MLAB 2434 –Microbiology Keri Brophy-Martinez Bacteremia MLAB 2434 –Microbiology Keri Brophy-Martinez

Definitions Pseudobacteremia False bacteremia Contamination of a blood culture during or after collection

Definitions Bacteremia – presence of bacteria in blood stream Some conditions have a period of bacteremia as part of the disease process (ex. Meningitis, endocarditis) Usually occurs due to a disruption of skin or mucosal barriers to bacterial invasion

Classifications of Bacteremia Classified by Site of Origin Classified by Causative Agent Classified by Place of Acquisition Classified by Duration

Classification by Site of Origin Primary Bacteremia Blood stream or endovascular 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) Fever of Unknown Origin (FUO) Source unknown

Classification by Causative Agent Gram positive bacteremia Gram negative bacteremia Anaerobic bacteremia Polymicrobial bacteremia

Classification by Place of Acquisition Community-acquired Health-care acquired/Nosocomial Defined as occurring 72 hours post admission

Classification by Duration 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 Organisms from an intravascular source that are consistently present in bloodstream

Sepsis & Septicemia Presence of active bacteria Results from continuous bacteremia Clinical signs and symptoms of bacterial invasion and toxin production Apply the SIRS criteria Systemic response to bacterial infection

Bacteremia Complications Septic shock Results from body’s reactions to bacterial bi-products Endotoxins: lipopolysaccharide Exotoxins Disrupts many body functions Hemodynamic changes, decreased tissue perfusion and compromised organ & tissue function Mortality 40% to 50%

Bacteremia/Septicemia Risk Factors Immunocompromised patients Due to decrease in circulating neutrophils Increased use of invasive procedures & indwelling devices Disrupts normal flora Age of patient Young: defect in humoral immunity Old: Decreased immune competency Administration of drug therapy Broad spectrum antibiotics decrease normal flora Increase in antimicrobial resistance

Sources of Bacteremia Pericarditis and Peritonitis Pneumonias Pressure sores Prosthetic medical devices Total hip replacement Skeletal system Skin and soft tissue Urinary Tract Infections

Clinical Signs and Symptoms Abrupt onset of chills, fever, or hypothermia and hypotension Prostration (exhaustion/weakness) and diaphoresis (perspiration) Tachypnea (rapid breathing) is an early sign of bacteremia Delirium, stupor, agitation Nausea, vomiting

Clinical Signs and Symptoms (cont’d) Laboratory Values in Bacteremia Thrombocytopenia Leukocytosis or leukopenia Acidosis Abnormal liver functions Coagulopathy DIC Elevations in CRP, haptoglobin, fibrinogen, ESR, procalcitonin

Specimen Collection Positive blood cultures Best Practice Critical value Physician correlates finding to clinical picture to verify septicemia Best Practice Collect specimen immediately PRIOR to rise in temperature Collect PRIOR to antibiotic therapy

Specimen Collection Aseptic collection procedure is critical Cleansing agents Tincture of iodine (1-2%) Leave on skin for 30 seconds Povidine-iodine (10%) Leave on skin 1.5 to 2 minutes Chlorhexidine/ChloraPrep 2% chlorhexidine gluconate + 70% isopropyl alcohol Cleansing Technique In concentric fashion, from inside to out After cleaning, wait 1.5-2 minutes Acceptable Contamination Rate 1-3%

Collection sites Preferred Less common Peripheral venous Arterial sites Less common Central venous catheters Arterial lines

Blood Collection Devices Traditional set Aerobic bottle Selects for aerobic & facultative anaerobes Anaerobic bottle Selects for obligate anaerobes ARD bottle (Antibiotic Removal Device) Used when patient is on antibiotics prior to blood collection SPS= Sodium polyanetholsulfonate

Blood Collection Devices Anticoagulants SPS= Sodium polyanetholsulfonate Function/Purpose Anticoagulant Neutralizes human serum Prevents phagocytosis Inactivates certain antimicrobial agents SAS(sodium amylosulfate) Similar to SPS, but less effective in neutralizing serum

