Anaerobic bacteria culture





Definition

An anaerobic bacteria culture is a method used to grow anaerobes from a clinical specimen. Obligate anaerobes are bacteria that can live only in the absence of oxygen. Obligate anaerobes are destroyed when exposed to the atmosphere for as briefly as 10 minutes. Some anaerobes are tolerant to small amounts of oxygen. Facultative anaerobes are those organisms that will grow with or without oxygen. The methods of obtaining specimens for anaerobic culture and the culturing procedure are performed to ensure that the organisms are protected from oxygen.


Purpose

Anaerobic bacterial cultures are performed to identify bacteria that grow only in the absence of oxygen and which may cause human infection. If overlooked or killed by exposure to oxygen, anaerobic infections result in such serious consequences as amputation , organ failure, sepsis, meningitis, and death. Culture is required to correctly identify anaerobic pathogens and institute effective antibiotic treatment.


Precautions

It is crucial that the health care provider obtain the sample for culture via aseptic technique . Anaerobes are commonly found on mucous membranes and other sites such as the vagina and oral cavity. Therefore, specimens likely to be contaminated with these organisms should not be submitted for culture (e.g., a throat or vaginal swab). Some types of specimens should always be cultured for anaerobes if an infection is suspected. These include abscesses, bites, blood, cerebrospinal fluid and exudative body fluids, deep wounds, and dead tissues. The specimen must be protected from oxygen during collection and transport and must be transported to the laboratory immediately.


Description

Anaerobes are normally found within certain areas of the body but result in serious infection when they have access to a normally sterile body fluid or deep tissue that is poorly oxygenated. Some anaerobes normally live in the crevices of the skin, in the nose, mouth, throat, intestine, and vagina. Injury to these tissues (i.e., cuts, puncture wounds, or trauma) especially at or adjacent to the mucous membranes allows anaerobes entry into otherwise sterile areas of the body and is the primary cause of anaerobic infection. A second source of anaerobic infection occurs from the introduction of spores into a normally sterile site. Spore-producing anaerobes live in the soil and water, and spores may be introduced via wounds, especially punctures. Anaerobic infections are most likely to be found in persons who are immunosuppressed, those treated recently with broad-spectrum antibiotics , and persons who have a decaying tissue injury on or near a mucous membrane, especially if the site is foul-smelling.

Some specimens from which anaerobes are likely to be isolated are:

  • blood
  • bile
  • bone marrow
  • cerebrospinal fluid
  • direct lung aspirate
  • tissue biopsy from a normally sterile site
  • fluid from a normally sterile site (like a joint)
  • dental abscess
  • abdominal or pelvic abscess
  • knife, gunshot, or surgical wound
  • severe burn

Some of the specimens that are not suitable for anaerobic cultures include:

  • coughed throat discharge (sputum)
  • rectal swab
  • nasal or throat swab
  • urethral swab
  • voided urine

Specimen collection

The keys to effective anaerobic bacteria cultures include collecting a contamination-free specimen and protecting it from oxygen exposure. Anaerobic bacteria cultures should be obtained from an appropriate site without the health care professional contaminating the sample with bacteria from the adjacent skin, mucus membrane, or tissue. Swabs should be avoided when collecting specimens for anaerobic culture because cotton fibers may be detrimental to anaerobes. Abscesses or fluids can be aspirated using a sterile syringe that is then tightly capped to prevent entry of air. Tissue samples should be placed into a degassed bag and sealed, or into a gassed out screw top vial that may contain oxygen-free prereduced culture medium and tightly capped. The specimens should be plated as rapidly as possible onto culture media that has been prepared.

Culture

Cultures should be placed in an environment that is free of oxygen, at 95°F (35°C) for at least 48 hours before the plates are examined for growth.

Gram staining is performed on the specimen at the time of culture. While infections can be caused by aerobic or anaerobic bacteria or a mixture of both, some infections have a high probability of being caused by anaerobic bacteria. These infections include brain abscesses, lung abscesses, aspiration pneumonia, and dental infections. Anaerobic organisms can often be suspected because many anaerobes have characteristic microscopic morphology (appearance). For example, Bacteroides spp. are gram-negative rods that are pleomorphic (variable in size and shape) and exhibit irregular bipolar staining. Fusobacterium spp. are often pale gram-negative spindle-shaped rods having pointed ends. Clostridium spp. are large gram-positive rods that form spores. The location of the spore (central, subterminal, terminal, or absent) is a useful differential characteristic. The presence of growth, oxygen tolerance, and Gram stain results are sufficient to establish a diagnosis of an anaerobic infection and begin antibiotic treatment with a drug appropriate for most anaerobes such as clindamycin, metronidazole, or vancomycin.

