- Email: [email protected]
A comparative study of blood and bone marrow cultures in cadaveric bone donation
M
journal of Hospital Infection ( 1999) 43: 305-308
A comparative study of blood and bone marrow cultures in cadaveric bone donation S. B. W. Vehmeyeflt, F?L. C. Petits
R. M. Bloem$$, R. L. M. DeijkerQ,
M. R.Veen$$,
*B/S Foundation, Plesmanlaan I 00,230 I CH Leiden, The Netherlands, f Department of Surgery, Westeinde Hospital, Lijnbaan 32, ‘s-Gravenhage, The Netherlands, # Department of Orthopaedic Surgery, Leiden University Medical Center,Albinusdreef2,2333 Z4 Leiden,The Netherlands, §Netherlands Bone Bank Foundation, Plesmanlaan I 00,230 I CH Leiden, The Netherlands, Department of Microbiology, She/and Hospital, Burg. Knappertlaan 253 I I6 B/3 Schiedam, The Netherlands.
Summary: The results of blood cultures taken from cadaveric bone donors were compared with bone marrow and also swab cultures of the procured grafts. In eight of the 95 donors evaluated, pathogenic micro-organisms were detected in the blood sample. In two, identical micro-organisms were cultured from the blood and bone marrow sample whilst the swab cultures were negative. Considering the low sensitivity of the swab culture, the organisms detected in the blood culture were likely to have spread haematogenously and considered to be present in the explanted grafts. Bacteriological screening of bone donors is extremely important since the transmission of microorganisms via an allograft can lead to serious complications in the recipient. Positive blood cultures provide important information on the presence of pathogenic micro-organisms in grafts obtained from cadaveric bone donors and are therefore essential in deciding whether a graft is bacteriologically acceptable for transplantation. 0 I999The
Keywords: Bone donation;
cadaver;
blood cultures;
most serious complication assouse of bone allografts.lW3 Careful aseptic donor procurement and screening of donors are essential these complications. Although
Received 6 January 1999; revised manuscript accepted 19 August 1999 Address for correspondence: S.B.W. Vehmeyer, BIS Foundation, PO. Box 2304, 2301 CH Leiden, The Netherlands, Tel.: 31-71-5795710 Fax: 31-71-5790903 E-mail: [email protected]
0195-6701/99/120305+04$12.00
Infection
Society
microbiology.
Introduction Infection is the ciated with the donor selection, bacteriological in preventing
Hospital
standards issued by the American Association of Tissue Banks (AATB),4 the European Association of Tissue Banks (EATB) and the European Association for MusculoSkeletal Transplantation (EAMST)S offer guidelines for tissue banks on most of these aspects, guidelines on the interpretation of the microbiological test results of a donor are not included. Such interpretation is often difficult. It is unclear whether positive cultures are caused by contamination during procurement, or whether they are the result of a haematogenous spread of micro-organisms preor post-mortem. 0 I999 The Hospital
Infection
Society
S. B.W. Vehmeyer
306
In a period of 26 months, bone marrow samples were aseptically obtained from the iliac crest prior to every procurement procedure. The samples were cultured for aerobic and anaerobic micro-organisms for five days in blood culture media. The results of the blood and bone marrow cultures, if positive, were compared with the results of the swab cultures. For the interpretation of the culture results a differentiation was made between micro-organisms of low pathogenicity including skin contaminants such as coagulase negative staphylococci and Propionibacterium acnes and micro-organisms generally considered of higher potential pathogenicity, including Staphylococcus aureus, Streptococcus spp and Candida spp.’
Although other culture methods have been described,” most tissue banks rely on swab cultures taken from the external surface of every graft. The aim of this study was to show that swab cultures alone provide insufficient information for the microbiological assessment of bone grafts, and that blood culture results should always be taken into account when interpreting the microbiological test results of a donor.
Materials
and methods
Bone and soft tissue were retrieved from cadaveric donors that met the standard selection criteria issued by the EAMST and the EATB.’ Exclusion criteria included all signs of clinical infections. Procurement was performed in operating rooms using aseptic techniques. Prior to the procedure, blood samples for blood cultures were taken under aseptic conditions through a cardiac puncture. The blood cultures (Bactec, Becton Dickinson Microbiological systems, Cockeysville, Maryland USA) were cultured aerobically and anaerobically for five days. During the procurement procedure every obtained graft was rinsed with a sterile saline solution. Subsequently the entire bone surface was carefully swabbed using a 15 cm polyestertipped applicator (Becton Dickinson), which was then put in transport medium (Port-A-Cul, Becton Dickinson). The swabs were inoculated within 24 h on to blood agar and chocolate plates and cultured under aerobic and anaerobic conditions for 48 h. The swab sticks themselves were cultured in a Brain-heart infusion culture broth (Oxoid Unipath Ltd., Basingstoke, Hampshire, UK) which was subcultured after 48 h. Table
I
Results Samples from 95 donors were used in this study, and the results of the blood cultures versus the bone marrow cultures are listed in Table I. In 65 donors (68%) the blood and bone marrow cultures were negative. Organisms of “low” pathogenicity were detected in the cultures of 19 donors (20%). In only one of these donors, the blood and bone marrow culture yielded an identical isolate, P acnes. Pathogenic micro-organisms were detected in the cultures of 11 donors (12%). In three of these, both the blood and bone marrow cultures were positive (Table I I). In nine donors, identical micro-organisms were retrieved from the swab culture of one of the grafts as well as from the blood or the bone marrow sample. In eight of these, coagulase negative staphylococci were found. In these donors, the bone marrow culture was negative.
