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SPS TUBES

SPS for blood culture specimen collections in microbiology

In order to provide quality results in microbiology, the laboratory must treat the specimen appropriately. Specimen selection and collection, proper transport, specific plating, incubation and processing all contribute to the successful isolation of potential pathogens.

SPECIMEN FROM A NORMALLY STERILE SITE

Specimen collection from these sites requires a needle puncture or surgical procedure. The following procedures are designed to reduce the risk of contamination with skin flora. Decontamination of Skin Clean the puncture site with 70% alcohol. Clean the puncture site with an antiseptic povidone-iodine preparation. Allow to remain on the skin for at least 1 to 2 minutes. Do not probe with your finger after puncture site has been decontaminated. Following venipuncture, remove iodine solution with 70% alcohol.

Collection of Clinical Material

Blood Culture - Since blood cultures are processed using special media, it is necessary to submit all blood cultures in appropriate plates. Most often, two separate sets of blood samples will suffice. - If a physician orders two separate sets of blood cultures, the phlebotomist would use two separate venipuncture sites, drawing one culture set from the right arm and one culture set from the left arm; in most cases both sets may be obtained without any time elapsing between sites. - Multiple plates filled from a single venipuncture site should be interpreted as a single blood culture set. - Preferably, blood for culture should not be drawn through an indwelling or intra -arterial catheter. - Draw the sample of each set of blood cultures with a needle and syringe. Before injecting the blood into the sps tubes, decontaminate the diaphragm tops by swabbing with 70% alcohol and allow to dry. - Blood for mycobacterial or fungal cultures can be transported in tubes containing SPS. Gently mix the tube(s) following inoculation. Do not vent or refrigerate.

We recommend the following procedure for the timing of blood cultures and optimal recovery of microorganisms present:

a) Before the use of systemic anti-microbials obtain 2 separate sets of blood cultures when there is a fever combined with significant leukocytosis or leukopenia. b) Systemic and localized infection - Suspected acute sepsis, meningitis, osteomyelitis, arthritis, or acute untreated bacterial pneumonia: Obtain two sets of blood cultures. - Fever of unknown origin: Initially obtain two sets of blood cultures; 24-36 hours later obtain 2 additional sets of blood cultures. Note: the yield beyond 4 sets of blood culture is often negligible. - Suspected early typhoid fever or brucellosis: Owing to the low grade bacteremia present in these infections, obtain 4 sets of blood cultures (the same venipuncture site may be used) over a 24-36 hour period. c) Infective endocarditis - Acute: Obtain 3 sets of blood culture during the first 1-2 hours of evaluation. - Sub Acute: Obtain 3 sets of blood cultures on the first day (ideally 15 or more minutes apart; the same venipuncture site may be used). If all 3 sets are negative 24 hours later, obtain 2 additional sets of cultures. - Culture negative endocarditis: If 5 negative sets of blood cultures are obtained, special culture techniques may be advised. º

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AISBACT A

For blood culture

The AISBACT A system is based on the lysis of blood elements using a mixture contained in a test tube, which does not alter either the viability or number of microorganisms present, and the subsequent centrifugation of the sample for cultivation the sediment obtained on agar plates selected according to the criteria of the Microbiologist.

CONTENS

Each box contains 20 ready to use tubes, containing one mL of the special lythic mixture. The tubes contain partial vaccum, facilitating their use for direct collection of the sample, taking care that at no time does the lythic solution touch the rubber stopper during sampling.

METHOD

Collection of the sample # Disinfect the stopper of the tube with iodine alcohol. # Disinfect the puncture area of the skin with iodine alcohol and cleanse with alcohol.Then using a sterile syringe or the AISBACT A tube, collect 10 mL of peripheral blood. If blood has been extracted by syringe, it should be introduced quickly into the 10 mL AISBACT tube. # Carefully, withdraw the needle. # Invert the tube five or six times until a homogenous mixture of the contens has been obtained (do not shake vigorously). # Transport the tube to the laboratory within 6 hours for processing, (if transportation is delayed more than 4 hours the resultant CFU/mL will not be representative). Equally, if the patient is under antibiotherapy, the processing should be caried out as quickly as possible.

METHOD

Processing the sample. # Once in the laboratory, shake the tube vigorously for 10 seconds. # Spin down in a free angle centrifuge for 30 minutes at 2,500-3,000 g, ensuring that the temperature does not exced 35º C. # Once the centrifugation has been completed, withdraw the tube and place it in a vertical position for processing. The following steps should, if possible, be carried out in a vertical flow cabinet. # Disinfect the AISBACT A stopper with iodine alcohol. # Aseptically remove the stopper and using a sterile pipette, carefully withdraw the supernatant until a volume equivalent to 1-1.5 mL (containing the sediment) is left in the tube. # Using a stirrer, mix the sediment vigorously and the remainder of the supernatant in the tube. # With a new sterile pipette, withdraw the ENTIRE contents of the tube and disperse in the centre of 3-4 agar plates in accordance with the instructions of the microbiologist. It is important that the plates shouldn't be moist as this increases the absorption of the inoculum. # Using a loop or the end of the same pipette, disperse the sample but this time in a direction perpendicular to that carried out formerly. # Place the plates upside down in the incubator set for the appropiate conditions ( aerobic, anaerobic, etc). # Examine the aerobic plates after 24 hours and the anaerobic after 48 hours.

