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Blood Culturing in Infections

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Dr.T.V.Rao MD

Dr.T.V.Rao MD

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What is a Blood Culture?

  • A blood culture is a laboratory test in which blood is injected into bottles with culture media to determine whether microorganisms have invaded the patient’s bloodstream.

Dr.T.V.Rao MD

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Need for Blood Culture?

No microbiological test is more essential to the clinician than the blood culture. The finding of pathogenic microorganisms in a patient’s bloodstream is of great importance in terms of diagnosis, prognosis, and therapy.”

- L. Barth Reller, Clin. Infect. Diseases, 1996

Dr.T.V.Rao MD

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Blood Culture is done to Detect Infectious Diseases

  • Blood culture is a microbiological culture of blood. It is employed to detect infections that are spreading through the bloodstream (such as bacteremia, septicemia amongst others). This is possible because the bloodstream is usually a sterile environment

Dr.T.V.Rao MD

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Proof in Blood borne Infection

  • A clinically suspected infection is ultimately confirmed by isolation or detection of the infectious agent. Subsequent identification of the microorganism and antibiotic susceptibility tests further guide effective antimicrobial therapy. Bloodstream infection is the most severe form of infection and is frequently life-threatening, and blood culture to detect circulating microorganisms has been the diagnostic standard.

Dr.T.V.Rao MD

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  • Bacteremia – presence of bacteria in blood stream

    • Some conditions have a period of bacteremia as part of the disease process (ex. Meningitis, endocarditis)

  • Septicemia – bacteremia plus clinical signs and symptoms of bacterial invasion and toxin production

Dr.T.V.Rao MD

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Dr.T.V.Rao MD

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Definitions (cont’d)

  • Primary Bacteremia – blood stream bacterial invasion with no preceding or simultaneous site of infection with the same microorganism

  • Secondary Bacteremia – isolation of a microorganism from blood as well as other site(s)

Dr.T.V.Rao MD

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Bacteremia and Fungemia Episodes

  • Transient

    • Comes and goes

    • Usually occurs after a procedural manipulation (ex. Dental procedures)

  • Intermittent

    • Can occur from abscesses at some body site that is “seeding” the blood

  • Continuous Bacteremia

Dr.T.V.Rao MD

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Bacteremia Complications

  • Warm shock – fever, increased pulse, hyperventilation, and warm, dry flushed skin

  • Cold shock – decreased blood pressure, increased pulse, and rapid, shallow respirations

  • Septic chock

    • Hemodynamic changes, decreased tissue perfusion and compromised organ & tissue function

    • Mortality 40% to 50%

Dr.T.V.Rao MD

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Bacteremia/Septicemia Risk Factors

  • Immunocompromised patients

  • Increased use of invasive procedures

  • Age of patient

  • Administration of drug therapy

Dr.T.V.Rao MD

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Sources of Bacteremia Spread

  • Pericarditis and Peritonitis

  • Pneumonias

  • Pressure sores

  • Prosthetic medical devices

  • Total hip replacement

  • Skeletal system

  • Skin and soft tissue

Dr.T.V.Rao MD

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Blood culturing most important and life saving Investigation

Needs optimal Methods for Diagnosis of Blood Borne Pathogens

Dr.T.V.Rao MD

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  • Aseptic collection procedure is critical Amount of blood

    • 1:10 ratio of blood to broth

    • Younger than 10 years – 1 ml of blood for every year of life

    • Over 10 years – 20 ml

Dr.T.V.Rao MD

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Blood Collection

  • Frequency of Collection

    • Depends if bacteremia is transient, intermediate or continuous

    • Number of cultures collected are usually inversely related to the type of bacteremia

    • Usually x3 from different body sites

Dr.T.V.Rao MD

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Blood Culture Methods

  • Conventional Broth Systems

    • One aerobic bottle and one anaerobic bottle per blood collection

    • Aerobic broth contains soybean casein digest broth, Tryptic or trypticase soy broth, Brucella agar or Columbia broth base

    • Anaerobic broth is usually the same as aerobic with addition of 0.5% cysteine in an aerobic environment

    • Must be subcultured and gram stained manually

Dr.T.V.Rao MD

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  • Venipunctureis the process of obtaining intravenous access for the purpose of intravenous therapy or obtaining a sample of venous blood. This procedure is performed by medical laboratory scientists, medical practitioners, some EMTs, paramedics phlebotomists and other nursing staff. Venipuncture is one of the most routinely performed invasive procedures and is carried out for two reasons, to obtain blood for diagnostic purposes or to monitor levels of blood components (Lavery & Ingram 2005).

