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Abscesses and Deep-Seated Wound Infections

1.0.Types of specimens:
Abscess pus Post-operative wound swab
Abscess swab Deep-seated pus swab
Wound exudate

2.0. SCOPE OF DOCUMENT

This SOP describes the processing and bacteriological investigation of specimens
from abscesses and from post-operative wound and deep-seated infections.
3.0. INTRODUCTION

3.1.0. Abscesses are accumulations of pus in the tissues. Any organism isolated
may be significant. Abscesses occur in many parts of the body as superficial
infections or as deep-seated infections associated with any internal organ.

3.1.1.0 BRAIN ABSCESS

Brain abscesses are serious and life-threatening.
3.1.1.1 Sources of abscess formation include:
1. direct contiguous spread from chronic otitic or paranasal
sinus infection
2. metastatic haematogenous spread either from general
sepsis or secondary to chronic suppurative lung disease
3. penetrating wounds
4. surgery
5. cryptogenic (i.e. source unknown)

3.1.1.2 Bacteria isolated from brain abscesses. The most commonly

isolated organisms include:
1. anaerobic streptococci
2. anaerobic Gram-negative bacilli
3. "Streptococcus anginosus" group
4. Enterobacteriaceae
5. Streptococcus pneumoniae
6. β-haemolytic streptococci
7. S. aureus

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Any organism isolated from a brain abscess must be regarded as clinically
significant.Organisms causing brain abscesses following trauma may often be
environmental in origin, such as Clostridium species, or skin derived, such as
staphylococci and Propionibacterium species.
Brain abscesses due to fungi are rare.

3.2.0 BREAST ABSCESS

Breast abscesses infections are commonly caused by S. aureus, but may also be
polymicrobial, involving anaerobes and streptococci. Signs include discharge from
the nipple, swelling, edema, firmness and erythema.
Abscesses may also be caused by Pseudomonas aeruginosa and Proteus species.

3.3.0CARBUNCLES, FURUNCLES, CUTANEOUS AND SOFT TISSUE

ABSCESSES
Common bacterial isolates include:
1. Oral streptococci
2. Streptococcus anginosus group
3. Fusobacterium nucleatum
4. Prevotella species
5. Porphyromonas species
6. Staphylococcus aureus
7. Clostridium species

3.3.1 Soft tissue abscesses they may arise from animal bites in which case common
isolates include Pasteurella and Actinobacillus species as well as other organisms of
the HACEK group (Haemophilus, Actinobacillus, Cardiobacterium, Eikenella and
Kingella species).
Burkholderia pseudomallei causes melioidosis.
Pyomyositis is a purulent infection of skeletal muscle, organism is S. aureus.

DENTAL ABSCESS
3.3.2 Dental abscesses - microorganisms colonizing the teeth may also become
responsible for oral and dental infections, leading to dentoalveolar abscesses and
associated diseases.
3.3.3 Periodontal disease involves the gingiva and underlying connective tissue, and
infection may result in gingivitis or periodontitis.
3.3.3.1 Organisms most commonly isolated in acute dentoalveolar abscesses are
facultative or strict anaerobes. Strict anaerobes predominate. Examples include:
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1. α-haemolytic streptococci
2. anaerobic Gram-negative bacilli
3. anaerobic streptococci
4. "S. anginosus" group
5. Actinobacillus actinomycetemcomitans
6. spirochaetes
7. Actinomyces species

3.4 LIVER ABSCESS

Liver abscesses can be amoebic or bacterial (so-called pyogenic) in origin or, more
rarely, a combination of the two.
3.4.1 Pyogenic liver abscesses The most common bacteria include:
1. Enterobacteriaceae
2. Bacteroides species
3. Clostridium species
4. anaerobic streptococci
5. "S. anginosus" group
6. enterococci
7. P. aeruginosa
8. B. pseudomallei (in endemic areas)

3.4.2 Hepatic candidiasis is an increasing problem in immunocompromised

patients
3.4.3 Amoebic liver abscesses arise as a result of the spread of Entamoeba
histolytica via the portal vein from the large bowel which is the primary site of
infection.
3.4.4 Hydatid cysts may also occur as fluid-filled lesions in the liver.

