1. Blood Culture in Sepsis
Prof. MD. Ahmet Başustaoğlu
Girne American University
Faculty of Health Sciences
Kyrenia/Northern Cyprus

2. Anand Kumar, et. All., Chest 2009; 136;1237-1248;

4. Each hour of delay in initiation of effective antimicrobials can increase mortality rates by 7.6%.

7. Sepsis: The medical syndrome
Hours to Days
% Mortality
% + blood culture 17 25 69
INFECTION
SIRS
SEPSIS
SEVERE SEPSIS
SEPTIC SHOCK
*Rangel-Frausta, JAMA 273:117-23

8. Standards and Guidelines
WHO guidelines on drawing blood: best practices in phlebotomy
Blood Culture Ordering and Collection Guidelines Vanderbilt University Medical Center
Johns Hopkins Medical Microbiology Specimen Collection Guidelines
Principles and Procedures for Blood Cultures; Approved Guideline M47-A CLSI (Clinical Laboratories Standards Institude)
Cumitech 1C; Blood Cultures, ASM (American Society for Microbiology)

9. Key Points from CLSI 47-A and Cumitech 1C
Timing of drawing blood cultures
Skin disinfection
Volume of blood to inoculate
Number of blood culture sets
How long to incubate bottles?

10. Common organisms associated with sepsis
Gram Positive
Staphylococcus aureus
Viridans Streptococci
Streptococcus pneumoniae
Streptococcus pyogenes
Enterococcus faecalis
Clostridium perfringens
Anaerobic streptococci
Yeasts
Candida albicans and other Candida spp.
Other yeasts e.g.
Cryptococcus neoformans,
Histoplasma capsulatum
Gram Negative
Salmonella typhi
Other Salmonella serovars
Brucella spp.
Haemophilus influenzae
Pseudomonas aeruginosa
Acinetobacter spp.
Klebsiella spp.
Escherichia coli
Proteus spp.
Bacteroides fragilis
Neisseria meningitidis

11. American Journal of Medicine September 2010

12. Blood Culture Indications
Fever (> 38°C)
Hypothermia (< 36°C)
Leukocytosis (> /µl)
Granulocytopenia (< 1.000/µl)
Hypotension
Regional infections: pneumonia, UTI, …
Renal failure
Immune deficiency
CUMITECH Blood Cultures IV, ASM Press 2005
Mylotte and Tayara EJCMID 2000 4

13. “When is the best time to draw blood cultures on a patient with suspected sepsis?”
After an influx of bacteria into the bloodstream there is a lag time of approximately 1 h, after which chills occur. Fever then follows. Draw blood cultures as close as possible to the episode of chills or fever. Do NOT delay, as recovery of microorganisms diminishes with time after the fever spike. Blood Cultures Obtained Prior to Antibiotic Administration
Chills
Blood Cultures
Temp
BACTEREMIA
LEVEL 30 60
Time (min)

14. MEDIA
Multiple nutritionally enriched broth-based culture media have been used successfully.
The most widely used medium is soybean-casein digest broth. Other media include brain heart infusion, supplemented peptone, Columbia, and brucella broths.
Medium formulations designed to enhance the detection of anaerobes, fungi, and mycobacteria are also marketed commercially
Some media with resin or charcoal to inaktivate the antibiotic.
System
Aerobic w/o
Antimicrobial Removal Device
Aerobic with Antimicrobial Device
Anaerobic with Antimicrobial Device
Anaerobic w/o Antinmicrobial Removal Device
Pediatric
Mycosis
Mycobacterium
Bactec
Standart Aerobic F
Plus
Aerobic/F
Plus Anaerobic/F
Lytic/10 Anaerobic /F
PEDS Plus/F
Mycosis IC/F
Myco/F Lytic
BacT/ALERT
-----
FA Plus
FN Plus
------
Peds Plus MP
Versa Trek
Redox 1
Redox 2
Myco

16. Methods of Obtaining Blood for Culture
Venipuncture
Intravascular devices
Venipuncture remains the technique of choice for obtaining blood for culture
Blood cultures obtained from intravascular access devices are associated with greater contamination rates than obtained by venipuncture through a properly prepared skin site
If needed to be obtained from intravenous lines and similar access devices, should be paired with another culture of blood obtained by venipuncture to assist in interpretation in the event of a positive result.