Specimen Collection: Blood Volume Ideal ratio of blood: broth 1:5 to 1:10 Dilution aids in preventing the bactericidal effect of WBCs & complement Volume Recommendations by Age Younger than 10 years- 1 mL of blood for every year of life Over 10 years- 20 mL Short draw? Inoculate anaerobic bottle first

Specimen Collection: Frequency of Collection Depends if bacteremia is transient, intermediate or continuous General guidelines Usually x2 from different body sites, when patient is spiking a fever Endocarditis 3 sets from 3 different sites within 1-2 hours of clinical presentation Fever of Unknown Origin (FUO) Initially 2 sets; 24-36 hours later, obtain 2 more

Specimen Collection: Frequency of Collection If a catheter-related bloodstream infection is suspected: One set drawn peripherally One set drawn via catheter

Blood Culture Methods Conventional Broth Systems 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, at 12, 24 and 48 hours Method not recommended due to risk of needlestick and contamination; not cost effective

Blood Culture Methods (cont’d) Biphasic Broth-Slide System Agar “paddles” attached to top of bottle; includes CA, MAC, malt extract agars Incubate at 35 OC for 7 days Allows for blind subcultures Closed system

Blood Culture Methods (cont’d) Lysis-Centrifugation Blood Culture Systems (Isolator) Used in the recovery of Fungus and AFB The Isolator is a special tube that contains saponin, a chemical that lyses cells and other anticoagulants Approximately 7.5-10 ml of blood is placed in the tube, then centrifuged to concentrate microorganisms; sediment is subcultured to fungal and/or mycobacterial media

Blood Culture Methods (cont’d) Automatic Blood Culture Systems BacTec 9000 Series Fluorescent light is used to detect changes in CO2 levels

Bactec 9000 Series

Automatic Blood Culture Systems (con’t) ESP( Extra Sensing Power) Now VersaTREK Measures consumption/production of gases; such as CO2 H2, N2 and O2 in the headspace of each bottle Detects a change in pressure

Automatic Blood Culture Systems (con’t) BacT-Alert Carbon dioxide production results in a pH change pH change results in color change detected by system as “positive”

Blood Culture Workup Incubation times Reporting results Routine aerobic/anaerobic 5-7 days Endocarditis 2 weeks Brucellosis/Fungemia/HACEK 21-28 days Reporting results Initial report is sent out at 24 hours Final report is sent out at 5-7 days for all no growth specimens

Blood Culture Workup Positive Cultures Gram stain the bottle to determine the morphology of the organism present Call the results of the gram stain to the physician or nurse, including how many sets etc., so that antibiotic therapy can be initiated Subculture to appropriate media Identify organism and perform sensitivity testing

Blood Cultures: Pathogens Staphylococcus aureus Streptococcus pneumoniae Haemophilus influenza Pseudomonas species Neisseria species Coagulase negative Staphylococcus species (immunocompromised) Group B Streptococcus (infants) Alpha hemolytic Streptococcus viridans group Gram negative rods Yeasts and molds Anaerobes

Blood Cultures: Contaminants Coagulase negative Staphylococcus Propionibacterium acnes Alpha hemolytic Streptococcus viridans group Bacillus species Diphtheroids Growth of multiple organism

Treatment & Prevention Empirical treatment, initially, with broad spectrum antibiotic Antisepsis therapy; physiological support, anticoagulation agents, glucocorticoids Adjunctive measures; draining fluids, removing catheters Prevention Vaccines; S. pneumo, influenza, varicella

References Broyles, M. (2013, June). A Closer Look at Sepsis. ADVANCE for Medical Laboratory Professionals, 25(5), 12-13. http://www.achats-publics.fr/Fournisseurs/BIOMERIEUX.htm http://www.bd.com/ds/productCenter/212536.asp http://www.bd.com/ds/productCenter/445718.asp http://www.temple.edu/medicine/microbiology_lab.htm Kiser, K. M., Payne, W. C., & Taff, T. A. (2011). Clinical Laboratory Microbiology: A Practical Approach . Upper Saddle River, NJ: Pearson Education. Mahon, C. R., Lehman, D. C., & Manuselis, G. (2011). Textbook of Diagnostic Microbiology (4th ed.). Maryland Heights, MO: Saunders.