A summary review provided by the

1
Bacterial Contamination in Blood ProductsRisks,
Prevention and Detection

• A summary review provided by the
• American Red Cross Blood Services Regions
• serving the North Atlantic Area
• July, 2003

2
Bacterial Contamination
in Blood Products

• Agenda
• What is the Problem?
• What are the Risks?
• What Organisms are Associated with Bacterial
  Contamination?
• What are the Sources of Contamination?
• What Corrective Actions are Planned?

3
Recent Advances in Testing Technology1990-2003

• Anti-HCV (1990)
• Multi-antigen anti-HCV (1992)
• Anti-HIV1/2, replacing anti-HIV-1 (1992)
• HIV-1 p24 antigen (1996)
• HCV/HIV NAT (IND) (1999)
• Licensed NAT (2003)
• West Nile Virus (IND) (2003)

4
Comparison of Residual Risks
1100
Transmission risk, per unit
HIV
11000
Bacterial Contamination (platelets)
HBV
110 000
Clinical Sepsis (platelets)
HCV
Septic Fatalities (platelets)
1100 000
11 000 000
1996
1994
1992
1990
1988
1986
1984
1998
2000
2002
Updated from Goodnough LT e t al. NEJM
1999341126-7
5
Bacterial Contamination of
Blood Products

• First recognized infectious risk of blood
  transfusion
• Risk greatly reduced in the 1960s by the use of
  closed, sterile systems for the collection and
  storage of blood
• Recent dramatic improvements in safety from viral
  screening and testing have reduced the risks from
  Hepatitis and HIV
• Bacterial sepsis is now the most common
  infectious disease event following transfusion

6
Bacterial Contamination of Blood Products

• Bacterial contamination occurs primarily in
  room-temperature stored products (platelets) but
  can occur in red blood cells and plasma also
• The blood banking community is taking steps to
  improve prevention and detection of bacterial
  contamination
• The American Association of Blood Banks, as well
  as the College of American Pathologists have
  established compliance criteria for transfusion
  services

7
Bacterial Contamination in Blood Products

• The American Association of Blood Banks has
  issued two new standards (March, 2003)
• 5.1.5.1 The blood bank or transfusion service
  shall have methods to limit and detect bacterial
  contamination in all platelet components.
• 5.1.5.1.1 Standard 5.1.5.1 shall be implemented
  by March 1, 2004
• 5.6.2 The venipuncture site shall be prepared so
  as to minimize the risk of bacterial
  contamination. Green soap shall not be used

8
Bacterial Contamination in
Blood Products

• College of American Pathologists Accreditation
  Checklist (December, 2002)
• TRM.44955 Phase 1 Does the laboratory have a
  system to detect the presence of bacteria in
  platelet components?

9
Bacterial Contamination of
Blood Products

• What are the Risks?

10
Risk of clinical sepsis
Pooling issues

• Ness et al, Transfusion 200141857-60.
• Identified clinical cases of transfusion
  associated sepsis over a 12 year period, with
  conversion from 51.7 random donor platelets to
  99.4 SDPs
• The donors/septic event remained constant at
  15,000 throughout the 12 year period, despite the
  conversion to SDPs
• Pooled random donor platelets were 5.5-times more
  likely to cause sepsis than SDPs due to pool size

11
Bacterial Contamination of Blood Products

• What Bacterial Organisms are associated with
  Blood Product Contamination?

12
Bacterial species in platelets implicated in
clinical sepsis
Compilation of data from Clin Micro Rev 1994
7290-302 Transfusion 2001411493-99
www.shot.demon.co.uk/toc
n 86
13
Bacterial species in platelets implicated in
septic fatalities reported to the FDA (1976-1998)
n 52
14
Differences between the species implicated in
septic morbidity and mortality in platelet
components

• S. epidermidis is less commonly observed in
  septic fatalities and more commonly observed in
  septic reactions
• Klebsiella is commonly observed in septic
  fatalities
• Gram negative organisms are implicated in more
  fatalities (60) than gram positive organisms
  (40) gram positives cause a majority of septic
  reactions (56)

15
Organisms implicated in sepsis
from platelets

• Approximately 30 are associated with normal skin
  flora
• Approximately 56 are gram positive
• All are aerobic or facultative anaerobes
• A rare (single case) exception Clostridium
  perfringens fatality from a pooled platelet unit
  Trans Med 1998819-22

16
Bacterial Contamination in Blood Products

• What are the Sources of Bacterial Contamination?