17. Skin Disinfection A positive blood culture represents
Cause of infection
Contamination
Failure to adequately cleaning of the skin increases the risk that microbial flora of the skin, will contaminate the blood culture.
Health care workers who obtain blood cultures are often in a hurry, do not understand the importance of antiseptic preparation contact time.
Cumitech; Blood Cultures, ASM

18. Skin Disinfection Povidone-iodine 2% Tincture of iodine,
0,2% Chloride peroxide,
5% Chlorhexidine gluconate
Tincture of iodine, chloride peroxide, and chlorhexidine gluconate are superior to povidone-iodine preparations
Tincture of iodine and chlorhexidine gluconate are probably equivalent
Chlorhexidine gluconate is recommended for older infants, children, and adults
30 seconds to min of contact time
Chlorhexidine preparations have the advantage of being both colorless and less irritating to skin.

19. Blood-to-Broth Ratio
Human blood contains substances capable of inhibiting microbial growth.
Diluting blood in culture broth reduces the concentrations of these inhibitory substances and concentrations of any antimicrobial agents that may be administered to patients.
Studies that have addressed the blood-to-broth ratio have recommended 5- to 10-fold dilution
Dilutions less than 1:5 may result in reduced yield

20. Methods of Obtaining Blood for Culture
“two-needle” method
“single-needle” method
“transfer set” or “a double-ended needle” method
Higher contamination rates with the single-needle method (3.7%) compared with the two-needle method (2.0%),
the higher rates are tolerated in order to reduce the risk of occupational needle-stick injuries

21. Methods of Obtaining Blood for Culture
Using a transfer set or a double-ended needle,
Low contamination risk
Low blood-borne pathogen risk
Taking the proper amount of blood

22. Blood Culture
At least 2 blood culture sets should be drawn in critically ill patients with suspected sepsis
Catheter-drawn blood cultures
Catheter-drawn blood cultures are equally likely to be truly positive (associated with sepsis), but more likely to be colonized (J Clin Microbiol 38:3393, 2001.)
Both drawn through vein PPV of 98%
One drawn through catheter and other though vein PPV of 96%
Both drawn from catheter PPV 0f 50%
Study of positive coagulase negative Staphylococcus cultures and sepsis (Clin Infect Dis. 39:333, 2004.)
Mike 22

23. Use of Anearobic blood culture bottles;
When one aerobic and one anaerobic bottle was used, a significantly increased yield of some agents was demonstrated;
Streptococci, Staphylococci, Enterococci
Enterobacteriaceae, Anaerobes
In a study by Akyar et al. (Acıbadem University) out of all the positive cultures;
172 (%24.3) was only in anerobic bottles,
310 (%43.7) was only in aerobic bottles,
227 (%32) was both aerobic and anerobic bottles

24. “The patient is already on antibiotics, would resin-based media help to recover the organism?”
Approximately 28-62% of patients are already on antibiotics when the first blood culture is obtained!!!
In patients overall, resin media are superior to media without resins
The difference is greatest in those patients already on antibiotics…and too many patients are on antibiotics when blood cultures are obtained!
Therefore, best practice would be to use resin media routinely!