3.5.0 LUNG ABSCESS

Lung abscesses involve the destruction of lung usually after necrotizing pneumonia.
Pneumonia caused by S. aureus and Klebsiella pneumoniae may show this picture.
other organisms may also occur.
Nocardia infection is most often seen in the lung where it may produce an acute,
often necrotizing, pneumonia.

3.6.0 PANCREATIC ABSCESS

Pancreatic abscesses are potential complications of acute pancreatitis. Infections are
often polymicrobial and common isolates include Escherichia coli, other
Enterobacteriaceae, enterococci and anaerobes.
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3.7.0 PERIRECTAL ABSCESS

Perirectal abscesses are encountered in patients with predisposing factors. These
include:
1. immunodeficiency
2. malignancy, particularly cancer patients
3. rectal surgery
4. ulcerative colitis
They are often caused by:
1. anaerobes
2. Enterobacteriaceae
3. streptococci
4. S. aureus

3.7.0 PILONIDAL ABSCESS

Pilonidal abscesses are common in children and result from infection of a pilonidal
sinus. Anaerobes and Enterobacteriaceae are usually isolated, but they may be
caused by S.aureus and β-haemolytic streptococci.
3.8.0 PROSTATIC ABSCESS
3.8.1 Prostatic abscesses may be caused by, or associated with:
1. acute and chronic prostatitis
2. instrumentation of the urethra and bladder
3. lower urinary tract obstruction
4. hematogenous spread of infection

3.8.2 Organisms that may cause prostatic abscesses include:

1. E. coli and other Enterobacteriaceae
2. anaerobes
3. Neisseria gonorrhoeae
4. S. aureus

3.9.0 PSOAS ABSCESS

3.9.1 Psoas abscesses occur secondarily to:
•Appendicitis diverticulitis osteomyelitis of the spine
•infection of a disc space bacteraemia perinephric abscess
3.9.2 Psoas abscesses are predominantly caused by:
1. Enterobacteriaceae
2. Bacteroides species
3. S. aureus
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4. streptococci
5. Mycobacterium species

3.9.0 RENAL ABSCESS

3.9.1 Renal abscesses are typically caused by Gram-negative bacilli and result
from ascending urinary tract infection, pyelonephritis or septicemia. S. aureus has
been reported infecting the renal cortex as a consequence of haematogenous
spread from a site elsewhere in the body.
3.9.2 Perinephric abscesses are relatively uncommon but serious extensions of
renal abscesses. Causative organisms are the same as those for renal abscesses.

3.10.0 SCALP ABSCESS

3.10.1 Scalp abscesses are a recognized complication of electronic monitoring with
scalp electrodes. Polymicrobial infections also occur, involving:
1. anaerobes
2. β-haemolytic streptococci
3. S. aureus
4. Enterobacteriaceae
5. enterococci
6. coagulase-negative staphylococci
3.10.2 Kerion is a pustular folliculitis of adjacent hair follicles, creating dense
inflamed areas of the scalp, usually caused by dermatophytes. Secondary bacterial
infection may occur.
3.11.0 SPINAL EPIDURAL ABSCESS
3.11.1 Spinal epidural abscesses may occur in patients with:
The most common isolate is S. aureus. Staphylococcus epidermidis may be
isolated in patients following invasive spinal manipulation. Streptococci (α-
haemolytic, β-haemolytic and S. pneumoniae), Enterobacteriaceae and
pseudomonads may also be isolated.

