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BLOOD CULTURINGIN

INFECTIOUS DISEASES

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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.

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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

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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

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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.

Proof in Blood borne Infection

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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

Definitions

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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)

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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

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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%

Bacteremia Complications

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Bacteremia/Septicemia Risk Factors Immunocompro

mised patients Increased use

of invasive procedures

Age of patient Administration

of drug therapy

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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

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Blood culturing most important and life saving

Investigation

Needs optimal Methods for Diagnosis of Blood Borne

Pathogens

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Blood Collection

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

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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

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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

Blood Culture Methods

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Venipuncture is 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).

Venipuncture

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Phlebotomy Definition

phle·boto·my (fli) noun the 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

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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.

LABELING THE SAMPLE

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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.

Principles for Collection

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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

What Materials We need

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The requisitions form should be completely filled out, and the

requisition must indicate the tests ordered.

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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

Steps 1 – 3, Check, Explain, Wash

1.Identify the patient 2.Explain the

procedure to the patient.

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

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Materials

• 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

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

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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

Obtaining Blood

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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.

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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

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The vein is anchored and the needle is inserted.

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The vacutainer tube is depressed into the needle to

begin drawing blood

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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. .

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

remove the needle.

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After the needle is removed from the vein, apply firm

pressure over the site to achieve haemostasis.

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Apply a bandage to the area.

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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.

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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).

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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

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Label the tubes, checking the requisition

for the proper identification.

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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

Give the all possible Medical Information

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Document the Medical Records

Document the following in the medical record:

–Date & time specimen obtained

–Site of specimen collection

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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

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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.

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• 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.

Traditional Methods in Blood cultures

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Bacteria and Fungi Are Identified by Phenotypic

Characters

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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

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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.

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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..

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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.

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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!

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.

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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.

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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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.

Automation Signals Bacteremia cases

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A vial 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.

The positives cases to be proceeded

without delay

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

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.

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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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.

The Contaminated Blood Culture

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The programme created by

Dr.T.V.Rao MD as Technical Series for Microbiologists in the Developing World

Email doctortvrao@gmail.com