Gram-negative anaerobes and some of the infections they produce include the following genera:

  • Bacteroides (the most commonly found anaerobes in cultures; intra-abdominal infections, rectal abscesses, soft tissue infections, liver infection)
  • Fusobacterium (abscesses, wound infections, pulmonary and intracranial infections)
  • Porphyromonas (aspiration pneumonia, periodontitis)
  • Prevotella (intra-abdominal infections, soft tissue infections)

Gram-positive anaerobes include the following:

  • Actinomyces (head, neck, pelvic infections; aspiration pneumonia)
  • Bifidobacterium (ear infections, abdominal infections)
  • Clostridium (gas, gangrene, food poisoning, tetanus, pseudomembranous colitis)
  • Peptostreptococcus (oral, respiratory, and intra-abdominal infections)
  • Propionibacterium (shunt infections)

The identification of anaerobes is highly complex, and laboratories may use different identification systems. Partial identification is often the goal. For example, there are six species of the Bacteroides genus that may be identified as the Bacteroides fragilis group rather than identified individually. Organisms are identified by their colonial and microscopic morphology, growth on selective media, oxygen tolerance, and biochemical characteristics. These include sugar fermentation, bile solubility, esculin, starch, and gelatin hydrolysis, casein and gelatin digestion, catalase, lipase, lecithinase, and indole production, nitrate reduction, volatile fatty acids as determined by gas chromatography, and susceptibility to antibiotics. The antibiotic susceptibility profile is determined by the microtube broth dilution method. Many species of anaerobes are resistant to penicillin, and some are resistant to clindamycin and other commonly used antibiotics.


Diagnosis/Preparation

The health care provider should take special care to collect a contamination-free specimen. All procedures must be performed aseptically. The health care professional who collects the specimen should be prepared to take two samples, one for anaerobic culture and one for aerobic culture, since it is unknown whether the pathogen can grow with or without oxygen. In addition, health care professionals should document any antibiotics that the patient is currently taking and any medical conditions that could influence growth of bacteria.


Aftercare

In the case of vein puncture for anaerobic blood cultures, direct pressure should be applied to the vein puncture site for several minutes or until the bleeding has stopped. An adhesive bandage may be applied, if appropriate. If swelling or bruising occurs, ice can be applied to the site. For collection of specimens other than blood, the patient and the collection site should be monitored for any complications after the procedure.


Risks

Special care must be taken by the health care team obtaining, transporting, and preparing the specimen for anaerobic culture. Poor methodology may delay the identification of the bacterium, may allow the patient's condition to deteriorate, and may require the patient to provide more samples than would otherwise be required. Patients may experience bruising, discomfort, or swelling at the collection site when tissue, blood, or other fluids are obtained.


Results

Negative results will show no pathogenic growth in the sample. Positive results will show growth, the identification of each specific bacterium, and its antibiotic susceptibility profile.


Patient education

A health care team member should explain the specimen collection procedure to the patient. If the patient is seriously ill, the team member should explain the procedure to the patient's family members. The patient and his or her family should understand that because bacteria need time to grow in the laboratory, several days may be required for bacterium identification.


Resources

BOOKS

Anel, Ramon L. and R. Phillip Dellinger. "Sepsis and Bacteremia." In Conn's Current Therapy 2001, edited by Robert E. Rakel and Edward T. Bope. Philadelphia: W.B. Saunders Company, 2001, pp.56–62.

Fischbach, Frances. "Blood Cultures." In A Manual of Laboratory & Diagnostic Tests. 6th ed. Philadelphia: Lippincott Williams & Wilkins, 2000, pp.542–545.

Henry, J.B. Clinical Diagnosis and Management by Laboratory Methods. 20th ed. New York: W.B.Saunders Company, 2001.

Wallach, Jacques. Interpretation of Diagnostic Tests. 7th ed. Philadelphia, PA: Lippincott Williams & Wilkens, 2000.


PERIODICALS

Brook, Itzhak. "Anaerobic Infections in Children." Advances in Pediatrics 47 (2000): 395–437.


ORGANIZATIONS

The American Society for Microbiology. 1752 N St. N.W., Washington, DC 20036. (202) 737-3600. http://www.asmusa.org .

National Center for Infectious Disease, Centers for Disease Control and Prevention. 1600 Clifton Road NE, Atlanta, GA 30333. (800) 311-3435. http://www.cdc.gov .


OTHER

National Institutes of Health. [cited April 5, 2003] http://www.nlm.nih.gov/medlineplus/encyclopedia.html .


