cream flat
colony 'ith
undulatin ede
#od $ $ % % $ % $ $ $ $ % Pseudom
aerogino
"in( colony
'ith oriod
shape
#od $ % $ $ % % $ % % % + Escheric
.coli
Cream mucoid
colony 'ith
irreular ede
#od $ % & & & % % % $ % + Proteus
Mirabilis
)hite and flat
colonies
#od $ % $ $ % % % % $ $ + Proteus
Vulgaris
Creamy on
blood aar
Cocci
in
cluste
rs
% % $ $ % % $ % % % +
Staphylo
s aureus
*ey $+ neati,e to the test-%+ "ositi,e
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Microbial prevalence and antimicrobial susceptibility of wound isolates from tertiary institution
Table 4. 2: "re,alence of indi,idual bacterial isolates from 'ound infection
Oranism o /0
Staphylococcus aureus 12/30
Escherichia coli 1/50
Proteus mirabilis 2/10
Proteus vulgaris 2/16
Klebsiella pneumonia 1/50
Pseudomonas aeruginosa 2/10
Total 2/1
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Table 4.3: Antibiotic sensitivity/resistance of the isolated organism
Isolates CTR GEN MEM IMP CEF ERY LEV AUG
Pseudomonas
aeruginosa
R R S S R R S S
Klebsiella
Pneumonia
R R R R R R R R
Proteus
vulgaris
R R R R S S S S
Proteus
mirabilis
S S R R S S S S
Staph. aureus R R R R R R R R
E coli R S S S R R S S
Key:
GEN = Gentamycin
AUG = Augmentin
MEM = Meropenem
IMP = Imepenem
CEF = Cefixime
ERY = Erythromycin
LEV = Levofloxacin
CTR = Cefixime
CHAPTER FIVE
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DISCUSSION AND CONCLUSION
Discussion of result
Epidemiological surveillance of antimicrobial resistance is indispensable
for empirical treatment infections, implementing control measures, and
preventing the spread of antimicrobial resistant microorganisms (Goosens and
Sprenger, 1998). Also Bacterial contamination of wounds is a serious problem
in the hospital, where the site of a sterile operation can become contaminated
and subsequently infected.
Our study demonstrated a high prevalence (62.5%) of pathogenic
bacteria in wounds. This high figure is consistent with that obtained in similar
studies in Nigeria as rightly reported by Wariso and Nwachukwu, (2003) and
Taiwo et al., (2001), but different from another study in East Africa reporting a
prevalence of 70.5% as authored by Mulugeta and Bayeh, (2011). These
differences may be due to study design. The rates might be equally high if only
wounds with a high suspicion of infection are investigated as opposed to all
wounds.
Although there was no association between the type of wound and the
type of micro-organism isolated, it is important to note that all swabs from
traumatic wounds yielded significant bacterial growth and were thus deemed to
indicate infection. However, two previous studies carried out by Otokunefor et
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Microbial prevalence and antimicrobial susceptibility of wound isolates from tertiary institution
al., (1990) and Okesola and Kehinde, (2008) associated specific micro-
organisms with particular wound types]. More studies are required to clarify
this observation.
As in previous studies, documented by Taiwo, et al., (2002) and Egbe et
al., (2011); Gram-negative bacteria were the most commonly isolated
pathogens. Our observation ofStaphylococcus aureusas the most common
pathogen in wound infections is in-line with the works of other authority in
Nigeria who reported thatStaphylococcus aureusis the predominant organism
isolated from wound (Egbe et al., 2011).Klebsiella pneumoniaewhich in our
study amounted for 20% of the prevalence rate of organism isolated from
wounds was observed as the most common pathogen in wounds in a study
conducted by Sule et al., (2002). This is evidence of the existence of local and
regional variability and shows that each health facility has to determine the
prevalent micro-organisms and other associated indices. Most of these studies,
including ours, are limited by the fact that anaerobic cultures were not done for
a variety of reasons, the main one being a lack of equipment and funds. Thus,
anaerobic bacteria, which are also important in wound infections, could not be
isolated.
Pathogenic isolates of have relatively high potentials for developing
resistance (Karlowsky etal., 2004). High resistance of organism to
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Microbial prevalence and antimicrobial susceptibility of wound isolates from tertiary institution
antimicrobial agents tested was observed in this study withStaphylococcus
aureusandKlebsiellapneumoniaresistant to all the tested antibiotic.
