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MICROBIOLOGY: Bacterial culture and antibacterial sensitivity testing.

The primary role of the Veterinary Clinical Laboratory is to provide information which veterinarians can use to diagnose and treat infectious diseases.

For this the two pieces of information needed by the clinician are:

  1. whether an infectious agent is present or not
  2. which antibacterial agent should provide adequate treatment.

In choosing a diagnostic laboratory, a laboratory which specializes in providing a veterinary diagnostic service is preferable. It should also have accreditation to a recognized body, such as the UKAS (United Kingdom Accreditation Service), which indicates the laboratory has standardized techniques and its staff are correctly trained and competent. It also indicates a high standard of customer service to clients. Such a laboratory should have an understanding of the bacteria involved in infectious diseases of animals and employ the correct methods to isolate and identify these bacteria. Correct identification is an important part of the service provided and can involve a variety of criteria e.g. growth on certain media, growth at certain temperatures, colonial morphology, microscopic appearance and biochemical reactions. The latter are now more commonly carried out using commercial identification systems which include multiple tests, and the identification is determined by comparing the results with an extensive computer data base of known isolates. These systems can be automated to allow large numbers of tests to be carried out efficiently and cost-effectively. These systems allow identification within 24 hours or less, and in some cases in a few hours. Newer techniques being trialed at the moment allow identification within minutes. Once the organism has been isolated and identified the next step is to carry out antibacterial sensitivity testing. Identification procedures and antibacterial sensitivity testing are often carried out in parallel to speed the process.

 

Antibiotic Sensitivity testing:

Antibiotic sensitivity testing is usually carried out by one of two methods; either disc diffusion, i.e. antimicrobial disc susceptibility testing, or broth dilution antimicrobial susceptibility testing.

With broth dilution antibacterial testing the test bacterium is grown in a range of broths containing different dilutions of the antibiotic. The minimal inhibitory concentration (MIC) is defined as the lowest concentration of antibiotic in micrograms per milliliter (µg/ml) that prevents in vitro growth of the bacterium. These techniques can be automated to read the growth of the organism continuously and base the results on growth kinetics, which can speed up the availability of results with same day results possible. This also allows greater control and standardization of the testing.

Results are reported with an MIC value for each antibiotic tested. Rather than just giving an answer as to whether an isolate is sensitive or resistant to particular antibiotic, the MIC value gives an idea as to how sensitive or resistant the organism is. The laboratory should provide support in interpreting and using this information. There are published guidelines for interpretation of MICs published by CLSI (Clinical and Laboratory Standards Institute).

bacteria_dish_man

Minimally the results should be reported as one of the following:

SENSITIVE – implies that the organism should respond to the usual doses of the antibiotic administered by an appropriate route, including orally.

INTERMEDIATE – implies that the isolate may be inhibited by the concentrations of the drug that are achieved when the maximum parenteral doses are given, but consideration should be given to other choices that may provide more optimal therapy.

RESISTANT – indicates that the bacterium will not inhibited by achievable concentrations of the antibiotic, and therefore the drug should not be chosen for treatment except in certain body fluids or topically where high concentrations of the antibiotic may accumulate.

With disc diffusion susceptibility testing an agar plate is inoculated with a suspension of the bacteria being tested. Antibiotic impregnated discs are placed on the plate and the antibiotics diffuse in to the surrounding medium. If the bacterium is susceptible to a particular antibiotic there will be a clear zone of inhibition around the disc where no bacterial growth occurs. Originally it was thought that the zone sizes correlated with the degree of susceptibility of the organism to these antibiotics, i.e. the larger the zone the more effective the antibiotic. However it is now recognized that other factors affect the diffusion of the antibiotic in to the medium and influence the results. Techniques should be standardized to reduce the influence of these variables and tests carried out and interpreted in accordance with published guide lines e.g. using CLSI guide lines. The CLSI guide lines are based on a correlation between zone sizes and MICs.  The zone size in itself has no meaning and the method is useful only for those organisms for which interpretive guide lines are available.

For both methods standardized procedures should be followed and adequate quality control procedures implemented using reference strains with known antibacterial sensitivities. Guidelines for antibacterial treatment of infections in different body systems have recently been published (e.g. BSAVA SAMSOC, ISCAID [International Society for Companion Animal Infectious Diseases], Pfizer Animal Health and Bayer Animal Health). Antibiotic sensitivity testing should take these guidelines in to account.

Fast, accurate isolation and identification of bacteria involved in infectious disease allows clinicians to choose the most optimal and effective treatment for the patient. The data can monitor emerging bacterial resistance patterns, and can be used to modify antibiotic prescribing.

The Bella Moss Foundation wishes to thank Larry Roberts for this contribution

For more on Data Idexx please contact
www.petresist.com

I am very pleased to be serving as an advisor to the Bella Moss Foundation. BMF has for several years been a unique international resource for pet owners worldwide who desperately need information and support regarding MRSA and its effect on their pets and family. The Bella Moss Foundation has now taken on a new goal to broaden their resources in America. I look forward to helping them to raise awareness in the U.S and Canada and will strive to improve the collaboration between the veterinary and human medicine professions in North America.

Dr Richard Oehler MD. University of South Florida College of Medicine http://health.usf.edu/medicine/ internalmedicine/infectious/

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