3.12.0 UNUSUAL CASES OF ABSCESS FORMATION

Unusual cases of abscess formation can occur in patients with many underlying
conditions and may be caused by a vast range of organisms. Any organism isolated
from abscess pus is potentially significant.
Actinomycosis is a chronic suppurative infection characterized by chronic abscess
"Sulphur granules" are sought in the pus specimen. These are discharged from
actinomycosis abscesses. Sulphur granules are colonies of organisms forming a
filamentous inner mass which is surrounded by host reaction. They are formed only
in vivo. They are hard, buff to yellow in color and have a clubbed surface.
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3.12.0 POST-OPERATIVE WOUND INFECTIONS
Postoperative wound infections - arise when microorganisms contaminate surgical
wounds during an operation or immediately afterwards. Colonized body sites are
frequent sources of pathogens although they may be transmitted via medical and
nursing staff or via inanimate objects from other patients or elsewhere in the hospital
environment.
3.12.1 Organisms most commonly isolated include,
1. S. aureus including MRSA
2. Bacteroides species
3. Clostridium species
4. Enterobacteriaceae
5. pseudomonads
6. β-haemolytic streptococci
7. enterococci
8. Peptostreptococcus species

9. Coagulase-negative staphylococci and coryneforms isolated from post-

operative sites overlying implants or prostheses may indicate infection.
3.12.2 Progressive bacterial synergistic gangrene usually follows as a post-
operative complication of abdominal or thoracic surgery. It results from
infection with a symbiotic mixture of aerobic and anaerobic organisms such as
S. aureus and anaerobic streptococci.
3.12.3 Infectious gangrene (gangrenous cellulitis) is a rare disease in which
bullous necrosis of the skin and underlying tissues may be associated with
bacteremia. There are several forms of infectious gangrene, differentiated by the
causative organism, anatomical site and predisposing conditions. These include
necrotizing fasciitis and gas gangrene (clostridial myonecrosis).
3.12.4 Gas gangrene is caused by clostridia, in particular Clostridium
perfringens. It’s associated with clinical shock, leakage of serosanguinous fluid,
tissue necrosis and presence of gas in the tissues.
3.12.5 Fournier’s gangrene is a form of necrotizing fasciitis affecting the male
genitalia: usually confined to the scrotum but it may spread to involve the
perineum, penis and abdominal wall.
Necrotizing fasciitis can occur post-operatively and is a serious, infrequently
occurring infection primarily affecting the subcutaneous fat and superficial
fascia of muscles and often the overlying soft tissues. It is caused by anaerobic
organisms , streptococci and others.

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3.13.0 INTRA-ABDOMINAL SEPSIS
Intra-abdominal sepsis is infection occurring in the normally sterile peritoneal cavity.
The term covers primary and secondary peritonitis, as well as intra-abdominal
abscesses.

4.0 PRACTICAL PROCEDURES

4.1.0 Safety Considerations
4.1.1 Collection Specimen
Avoid accidental injury when pus is aspirated
4.1.2 Specimen Transport and Storage
Pus in sterile leakproof container in a sealed plastic bag
4.1.3 Specimen Processing
4.1.3.1 Should be performed in a microbiological safety cabinet
4.1.3.2 Prior to staining, fix smeared material by placing the slide on
an electric hotplate (65 to 75 °C), under the hood, until dry. Then
place in a rack or other suitable holder
NOTE: Heat-fixing may not kill all Mycobacterium species. Slides
should be handled carefully
Centrifugation must be carried out in sealed buckets, which are
subsequently opened in a microbiological safety cabinet
Specimen containers must also be placed in a suitable holder
Laboratory procedures that give rise to infectious aerosols
must be conducted in a microbiological safety cabinet

4.2.0 Specimen Collection

4.2.1 Optimal Time of Specimen Collection
Before antimicrobial therapy where possible

4.2.2 Correct Specimen Type and Method of Collection

The specimen will usually be collected by a medical practitioner
4.2.3 Samples of pus are preferred to swabs. However, pus swabs are
often received. If swabs are used, sample the deepest part of the wound
trying to avoid the superficial microflora