Dr.T.V.Rao MD

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Phlebotomy Definition

  • phle·boto·my (fli) nounthe act or practice of bloodletting as a therapeutic measure

  • Phlebotomy from Greek words, phlebo, relates to veins, tomy, relates to cutting.

  • Opening a vein to collect blood

Dr.T.V.Rao MD

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  • Properly labelled sample is essential so that the results of the test match the patient. The key elements in labelling are:

    • Patient's surname, first and middle.

    • Patient's ID number.

    • NOTE: Both of the above MUST match the same on the requisition form.

    • Date, time and initials of the phlebotomist must be on the label of EACH tube.

Dr.T.V.Rao MD

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Principles for Collection

  • Gloves will be worn in accordance with standard precautions.

  • •Appropriate verification of the patient's identity, by means of an armband or area specific procedure, will occur before the specimen collection.

  • •Cultures should be drawn before administration of antibiotics, if possible. ???

  • • blood cultures should be drawn from lines, but should be drawn viavenipuncture.

Dr.T.V.Rao MD

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What Materials We need

  • Chlorhexidine swabs (1-2 packages)

  • Alcohol swabs

  • Blood culture bottles (2 bottles per set)

  • 2 syringes (adult: 20 cc, paediatric: 5 cc)

  • 2 needles (adult: 22 gauge or preferably larger butterfly or standard needle; pediatric: 25 or 23 gauge butterfly or standard needle)

  • Gloves (sterile &nonsterile)

  • Tourniquet

  • Sterile gauze pad

  • Adhesive strip or tape

  • Self-sticking patient labels

  • Plastic zip lock specimen bags

Dr.T.V.Rao MD

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The requisitions form should be completely filled out, and the requisition must indicate the tests ordered.

Dr.T.V.Rao MD

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Self Protection

A few ways to make sure your role in the collection process is carried out with efficiency, orderliness and safety

Dr.T.V.Rao MD

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Steps 1 – 3, Check, Explain, Wash

  • 1.Identify the patient

  • 2.Explain the procedure to the patient.

  • 3.Wash hands with soap and water withfriction for 15 seconds or use alcohol based hand rub

Dr.T.V.Rao MD

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  • Chlorhexidine swabs (1-2 packages)

  • Alcohol swabs

  • Blood culture bottles (2 bottles per set)

  • 2 syringes (adult: 20 cc, paediatric: 5 cc)

  • 2 needles (adult: 22 gauge or preferably larger butterfly or standard needle; pediatric: 25 or 23 gauge butterfly or standard needle)

  • Gloves (sterile &nonsterile)

  • Tourniquet

  • Sterile gauze pad

  • Adhesive strip or tape

  • Self-sticking patient labels

  • Plastic zip lock specimen bags

Dr.T.V.Rao MD

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. Barrier protection for the phlebotomist consists of the latex gloves.

Dr.T.V.Rao MD

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Obtaining Blood

  • Locate the vein

  • Prep kit

    • Alcohol 5 sec. Dry 30-60 sec ( resource poor conditions )

    • Ideal to collect with alcohol swabs containing 2% Chlorhexidine and 70% isopropyl alcohol

  • Remove caps, clean with alcohol

  • Put on gloves

  • Without palpating, draw 20 ml and put 10 in anaerobic and 10 in aerobic bottle

  • Dispose of syringe in sharps container

  • Label bottles and send to lab

Dr.T.V.Rao MD

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Method of Blood Collection

  • A minimum of 10 ml of blood is taken through venipuncture and injected into two or more "blood bottles" with specific media for aerobic and anaerobic organisms.