Staphylococcus aureusis a very common cause of infection thus it is not
surprise to have isolated it in wounds.
Staphylococcus aureuswas found to be a frequent isolate in wound sepsis
(Emmerson, 1994). A study by Ndip,et al., (1997) at Ilorin, Nigeria reported
wound infections of 38% as the highest frequency of S. aureus isolates. This
agrees with the result in the present study where S. aureus also had the
highest isolate of 30%.Staphylococcus aureusdevelops resistance very quickly
and successfully to different antimicrobials over a period of time. The highest
frequency of S.aureusoccurred with susceptibility to antimicrobial agent
Levofloxacin.
The high level resistance could be associated with earlier exposure of
these drugs to isolates which may have enhanced development of resistance.
There is high level antibiotic abuse in this environment arising from self-
medication which is often associated with inadequate dosage and failure to
comply to treatment, and availability of antibiotics to consumers across the
counters with or without prescription. It had been observed that the
indiscriminate use of antibiotics without prescriptions in the developing
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Microbial prevalence and antimicrobial susceptibility of wound isolates from tertiary institution
countries such as Nigeria where there are no regulatory policies in this respect
has rendered the commonly used antibiotics completely ineffective in the
treatment ofStaphylococcus aureusinfections.
This is similar to what was observed by Aibinuetal.,(2004) who reported
100% resistance of their E. coliisolates to ampicillin and amoxicillin.
Resistance to Augmentin and levofloxaxin observed in this study was similar to
what was observed in South Africa, Israel, (62% - 84%) and Hong Kong,
Philippines (64 - 82%) (Stellinget al., 2005). Densenclosetal. (1988)) reported
53% of theirE. coliisolates were resistant cotrimoxazole and 67% to
tetracycline. Their finding is in harmony with the report of this study, showing
69% and 88% resistance to antibiotics agents. The reason for this high
resistance to commonly used antibiotics may be due to widespread and
indiscriminate use in our environment.
Antibiotic resistance by the isolates to commonly prescribed antibiotics
was high. This high level of resistance is a cause for concern. The absolute
resistance to levofloxacin was not unexpected considering the fact that
levofloxacin is a component of Ampiclox, an antibiotic frequently implicated in
self-medication in Nigeria (Yah et al., 2008). Augmentin, which are among the
least prescribed antibiotics in Nigeria (Yah et al., 2008), are neither widely
abused in this country nor easily affordable by the patients in the Niger Delta
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Microbial prevalence and antimicrobial susceptibility of wound isolates from tertiary institution
region. The development of resistance to Augmentin observed in this study is
thus a wake-up call for action on antimicrobial resistance. The poor availability
of antibiotics, as well as their unregulated use and misuse, has been shown to
contribute to increasing antimicrobial resistance in developing countries (Hart
and Kariuki, 1998). The lack of diagnostic facilities in these developing regions
encourages empiric treatment and overtreatment, which contribute to the
increased resistance (Hart and Kariuki, 1998).
Conclusion
Severe antimicrobial resistance in wound infections was observed among
patients in Delta State University Teaching Hospital (DELSUTH), Oghara, Delta
State of Nigeria. There is a need for serious and urgent intervention to stem the
spread and further evolution of this resistance. A rigorous infection control
policy combined with rational drug use play an important role in this fight
against antimicrobial resistance..
Recommendations
Multiple antibiotic resistance in bacterial populations is a great challenge
in the effective management of wound infections. This calls for monitoring and
optimization of antimicrobial use. We suggest a multidisciplinary approach to
wound infection management involving both clinicians and microbiologists.
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Strengthening of laboratory services at local and national levels will ensure
effective surveillance of antimicrobial resistance. We also advocate routine
microbiological surveillance of wounds and testing for antimicrobial
susceptibility before drug use. Thus the inclusion of anaerobic culture in
routine microbiology culture investigations will be of immense contribution
also. Finally, Since antimicrobial resistant patterns are constantly evolving, and
present global public health problem, there is the necessity for constant
antimicrobial sensitivity surveillance. This will help clinicians provide safe and
effective empiric therapies.
Recommended