4.2.4 Adequate Quantity and Appropriate Number of Specimens

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Ideally, a minimum volume of 1 ml of pus Swabs should be well soaked
in pus

5.0 Specimen Transport and Storage

5.1 Time between Specimen Collection and Processing

Specimens should be transported and processed as soon as possible
The volume of specimen influences the transport time. Large volumes
of purulent material maintain the viability of anaerobes for longer
The recovery of anaerobes is compromised if the transport time exceeds
3h

5.2 Special Considerations to Minimize Deterioration

Swabs should be transported in Amies transport medium with charcoal
If processing is delayed, refrigeration is preferable to storage at ambient
temperature. Delays of over 48 h are undesirable

6.0 Specimen Processing

6.1 Test Selection

Divide specimen on receipt for appropriate procedures such as
examination for parasites and culture for Mycobacterium species,
depending on clinical details

6.2 APPEARANCE
Describe presence or absence of sulphur granules (if sought)

6.3 MICROSCOPY
6.3.1 Standard
6.3.1.1 Swab
Prepare a thin smear on a clean microscope slide for Gram staining
after performing culture.
6.3.1.2Pus
Using a sterile pipette place one drop of neat specimen or
centrifuged deposit, as applicable, on to a clean microscope slide
Spread this using a sterile loop to make a thin smear for Gram
staining

6.4 Supplementary
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6.4.1 Gram stain of sulphur granules
With care, either squash the washed sulphur granule between
two slides using gentle pressure or use the homogenized
granules and make a thin smear for Gram staining.
NOTE: Any grinding of sulphur granules should be performed in
a microbiological safety cabinet
Microscopy for fungi, Mycobacterium species and parasites

7.0 Culture and Investigation

7.1 preparation of samples
7.1.1 Exudates
Centrifuge in a sterile, capped, conical-bottomed container at 1200x g for 5-
10 mins Transfer the supernatant with a sterile pipette, leaving
approximately 0.5 mL, to another sterile container for additional testing if
required Resuspend the deposit in the remaining fluid

7.2.2 Sulphur granules

Wash any sulphur granules that are present
Suspend an aliquot of pus containing sulphur granules in sterile water or
saline in a sterile container. Agitate gently to wash pus from the granules
Grind the washed granules in a small amount of sterile water or saline Use
this homogenized sample to make a smear for Gram staining and to inoculate
agar plates
NOTE: Any grinding of sulphur granules should be performed in a
microbiological safety cabinet
8.0 Specimen processing
8.1 Pus
Inoculate agar plates and enrichment broth with the pus or centrifuged
deposit with a sterile swab. Sulphur granules should be ground before
inoculated.
For the isolation of individual colonies, spread inoculum with a sterile loop.
Use aspirate for anaerobic culture (not swab)

8.2 Swabs
Inoculate each agar plate with swab.
For the isolation of individual colonies, spread inoculum with a sterile loop

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8.3 Culture media, conditions and organisms For all specimens:

Clinical Standard media Incubation Cultures Target organism(s)

details/ read
Temp Atmos Time
°C
Blood agar 35-37 5-10% 40-48h daily S. aureus
CO2 β-haemolytic strept.
Enterococci
All MacConkey agar 35-37 air 16-24h >16h Enterobacteriaceae
specimens Or CLED Pseudomonads
Anaerobic culture 35-37 anaerobic 5d ≥40h and Anaerobes
at 5d

For these special situations, add the following:

Clinical details/ Supplementary Incubation Culture Target

conditions media s read organism(s)
Tem Atmos Time
p °C
Submandibular abscess Anaerobic 35-37 anaerobi 5d ≥40h Anaerobes
Empyema culture with c and at
Normally sterile sites such as: metronidazole 5d
Brain abscess -Liver abscess 5 µg disc
Lung abscess -Psoas abscess
Spinal abscess
All pus and exudates (not anaerobic brothor 35-37 air as 40-48h N/A Any organism
swabs) blood culture 35-37 above 40-48h >40h
bottle then
subcultured where
appropriate on to
above media
(excluding CLED
Actinomycosis (or where agar)
Blood agar 35-37 anaerobi 10 d ≥ 40h, Actinomyces spp.
microscopy suggestive of supplemented c at 7d
Actinomycetes) with and 10d
metronidazole and
nalidixic acid

Nocardiosis Blood agar 35-37 air up to 7d at 3d Nocardia species

and 7d
Prostatic abscess Primary Chocolate agar 35-37 5-10% 40-48h >40h N. gonorrhoeae
peritonitis in females GN medium 35- CO
anaerobi
2 5d 48 hr Gram-negative
(NAV) 37C c and 5d anaerobes

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 KINGDOM OF SAUDI ARABIA

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Applies To: Bacteriology Staff Approved Hospital Director
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Routinely better to use both blood agar, chocolate agar, MacConkey, Sabourauds, beside
anaerobic culture for wound specimens. And to ude metronidazole 5 µg disc for anaerobic
plate.

8.4 Identification
8.4.1 Minimum level in the laboratory

Organisms Minimal level

Anaerobes Anaerobes level
CNS Coagulase-negative level
Enterobacteriaceae Coliforms level
Fungi Species level
Hemophilus Species level
Streptococci Group A, B, C & G Lancefield Group level
Moraxella Species level
Neisseria Species Level
Pseudomonads Pseudomonads level
Staph aueus Species level
Strept anginosus group Species level
Yeasts Yeast level
Mycobacterium
Parasites

Organisms may be further identified if clinically or epidemiologically indicated

8.4.2 Referral to Reference Laboratories
-Actinomyces for identification (if unable to perform locally)
-Mycobacterium culture and susceptibility testing
-Fungi culture and identification and/or susceptibility testing
-Isolates associated with outbreaks and where epidemiologically indicated

8.4.3 Antibiotic Susceptibility Testing: see AST

9.0 Reporting Procedure

9.1 Microscopy
Report on WBCs and organisms detected
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Microscopy for fungi, Mycobacterium species and parasites
9.1.1 Microscopy reporting time
Urgent microscopy results to be telephoned or sent electronically
Written report, 16 – 72 h

9.2 Culture
Report clinically significant organisms isolated or
Report other growth or
Report absence of growth
Report on the presence of sulphur granules
Also, report results of supplementary investigations

9.2.1 Culture reporting time

Clinically urgent culture results to be telephoned or sent electronically
Written report, 16 – 72 h stating, if appropriate, that a further
report will be issued Supplementary investigations: Fungi,
Mycobacterium species and parasites

9.3 ANTIBIOTIC SUSCEPTIBILITY TESTING

Report susceptibilities as clinically indicated according to protocol

10.0 REPORTING TO THE INFECTION CONTROL

According to specific organisms especially MDR or ESBLs organisms or MRSA
Attachment: ABSCESS FLOWCHART

11.0. REFERENCES
11.1. national public health services of wales –UK- SOPS
11.2. Isenberg HD; Clinical Microbiology Procedure Handbook. American Society of
Microbiology, Washington DS, 2004
11.3. Murray P.R., et al. Manual of Clinical Microbiology – 8 th edition 2003, American
Society of Microbiology, Washington DC.

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APPROVALS:

NAME TITLE SIGNTURE DATE

Prepared
& NAEEM HANNOUN Specialist
Reviewe
d by

Approved
By DR.MUNA ALATRABI Section Head

Approved DR.MUNA ALATRABI Lab. Director

By

Reviewed
By
A'RAHMAN ALYAESH TQM Director

Reviewed Medical
Dr. MOHAMMED FAHMY
By Director

Approved DR.KHALID Hospital

By ALABDULWAHAB Director

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