RESULTS AND INTERPRETATION

# The appearance in the inoculated area of one or more colonies, irrespective of species, should be considered positive. # The appearance outside the cultivated area of one or more colonies should be considered as contaminated. # The presence of different colonies inside and outside the culture area should be considered as positive inside the area and evidence of contamination outside. # The presence of the same type of colony both inside and outside the cultivation area should be interpreted in terms of clinical manifestations, although this generally corresponds to contamination.

CALCULATION OF THE COLONY FORMING UNITS PER mL (CFU/mL)

If the cultivation has been carried out within four hours of collecting the sample and not been stored in a refrigerator, the AISBACT A system makes possible to obtain the number of CFU/mL and allows this to be taken into account both in the overall assessment of the clinical picture and in determining patient treatment. To calculate the Colony Forming Units per mL, proceed as follows : Total nº of colonies Total number of on all the plates cultivated plates ---------------------------- X ----------------------------- = CFU/mL Nº of plates on which Total volume of the germ is to grow blood

ADVANTAGES OF THE SYSTEM

The AISBACT A blood culture system’s immediate processing offers the following advantages : * A more rapid isolation and identification of the casual germs. * Determination of CFU´s per mL. * Increased number of positive results.

REFERENCES

Zierdt,Ch.;Peterson,D., and alt. J.Clin.Microbiol. 15,74,1982. Zierdt,Ch. J.Clin.Microbiol. 17,628,1983 Henry,N.;McLimans,C., and alt. J.Clin.Microbiol. 17,864,1983 Kiehn,T.;Wong,B., and alt. J.Clin.Microbiol. 18,300,1983 Gill,V.;Zierdt,Ch., and alt. J.Clin.Microbiol. 20,927,1984 Marí,M.;Vich,J.M. Datos de archivo,1989.

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AISBACT-i

System for pediatric use.

The AISBACT-i system is based on the lysis of blood elements using a mixture contained in a test tube, which does not alter either the viability or number of microorganisms present, for the subsequent cultivation on agar plates selected according to the criteria of the Microbiologist of the entire blood sample treated.

CONTENS

Each box contains 20 ready to use sterile tubes, each containing a special lythic mixture. The tubes contain a partial vaccum.

METHOD

Collection of the sample

Disinfect the stopper of the tube with iodine alcohol. Disinfect the puncture area of the skin with iodine alcohol and cleanse with alcohol. Then using a sterile syringe collect 1-2.0 mL of peripheral blood. Carefully, withdraw the needle Invert the tube five or six times until a homogenous mixture of the contens has been obtained (do not shake vigorously). Transport the tube to the laboratory within 6 hours for processing (if transportation is delayed more than 4 hours the resultant CFU/mL will not be representative).Equally if the patient is under antibiotherapy, the processing should be caried out as quickly as possible.

Processing the samples

Once in the laboratory, shake the tube vigorously for 10 seconds Disinfect the AISBACT I stopper with iodine alcohol. Ussing a disposable sterile syringe, prick the stopper and with the tube placed in an upside down vertical position, extract the contents Distribute the contents of the tube in the centre of 3-4 Petri dishes (the amount should not exceed 0,4 mL per plate).It is important that the plates shouldn't be moist as this increases the absorption of the inoculum. Using a loop spread the sample on the plates (it is not necessary to sterilise the spatula plate by plate for the same sample). Place the plates upside down in the incubator set for the appropiate conditions (aerobic, anaerobic). Examine the aerobic plates after 24 hours and the anaerobic after 48 hours.

RESULTS AND INTERPRETATION

The appearance in the inoculated area of one or more colonies, irrespective of species, should be considered positive. The appearance outside the cultivated area of one or more colonies should be considered as contaminated. The presence of different colonies inside and outside the culture area should be considered as positive inside the area and evidence of contamination outside. The presence of the same type of colony both inside and outside the cultivation area should be interpreted in terms of clinical manifestations, although this generally corresponds to contamination.

CALCULATION OF THE COLONY

FORMING UNIT PER mL (CFU/mL)

If the cultivation has been carried out within four hours of collecting the sample and not been stored in a refrigerator, the AISBACT-i system makes possible to obtain the number of CFU/mL and allows this to be taken into account both in the overall assessment of the clinical picture and in determining patient treatment. To calculate the Colony Forming Units per mL, proceed as follows : Total nº of colonies Total number of on all the plates cultivated plates ------------------------------- X --------------------------- = CFU/mL Nº of plates on which Total volume of the germ is to grow blood

ADVANTAGES OF THE SYSTEM

The AISBACT-i blood culture system's immediate processing offers the following advantages :

* A more rapid isolation and identification of the casual germs * Determination of CFU´s per mL * Increased number of positive results

REFERENCES

Zierdt,Ch.;Peterson,D., and alt. J.Clin.Microbiol. 15,74,1982. Zierdt,Ch. J.Clin.Microbiol. 17,628,1983 Henry,N.;McLimans,C., and alt. J.Clin.Microbiol. 17,864,1983 Kiehn,T.;Wong,B., and alt. J.Clin.Microbiol. 18,300,1983 Gill,V.;Zierdt,Ch., and alt. J.Clin.Microbiol. 20,927,1984 Marí,M.;Vich,J.M. Datos de archivo,1989.

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