17
Sources of Bacterial Contamination

• Skin Surface Contamination
• Phlebotomy Core
• Donor Bacteremia
• Containers and Disposables
• Environment

18
Skin source
Avoiding Skin Contamination

• Diversion of the initial blood flow
• Improvement in pre-phlebotomy skin cleansing

19
Diversion of initial blood flow

• Diversion of initial blood flow into sampling
  tubes
• Reduces the load of skin-associated bacteria
  entering blood container
• Phlebotomy core directed into sampling pouch
  instead of blood container

20
Clinical data supporting diversion of initial
blood flow

• de Korte et al. Vox Sang 20028313-16
• Collected blood normally or diverted the first
  10mL of whole blood into a satellite bag
• Performed bacterial testing by automated blood
  culture (BacT/Alert) in a laminar flow hood

21
Skin disinfection methods

• Some agents may reduce the number of surface
  bacteria more than others
• Method of application and applicator may have
  some impact on the extent of reduction of surface
  bacteria
• Minimum scrub of 30 seconds required to be
  effective

22
Impact of Skin Disinfection on surface bacteria
63
Goldman et al, Transfusion 199737309-12
23
Recurrent contamination from the dimpled skin of
one plateletpheresis donor

• Anderson et al., Am J Med 1986405-11.
• One donor gave 17 plateletpheresis donations from
  a scarred dimpled site in the right antecubital
  fossa
• Two units were implicated in septic events traced
  to this donor
• Four units, including the two units linked to the
  septic event, were culture positive with
  coagulase negative Staphylococcus
• Follow up blood samples obtained from the
  non-scarred left antecubital fossa were routinely
  culture negative

24
Recurrent contamination from an asymptomatic
bacteremic donor
Donor bacteremia

• Rhame et al., Ann Intern Med 197378633-41.
• One plateletpheresis donor was linked to 7 cases
  of Salmonella cholerae-suis transfusion
  associated bacterial sepsis 2 cases were fatal
• Three of the cases were linked to positive
  culture of the platelet units
• The donor had a low-grade bacteremia and
  unknowingly had Salmonella osteomyelitis of the
  tibia

25
Multiple cases of sepsis from
contaminated blood containers
Container source

• Heltberg et al. Transfusion 199333221-7
• Högman et al. Transfusion 199333189-91.
• Serratia marcescens was cultured from three
  septic patients and their implicated units in
  Denmark
• All units were collected using the same lot of
  blood containers
• 11 of 1,515 blood products collected using the
  implicated lot were positive for Serratia
  marcescens
• An organism of the same ribotype was isolated
  from the manufacturing plant
• The same containers were implicated in Sweden

26
A fatal case of Clostridium perfringens sepsis
from a platelet pool
Environmental source

• McDonald et al., Transfusion Medicine
  1998819-22.
• C. Perfringens is a spore forming facultative
  anaerobe, found in soil and human intestinal
  tract
• Organism recovered from platelet pool septic
  recipient was on antibiotics no organism
  recovered
• Patients death was considered a septic event
• The same serotype of Clostridium was isolated
  from the arm of 1 of the 4 donors a subsequent
  culture of the same arm 6 months later yielded
  fecal flora
• The donor was a mother who carried her two
  toddlers in the crook of her arm

27
Bacterial Contamination in
Blood Products

• What Options exist to Prevent and Detect
  Bacterial Contamination?

28
Bacterial Contamination of PlateletsPrevention
and Detection Options

• Donor screening not feasible except for arm
  screening. Cant detect asymptomatic bacteremic
  donors
• Arm Preparation-Limited effectiveness of arm
  scrub
• Pathogen reduction not yet available. May not
  inactivate spore forming organisms
• Better phlebotomy methods and initial blood
  diversion
• Bacterial detection offers best confirmatory
  option

29
Bacterial Detection Options in Platelet Products

• Visual examination for discoloration, clumping or
  abnormal morphology
• Microscopy
• Gram stain
• Acridine orange
• Measuring Biochemical changes
• Lowered pH
• Reduced Glucose
• Bacterial culture
• Detection through oxygen consumption
• Detection through CO2 production

30
Bacterial Detection Options in Platelet Products

• Visual Examination
• Inspect product prior to transfusion for
  discoloration or abnormal clumping
• Perform swirl procedure to detect morphologic
  changes in platelets
• Normal shaped platelets will align with fluid
  flow and shimmer when swirled
• Contaminated platelets, among others, lose
  discoid shape and do not shimmer when swirled
  Not a specific marker for contamination

31
Swirling
Alignment with flow
SENSITIVITY 75 SPECIFICITY 95
No alignment with flow
Low pH Metabolic disturbance
Leach MF et al. Vox Sang 199874(suppl 1)1180.
32
Bacterial Detection Options in Platelet Products

• Microscopic Methods
• Gram Stain or Acridine Orange preferred methods
• Limitations
• Must be performed by the Transfusion Service
  prior to product issue for transfusion
• Lack sensitivity with low bacterial load