25. Checklist for Blood Cultures before Leaving the Patient’s Bedside
As with all specimens submitted to the laboratory, blood culture vials should be labeled with the appropriate identification information showing
date and time of collection,
the identification of the patient (barcode),
All bottles or tubes and requisitions are labeled with collector’s employee name, number, or code.
The specific site of collection (which vein, which arm, etc.) is recorded for each set of bottles or tubes obtained
Examples of such a code are as follows:
AL, arterial line;
DIALL, dialysis catheter left; and
MEDL, mediport left

26. Transport to the Laboratory and Handling and Moving within the Laboratory
Timing. Blood should be transported as quickly as possible to the laboratory, preferably within 2 h, and should be placed into the incubator as quickly as possible.
Temperature. Blood culture should never be refrigerated or allowed to cool. Keeping the bottles warm (no warmer than 37°C) is preferable to leaving them at room temperature.
Safety for transport. Blood culture bottles should be carried in some sort of container that will protect them from dropping and from knocking against each other.

27. Rejection Criteria
Staff who receive blood cultures in the laboratory should check the bottles and requisitions carefully to detect a number of problems or errors;
Depending on laboratory and hospital guidelines, specimens with improper labeling may need to be rejected
Bottles with no labels are usually rejected at all times
Patient identification data on the culture bottles and requisition must match, and mismatched specimens may have to be rejected
Whenever rejection is being considered, the physician and/or nursing unit must be notified immediately so they can recollect the samples or discuss the options with a laboratory director or supervisor.

28. Length of Incubation of Blood Cultures
In routine circumstances, using automated continuous monitoring systems, blood cultures need not be incubated for longer than 5 days
For laboratories using manual blood culture systems, 7 days should be sufficient.
99.5% of nonendocarditis BSIs and 100% of endocarditis episodes were detected within 5 days of incubation
Extended incubation periods recommended in the past for Brucella, Capnocytophaga, Campylobacter spp. and the HACEK group (Haemophilus, Actinobacillus, Cardiobacterium, Eikenella, and Kingella spp.) are not necessary for the new continious monitoring systems.

29. BLOOD CULTURE
AEROBIC
ANAEROBIC
TELEPHONE REPORT
GRAM STAINING
GRAM STAINING
BLOOD AGAR
CHOCOLATE AGAR
McCONKEY AGAR
BLOOD AGAR
ID+ADT
A recent study has shown that a telephone report of a positive blood culture with Gram stain results has a greater influence on antimicrobial therapy.

30. Interpretation of Culture Results
Time to detection of positive cultures
Most positive cultures detected in first 48 hours of incubation
S. aureus detected in <24 hours; other staph >24 hours
Culture routinely held 5-7 days
Spectrum of organisms recovered in blood cultures
10-15% of cultures typically positive
Most common isolates are: CNS, S. aureus, E. coli, Enterococcus, Klebsiella, S. pneumoniae
Significant vs. contaminated culture
Most isolates of S. aureus, S. pneumoniae, ß-hemolytic streptococci, Enterococcus, Enterobacteriaceae, Ps. aeruginosa, gram-neg. anaerobes, and yeasts are significant
Most isolates of CNS, Corynebacterium, Proprionibacterium, and Bacillus are insignificant
Significant isolates of CNS are typically associated with a contaminated line or other foreign body.

31. Reasons for Negative BC in Sepsis Patients
Infection is contained locally
Host defenses are containing the infection at the 1°site
Poor timing of collection
A potential issue with intermittent bacteremias
Too low blood volume collected
A frequent issue
Patient on antibiotics
An increasingly important issue
Not loaded on time

32. Contaminants versus Pathogens
Some organisms more associated with colonization
E.g. coagulase negative staphylococci, diphtheroids
Others are always / mostly pathogenic E.g. E.coli, MRSA, Pseudomonas spp., Acinetobacter spp., Legionella, Shigella, Listeria
But most fall in between so:
Consider the body site
Look to see if there are multiple specimens with same organism
Look at the gram stain …….