  • The blood is collected using clean technique. This requires that both the tops of the culture bottles and the venipuncture site of the patient are cleaned prior to collection with alcohol swabs containing 2% Chlorhexidine and 70% isopropyl alcohol.

Dr.T.V.Rao MD

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The area of skin is cleaned with a disinfectant, or an alcohol swab.

  • Using sterile gloves, do not wipe away the surgical solution, touch the puncture site, or in any way compromise the sterile process. It is vital that the procedure is performed in as sterile a manner as possible as the persistent presence of skin commensals in blood cultures could indicate endocarditis but they are most often found as contaminants

Dr.T.V.Rao MD

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The vein is anchored and the needle is inserted.

Dr.T.V.Rao MD

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The vacutainer tube is depressed into the needle to begin drawing blood

Dr.T.V.Rao MD

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Additional vacutainer tubes can be utilized. Determine what tests are ordered and what tubes will be necessary BEFORE you begin to draw blood, and determine the order of draw for the tubes..

Dr.T.V.Rao MD

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When the final tube is being drawn, release the tourniquet. Then remove the tube, and remove the needle.

Dr.T.V.Rao MD

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After the needle is removed from the vein, apply firm pressure over the site to achieve haemostasis.

Dr.T.V.Rao MD

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Apply a bandage to the area.

Dr.T.V.Rao MD

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Preparation of Cap before Injecting Blood

  • Prep the rubber cap of the blood culture bottles with an alcohol pad in a circular motion. Allow the alcohol to dry.

Dr.T.V.Rao MD

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Inject the Blood …..

  • Inject the blood into the Selected Media

  • Gently rotate the bottles to mix the blood & the broth (do not shake vigorously).

Dr.T.V.Rao MD

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Follow the universal precautions when disposing Needle

  • Dispose of needle in sharps container and dispose of other waste in proper container

Dr.T.V.Rao MD

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Label the tubes, checking the requisition for the proper identification.

Dr.T.V.Rao MD

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Give the all possible Medical Information

  • Patient’s name

  • • Hospital number (Patient ID)

  • • Patient’s location (room and bed #)

  • • Date and time of collection

  • • Collector’s initials

  • • Site of venipuncture

  • • Or other information as per facility

  • Include you Mobile Contact No – A vital information can be delivered any time

Dr.T.V.Rao MD

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Document the Medical Records

  • Document the following in the medical record:

  • –Date & time specimen obtained

  • –Site of specimen collection

Dr.T.V.Rao MD

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Frequency of Collection

  • Frequency of Collection

    • Depends if bacteremia is transient, intermediate or continuous

    • Number of cultures collected are usually inversely related to the type of bacteremia

    • Usually x3 from different body sites

Dr.T.V.Rao MD

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Second Set

  • If 2 or more sets of blood cultures have been ordered, obtain the second set in the same manner as the first, making a new venipuncture at a different site.

Dr.T.V.Rao MD

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Traditional Methods in Blood cultures

  • Most microbiological culture procedures require the use of solid media, like blood agar and Mac Conkeys agar plates that need to be visually monitored by trained personnel at intervals of 24 hours. These conventional cultures using normal media take at least a minimum of 72 hours to isolate the pathogen and carry out susceptibility test to know the efficacy of antibiotics on simple aerobic bacteria.

Dr.T.V.Rao MD

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Bacteria and Fungi Are Identified by Phenotypic Characters

Dr.T.V.Rao MD

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Biochemical Tests gives Better Clues in Identification

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Newer Blood Culture Methods

  • Newer Blood Culture Systems

    • Biphasic Broth-Slide System

      • Agar “paddles” attached to top of bottle

      • Closed system

    • Continuous Monitoring Blood Culture Systems

      • BacTec – measures 14CO2

      • BacTec 9000 Series – measures CO2

      • ESP – measures consumption of gases

      • BacT-Alert – measures change in pH

Dr.T.V.Rao MD

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Automation reduces the time requirement

  • But this can be completed within 30 hours by using automated techniques. This is especially useful when large number of specimens needs to be cultured, as the instrument, which has been programmed for the same, automatically screens these.