Linda D. Jones, BA, PBT (ASCP) Mark A. Best, M.D., MPH, MBA



User Contributions:

mat treganza
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Nov 21, 2007 @ 7:07 am
How long do you normally (incubate) befor calling it no ana isolated. We are keeping it 7day. But in my experience at other labs, we keep anaerobes 4-5 days. I have been noticeing the amount of contamination, especially in thio's for anaerobic growth, part two of my question is, How long to keep thio's and sorce of thios should be set on. Also at my current position we are setting up to many thios, on non-sterial sites.
can you help me with this issue.

Mat Treganza (ASCP)
ATHENS REGIONAL MEDICAL CENTER.
ATEHNS,GA.
mulatu
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Dec 27, 2007 @ 1:01 am
it is an interesting note. but I would like to get more explanation about those anaerobic bacteria.what happens when this bacteria exposed to the oxygen that comes from their metabolic end product?
Anil
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Jan 1, 2008 @ 11:23 pm
I want know in which media one has to maintaine the Propionibacterium acnes bacteria. I can grow bacterium once in anaerobic jar but for maintaining the culture i cant keep subcultured tube or slant for months in the jar. can we use cooked meat medium inoculated with P.acnes followed by an oil overlay to create anaerobic conditions and can I keep it in refrigirator below 4C for one month for maintenance.

Anil
Keith
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Mar 11, 2008 @ 3:15 pm
Hi, I am curious about this Anaerobic bacteria. Reason being I have had issues with my skin around the facial area, and seems to have started only after a removal of 4 wisdom teeth. As for the issues well extremely dry skin, with fissures forming, and an odd burning sensation. Dryness is odd because my skin is of an oily type. This has now be an issue for about 5 years and I am having other issues such as weigh gain problems and odd urine being cloudy and with an unusual odor. So I ask, if this was anaerobic bacteria, could it last 5 years without a more serious symptom ?

I have made attempts with my doctor and dermatologist to have this looked at to no avail.But these symptoms are quite general and difficult to pinpoint. Would this not show in a blood examination ? Take care and thank you for your time.
hidayah
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Aug 7, 2008 @ 12:00 am
GOOD INFO!the content is sufficient and the explanation is accurate. includes examples and pictures for student or server to understand better
afreen
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Sep 8, 2008 @ 5:05 am
I want to know the selective media for culturing of propionibacterium acnes isolated from acne patients.because i want to do some applied work on p acnes.kindly do the needful.

Thanking you
Brajesh Kumar Jha
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Sep 16, 2008 @ 10:22 pm
i to want this type of article to teach medical student.so kindly send me this type of article to me. thanking you.
vinothini
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Nov 15, 2008 @ 9:09 am
I need more explation about those obligate anaerobic bacteria.what happens when this bacteria exposed to the oxygen ? Whether its die/ WHY?
Vinothini
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Nov 15, 2008 @ 9:09 am
it is an interesting note. but I would like to get more explanation about those anaerobic bacteria.what happens when this bacteria exposed to the oxygen that comes from their metabolic end product?
Madhu Pillai
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Apr 9, 2009 @ 3:03 am
i am pleased to go through the site and is very informative. i would like to know the application/effectiveness of TSB & FTG media on anaerobe growth.
rachel
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Jun 15, 2009 @ 4:16 pm
HI IM RACHEL,
I WANT TO KNOW ABOUT ANAEROBIC BACTERIA. HOW THEY SURVIV WITH OUT OXYGEN IN THE HUMAN BODY.AND I NEED COMPLETE INFORMATION.
THANK YOU.
nagib
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Jul 31, 2009 @ 4:16 pm
hi this i files you need st be read it carfuly files you need st be read it files you need st be read it files you need st be read it
Dianna
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Aug 7, 2009 @ 1:01 am
Hi, Thanks for explanation about bacteria. I have removed my tooth after treating by Clindamycin for 10 days -300 mg. evry 6 hours . Didn't work for infection shown on ex -ray.I deciced to remove the tooth in order to avoid Anaerobic bacteria to be progressing ..if it is there . I do knot know yet. But I have my neck hurts,lymph nodes,throat. After tooth is removed I still see infection in the hole ! I just reading this info on line trying to help myself ! Endodontics, and Oral surgion so far didn't help me yet. If you know someone in DC/VA area Infectionist with Experience , I would appreaciate INFO ! Thanks D.
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Apr 24, 2010 @ 1:01 am
hi i am sumit, this is very consolidated summary of the anaerobic bacteria, such a nice information about anaerobic bacteria. and i think its helpful for all.
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May 5, 2010 @ 4:04 am
hi
i want to know, why is it that incubation is done for at least 48 hrs and not less?? Any particular reason??
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Sep 14, 2010 @ 4:04 am
hi
i would like to know whether this is applicable for aerobic bacterial culture too ?
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Sep 20, 2010 @ 7:19 pm
Hai,
i want to know how to maintain anaerobic culture for dieertation purpose more than month?
Maryam
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Oct 13, 2010 @ 11:11 am
on what type of media we can culture these anaerobic bacteria either soft or hard media
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Nov 17, 2011 @ 7:19 pm
I HAVE AN ANROBE TYPE BUG AT MY SUPRA PUBIC CAFATOR SITE I AM ON CLYINDAMISON SORRY IF I HAVE SPELT THE ANTIBIOTIC I HAVE HAD TO GO TO THE XDENTIST BECAUSE I HAD 2 ABSESSES UNDER MY 2 BACK TEETH AND HAD TO HAVE THE 2 TEETH TAKEN OUT WAS IT THE INFECTION THAT CAUSED MY TEETH TO BE TAKEN OUT CAN SOMEONE REPLY BACK TO ME
MANY THANKS