33. False-Positive Blood Cultures (“Contaminants”)
Contamination
True pathogen
Contamination is “a single culture positive for coagulase-negative staphylococci or coryneform gram-positive rods or Micrococcus or Propionibacterium, or a single bottle with Bacillus species, not anthracis.”
These isolates are also perfectly capable of causing serious infections, catheter- associated septicemia.
Differentiating “contaminants” from true pathogens is extremely difficult, especially because the same species can easily be found in either situation.
Obviously, this can be accomplished most effectively by paying strict attention to the process of skin antisepsis, venipuncture, and specimen transfer to blood culture bottles.

34. What is an “acceptable” blood culture contamination rate*?
Mike
Berkeris LG, JA Toworek, MK Walsh, PN Valenstein. Trends in Blood Culture Contamination.
Arch Pathol Lab Med 129: , 2005 34

35. How to calculate the contamination rate;
Should we calculate it?
Yes
What parameters should we use to calculate it?
The ratio of positive results
Rate of real positivity
Rate of contamination
How should we calculate?
A LOW CONTAMINATION RATE ATTRIBUTES TO THE SUCCESS OF NURSES
Contaminated isolates
Total number of blood cultures

36. False-Positive Blood Cultures (“Contaminants”)
If, a potential contaminant is recovered from one or both bottles and without a second blood culture for comparison, interpretation of the clinical relevance of that positive culture is impossible.
Physicians may be forced to initiate treatment for practical and legal reasons if susceptibilities are reported by the laboratory.
Therefore, the evaluation of an isolate with low virulence potential recovered from a single blood culture set (one or both bottles) should be limited to the extent to which phenotypically similar but medically important organisms can be safely excluded from the identification.
Routine susceptibility testing is not necessary for suspected contaminants, but all isolates should be saved so that additional studies can be performed if an identical organism is recovered from a subsequent blood culture from the same patient.
At that point, full identification of both isolates along with susceptibility testing should be initiated.

37. QUALITY CONTROL / QUALITY ASSURANCE
CLIA and the Clinical and Laboratory Standards Institute (CLSI; formerly NCCLS) both state that commercially produced blood culture media do not require additional in-laboratory QC testing beyond visual inspection if the manufacturer follows CLSI guidelines.
Users should, however, keep accurate records of lot numbers, dates received and expired, and the product inserts certifying the proper QC by the manufacturer.
Blood culture accessioning, processing, incubating, and all the activities that occur within the laboratory for identification of isolates, susceptibility testing, and results formatting are activities for which QC parameters should be measured and monitored

38. EVALUATION OF BLOOD CULTURE PRACTICES: USE OF EPICENTER DATA
Ahmet Başustaoğlu1, Serap Süzük2, İpek Mumcuoğlu3,
Z. Ceren Karahan4, Dilara Öğünç5, İlknur Kaleli6, Şenol Kurşun3,
Ebru Evren4, Betil Özhak Baysal5, Melek Demir6, Nermin Hamurcu7
Patrick Murray8
Girne American University, Girne, Northern Cyprus
MOH. Public Health Agency of Turkey, Ankara, Turkey
Ankara Numune Training and Research Hospital, Ankara,Turkey
Ankara Uni. Faculty of Medicine İbni Sina Training and Research Hosp., Ankara, Turkey
Akdeniz Uni. Faculty of Medicine , Antalya, Turkey
Pamukkale Uni. Hospital, Denizli, Turkey
BD Diagnostic Systems , Istanbul, Turkey
BD Diagnostic Systems , Maryland , ABD.

39. EVALUATION OF BLOOD CULTURE APPLICATIONS BY EPICENTER DATA

40. The aim of the study :
To use the data obtained from proper blood culture practices for;
Demonstrating of the traceability of the data via the statistical analysis program of the EpiCenter operating system
Revealing the efficacy of proper blood culture practices on correct diagnosis through the data obtained and offer areas for improvement within this process
Raising awareness about the use of this program.