33
Bacterial Detection Options in
Platelet Products

• Measuring Biochemical Changes
• Measure changes in glucose consumption against a
  control. Variances of gt2 S.D. may indicate
  bacterial contamination
• Dipstick testing
• Limitations
• Both this method and staining methods are
  subjective, require high levels of contamination,
  and must be performed prior to issue by the
  Transfusion Service

34
Detecting Bacteria in Platelets Biochemical
Changes
Glucose, Day 0
-2 SD
Storage Time, d
after Burstain JM et al. Transfusion
199737255-8.
35
Chemical Tests - Dipsticks
Must be performed immediately before issue
because of its relative insensitivity and the
need for high bacterial counts
36
Bacterial Detection Options in Platelet Products

• Blood Culture Methods
• Two methodologies presently approved by FDA for
  Quality Control use
• bioMeriuex BacT/Alert System
• Pall Biomedical BDS System

37
Bacterial Detection Options in Platelet Products

• bioMeriuex BacT/Alert System
• Detects bacterial growth in culture bottles by
  measuring CO2 production
• Automated reader continuously monitors samples
• Sampling interval of gt24 hours post phlebotomy
• Culturing interval of gt24 hours post sampling
  (aerobic and anaerobic cultures)
• Cultures incubate for 5-7 days may identify
  positive cultures post-transfusion
• FDA-Approved for Q.C. purposes only on
  Leukoreduced Apheresis Platelets

38
Practical Application of Culturing in a
Transfusion Service Laboratory Aubuchon, Dartmouth
Experience in first 3 years 3,927 apheresis
units cultured (5 mL into aerobic bottle,
BacT/Alert automated system) 23 initial
positives (0.5) in 28 h (10-69) 14 not
confirmed on repeat culture 5 not able to be
recultured 4 confirmed positives
RATE 1/1,000 units (95 CI to 1/600)
39
Detecting Bacteria in Platelets Detection of
Growth by O2 Consumption Pall BDS system
Measure O2 in headspace
24 h
Limit 19.5
Filter Stops WBCsPlts Passes Bacteria
24 h at 35C
Gas impermeable bag
40
Bacterial Detection Options in Platelet Products

• Pall Biomedical BDS System
• Detects bacterial contamination by measuring O2
  consumption
• Automated reader measures O2 levels in headspace
  of culture pouch
• Sampling interval of gt24-48 hours
• Culture performed for gt24-30 hours
• FDA-Approved for Q.C. on leukoreduced platelet
  concentrates and leukoreduced apheresis platelets

41
Bacterial Detection Options in Platelet Products

• Limitations of Blood Culture Methods
• Early sampling/testing may not detect small
  bacteria per bag. Approved methods require 24-30
  hour wait before sampling
• Two FDA-Approved methods require bacteria to grow
  up after sampling to detectable levels, so
  culture must be done well before planned
  transfusion (Blood Center)
• The two time intervals (collection to sampling
  and sampling to release/transfusion) dominate the
  logistic considerations

42
Bacterial Detection Options in Platelet Products

• Limitations of Blood Culture Methods
• Both options require leukoreduced platelets
• BacT/Alert requires continued culture after
  product release
• Release and recall (BacT/ALERT) or hold to end of
  culture to release (PALL BDS)

43
Bacterial Detection Options in Platelet Products

• Limitations of Blood Culture Methods
• Need to balance the risk of platelet shortages
  versus the risk of platelet contamination
• The two available devices are FDA-Approved for
  Q.C, and not approved as pre-release tests
• Cost
• Probable negative impact on outdates
• Possible extension of platelet storage to seven
  days or pooling/storing whole blood derived
  platelets

44
Bacterial Contamination in Transfusable Blood
Products

• AABB Guidance
• Association Bulletin 03-07 issued May 16, 2003
• Provides guidance for methods to limit
  contamination and to detect contamination

45
AABB Association Bulletin 03-07May 16, 2003

• Methods to Limit Contamination
• Careful phlebotomy No green soap prep
• Iodine based scrub recommended
• Consider phlebotomy diversion sample first
  technologies
• Consider increased use of apheresis platelets

46
AABB Association Bulletin 03-07May 16, 2003

• Methods to Detect Contamination
• Culture methods optimal. Two approved products
  cited. Other culture methods can be validated. No
  label claims allowed
• Due to insensitivity, staining and dipstick
  methods should be used as close in time to issue
  as possible
• Validation of all methods is required
• Swirl procedure useful for inspection but does
  not by itself meet AABB Standard 5.1.5.1

47
Bacterial Contamination in Blood Products

• American Red Cross Bacterial Prevention and
  Detection Strategy
• Implement prevention and detection strategies to
  meet the requirements and timelines of the AABB
  and CAP
• Solicit customer feedback to develop efficient
  and cost-effective implementation strategies
• Keep customers well-informed during the
  pre-implementation period