Dr.T.V.Rao MD

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BacT/AlerT 3D culture system

  • BacT/AlerT 3D culture system. This is the first automated non-radiometric and non-invasive culture system that continuously monitors system for culture of bacteria (both aerobic and anaerobic), fungi and mycobacteria. All these bacteria can be cultured using different media as prescribed..

Dr.T.V.Rao MD

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bioMérieux BacT/ALERT® 3D

  • The bioMérieux BacT/ALERT® 3D provides an optimal environment for the recovery of a wide range of pathological organisms, including bacteria, yeasts and mycobacteria; utilizing proprietary plastic culture bottles ensuring added safety to the user.

Dr.T.V.Rao MD

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BacT/ALERT® 3D Microbial Detection System

This newest generation of the time-tested BacT/ALERT system offers advantages in every dimension of testing. From its space-saving modular design to its easy touch-screen operation and flexible data management options, every laboratory will find something to love about the BacT/ALERT 3D!

Dr.T.V.Rao MD

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Principles of functioning of BacT alert Monitors

  • Microorganisms multiply in the media, generating CO2. As CO2 increases, the sensor in the bottle turns a lighter colour.

  • Measuring reflected light, the BacT/ALERT 3D monitors and detects color changes in the sensor.

  • Algorithms analyze the data to determine positivity, and the laboratory is notified immediately with visual and audible alarms.

Dr.T.V.Rao MD

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Principles in BacT/AlerT 3D culture system

  • This is a closed system and works on the colorimetric principle of detection of CO2 produced by the organisms. The CO2 causes a lowering of the pH of the medium, which in turn produces a colour change in a sensor attached to the CO2-sensitive base of each bottle.

Dr.T.V.Rao MD

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Automation improves quality of services

  • Overall, laboratories transitioning from conventional to automated processes find that technologists and microbiologists are more open to innovation and improved quality.

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Automation Signals Bacteremia cases

  • After inoculating the culture vials, they are sent to the clinical pathology microbiology department. Here the bottles are entered into a blood culture machine, which incubate the specimens at body temperature. The blood culture instrument reports positive blood cultures (cultures with bacteria present, thus indicating the patient is "bacteremia"). Most cultures are monitored for 5 days after which negative vials are removed.

Dr.T.V.Rao MD

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The positives cases to be proceeded without delay

  • Avial is positive, a microbiologist will perform a Gram Stain on the blood for a rapid, general ID of the bacteria, which they will report to the attending physician of the bacteremic patient. The blood is also subcultured onto agar plates to isolate the pathogenic organism for culture and susceptibility testing, which takes up to 3 days. This culture & sensitivity (C&S) process identifies the species of bacteria. Antibiotic sensitivities are then assessed on the bacterial isolate to inform clinicians on appropriate antibiotics for treatment.

Dr.T.V.Rao MD

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Culturing Mycobacterium from Blood

  • Mycobacterial growth is generally observed within a week in case of smear (1+) positive.

  • Speciation into mycobacterium tuberculosis complex and mycobacteria other than tuberculosis takes an additional three days.

  • An important Investigation in AIDS and other Immunosuppressed patients

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Testing drug resistance in Tuberculosis a priority

  • Susceptibility testing to primary line of anti-tuberculosis drugs viz streptomycin, isoniazid, rifampicin, ethambutol and pyrizinamide and secondary line of drugs viz kanamycin, para-amino salicylic acid, cycloserine, ethionamide, capreomycin etc requires 5-10 days.

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Rapid Susceptibility Testing

  • Rapid susceptibility will be carried out for gram negative and staphylococcal isolates and other isolates on request. These will be reported within 12 hours using API systems. Automation has made it easier to arrive at a precise laboratory diagnosis of infection 

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The Contaminated Blood Culture

  • If the skin is not adequately cleansed before drawing blood for culture, bacteria on the skin will be injected into the bottle, producing a false positive blood culture

  • It is difficult for the physician to determine whether the bacteria growing in the blood culture is a real pathogen causing bloodstream infection or whether bacteria on the skin have contaminated the culture. This can lead to excess use of antibiotics and prolongation of hospital stay.

Dr.T.V.Rao MD

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  • The programme created by Dr.T.V.Rao MD as Technical Series for Microbiologists in the Developing World

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