GILLIAN BELL
megan
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Jul 23, 2012 @ 11:23 pm
what do anaerobic bacteria need to groe appart from moisture temperture and time
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Jul 30, 2012 @ 9:09 am
thanking u sir.Sir i m doing a master degree thesis on anaerobic bacteria in TU Teaching Hospital in nepal. I want to know all the things regarded anaerobic culture which is possible in nepal contest. I have also got a lot of knowledge from this chapter
admire mzezewa
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Aug 30, 2012 @ 12:00 am
Is it necessary to test skimmed milk powders for thermophilic anaerobic spores prior use in margarine production?This question follows my consideration of exposure of the powder to oxygen at various stages in production.
powell mwenda
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Sep 13, 2012 @ 1:01 am
I AM THANKFUL FOR ALL YOUR EFFORTS IN EDUCATING US ON ANAEROBES,BESIDES I WOULD LIKE TO KNOW THE
PROCEDURE OF OBTAINING THIS ANAEROBIC CONDITIONS ON THE VARIOUS METHODS STATED
Jerry
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Apr 5, 2013 @ 8:20 pm
Hi, how should I maintain the anaerobe cultures once I successfully isolates them from the samples I work on?
Thanks
tasmima
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Apr 15, 2013 @ 7:07 am
HI,
I have some questions. what is the selective media for clostridium species? After isolation of different clostridium species how can I identify each species?
Shaky
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Jun 3, 2013 @ 12:12 pm
Can anaerobic bacteria survive in prostate? Can we culture anaerobe in prostate?
mike
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Jun 30, 2013 @ 12:12 pm
CAN YOU HELP,I HAD A JAW INFECTION AT 19YRS OLD IT WAS TREATED WITH CLINDAMYCIN, THE DOCTOR SAID I WOULD NEED A OPORATION WITHIN FIVE YRS IT WILL NOT BE FUNDED BY NHS. NOW FORTYTWO AND KEPT MYSELF IN GOOD HEALTH IT HAS RE-APEARED, THE INFECTION IS SAID TO BE BONE ABBCESS I HAVE HAD VARIOUS ANTI-BIOTICS OVER A 1YR, WHICH I TOLD THE DOCTOR WOULDNT WORK. HE SEEMS UN-EDUCATED ABOUT ANAROBES I WAS SENT TO PHYSIO I NEW IT WAS A WASTE OF TIME, I HAVE HAD TWO DIFFRENT SCANS AND THEY SHOWED NOTHING WAS WRONG I ASKED THE DOCTOR WOULD HE GIVE ME CLINDAMYCIN, BECAUSE THE DOCTOR FROM YRS AGO SAID IT WAS THE ONLY ANTI-BIOTIC THAT WOULD GET RID OF IT. HE SAID NO BECAUSE IT WAS TO HEAVY ON THE GI TRACT, SINCE I HAVE LOSS OF FEELING AROUND EAR AND CHEEK THIS TIME IT PUT ME IN BED FOR FOUR DAYS. I AM VERY CONCERNED BECAUSE I WAS ALSO TOLD IT WAS VERY DANGEROUS TO MY LIVER AND KIDNEYS I DONT SEAM TO BE GETTING ANYWHERE WITH IT I HAVE FLUCTUATING FLU LIKE SYMPTOMS THAT SEAM TO BE FADING ALSO FACIAL SWELLING AND FEELINGS OF BODY NOT QUITE RIGHT AND HAD LOSS OF HEARING AT ONE POINT, I HAVE BEEN TAKING PROBIOTICS AND HAVING YOGURTS ANY SUJESTIONS PLEASE.

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