41. (Aerobe/Anaerobe/Mycotic)
Hospital
Annual Usage
(Aerobe/Anaerobe/Mycotic)
No. Of beds
2013
2014
2015
Akdeniz University Faculty of Medicine Hospital
18.953
28.497
29.786
983
Pamukkale University Faculty of Medicine Hospital
10.433
12.184
10.397
800
Ankara University İbni Sina Faculty of Medicine Hospital
9.174
5.877
9466
900
MOH. Ankara Numune Hospital
Before BD
13.513
17.163
1140
Total
38.560
60.071
66.812

43. Data entered to EpiCenter
Usage area of the data
Data Flow*
Source of the Data
Patient’s ID No.
Monitoring all data for the patient +
LIS
Patient’s ward
Distribution of data as per ward
- / +
Sample type
Classification as per right-left arm differentiation, catheter and non-blood samples
Time of culture collection
Evaluation of the process from culture collection to the laboratory
Delivery time to laboratory
LIS Manuel
Time of loading to the system
From the Bactec
Number of single vial collected
In the control of the compliance to proper blood culture collection techniques
Number of sets collected
Vial distribution on the basis of set (aerobic-anaerobic, mycotic, etc.)

44. Important points that arouse upon the analysis of EpiCenter data;
Preanalytical phase:
Data requested on EpiCenter that can support hospital quality systems/practices such as
the name of the clinic,
the name of the requesting physican
the name of the person that collected the sample,
the patient’s room number
which are important with respect to monitoring hospital infections and epidemiology have not been entered into the HIS and not transferred to the system.
Times of sample collection for blood culture were mostly not entered into the HIS and/or not written on the request form and consequently were not transferred to the EpiCenter database.

45. able to be taken from EpiCenter Area of usage of statistics
Data
able to be taken from EpiCenter
Area of usage of statistics
Data Flow*
Source of the data
Contamination rate
Driving the right treatment and/or retest according to the proper blood culture collection practices
- / +
Manual
Aerobe/Anaerobe bottle first positivity time (h) (on the basis of set)
Assessment/interpretation of duration of aerobe/anaerobe bottles to become positive on the basis of set and isolate +
From the Bactec device
Distribution of bacteria from aerobe/anaerobe bottles (on the basis of set)
Used in determining the contribution of set usage to the isolation
From the Bactec/Phoenix device
Rate of consistency between culture result and gram staining
Evaluation of the accuracy of the information flow to the clinic that is driving the treatment
Daily distribution of time for bottle loading, positivity and removing
Efficacy of rapid transportation-rapid loading on positivity by revealing the difference between the times of sample collection and sample loading

46. Adopting of appropriate sample collection techniques through necessary trainings ensures drop in contamination rate for blood cultures and increases the chance of obtaining correct results. .
Contamination rates for the hospitals that were included in this study were (6,2- 8,9%),
The reasons behind not being able to calculate contamination rates were ;
the site of blood collection was not specified,
more than one vial/set was not taken,
mistakes in the interpretation of the isolated agent were made
there was a lack of clinic-laboratory cooperation
Awareness should be raised in hospitals of the importance of monitoring these rates

47. Number of bottle/set loaded to the device and rate of positivity

48. Tüm hastanelerdeki kullanım ortalaması alındığında 2013 yılında, 24 saatte hastadan alınan kan kültür şişe sayısının 1 şişe olması durumunda Tablo 6’da görüldüğü gibi pozitiflik %24 iken 4-5 şişe kan kültür alınan hasta grubunda % 46’ya, yılında % 26’dan % 51’e 2015 yılında ise %23’den % 47’ye çıkmıştır

49. Loading times

50. Unloading times

52. Times of sets/bottles loading to the device, detecting positivity and removal of positive bottles within 24 hours.

53. Anand Kumar, et. All., Chest 2009; 136;1237-1248;
Each hour of delay in initiation of effective antimicrobials can increase mortality rates by 7.6%.
Anand Kumar, et. All., Chest 2009; 136; ;