Results ofblood cultures versus bone marrow cultures
Negative
Bone marrow Low pathogenicity
cultures High pathogenicity
Total
Blood cultures Negative ‘Low’ pathogenicity ‘High’ pathogenicity Total
65 13 5 83
et al.
5 I 0 6
3 0 3 6
73 I4 8 95
Culture
of cadaveric
Table
307
bone
II
Pathogenic micro-organisms in blood and bone marrow cultures
Donor
Blood cultures
Bone marrow
I 2 3 4 5
Negative Negative Negative K. oxyloca Viridaus streptococci, E cloacae 5. sanguis Viridans streptococci Viridans streptococci 5. haemolyticus, Neisseria sp Viridans streptococci S. agaloctiae Viridans streptococci, S. aureus Gram negative cocci*
C. albicans E. fiecalis S. oureus Negative
6 7 8 9 IO II
*= could not be cultured
cultures
Negative Negative Negative Negative Viridans streptococci S. agalactiae Gram negative rods*
for determination
In one donor, S. aureus was detected in the bone marrow sample as well as in the swab culture of the proximal humerus, whilst the blood culture was negative.
Discussion Micro-organisms, of either “low” or “high” pathogenicity, can be detected in the tissue cultures of bone allografts.6j8-‘0 Moreover, Veen et al. found a contamination rate of 92%, mainly ” Organisms originate of low pathogenicity. from (i) the procurement personnel; (ii) the skin of the donor; (iii) the environment or (iv) an endogenous source. The presence of organisms of low pathogenicity is mainly associated with external factors.” The only predisposing factor for blood cultures with micro-organisms of high pathogenicity found by Deijkers et al., was a traumatic cause of death.’ Martinez found a similar correlation with gunshot injuries.” Other tissue banks have reported high rates of pathogenic micro-organisms of associated with procurement of a donor in a mortuary.6,sa’0 But as all donors in the study of Deijkers et al. and this study were procured in operating theatres using standard aseptic techsource for these niques, the most probable organisms is endogenous. The absence of signs
of clinical infection in the donors with positive cultures seems to imply a haematogenous spread of such organisms, pre- or post-mortem. In eight donors (8%), the blood culture was micro-organisms, positive with pathogenic comparable to the study of Martinez et a1.l’ No comparable organisms were cultured from any of the grafts obtained from these donors. In two donors, identical micro-organisms, viridans steptococci and S. agalactiae, were cultured and in a third donor, mixed flora were detected in the blood sample and Gram-negative rods were cultured from the bone marrow sample (Table I I). Although we can not exclude the possibility of contamination of the samples of these three donors, an endogenous source seems most likely. Of these three donors, only one had a traumatic cause of death, which could explain the mixed flora detected in the blood and bone marrow samples, indicating mucosal damage. In the second donor, from which S. agalactiae were cultured, a non-heart-beating nephrectomy was performed, prior to the bone procureIn this case, a catheter was ment procedure. placed in the right femoral artery shortly after circulatory arrest, and could have acted as a possible source. In the third donor, from which viridans streptococci were cultured from the blood and bone marrow, no predisposing factors could be found.
S. B.W. Vehmeyer
Pathogenic organisms were detected in the bone marrow culture of six donors in three of whom the blood culture was negative, whilst all swab cultures were negative. The organisms isolated from bone marrow alone were C. albicaus, E. faecalis and S. aureus Whilst there results might be indicative of the importance of bone marrow cultures, we feel, however, they are not convincing enough to justify a recommendation to routinely perform bone marrow cultures. In two donors, identical pathogenic organisms were found in the blood sample as well as in the bone marrow sample, whilst the swab cultures, from the external surface, were negative. This confirms the limited sensitivity of swab and also cultures as found by Veen et al.,” stresses the importance of blood cultures should a tissue bank rely on swab cultures alone, For some surgical procedures it would be preferable to use large unprocessed bone allografts of which the essential characteristics are still intact. To ensure the safety of these grafts, careful donor selection, vigorous virological screening and microbiological surveillance are of extreme importance. The Netherlands BoneBank Foundation provides unprocessed allografts for limited indications, of which the swab cultures and, most importantly, the blood cultures should always be negative. If the swab culture of a graft or the blood culture of a donor is positive with pathogenic micro-organisms, all grafts of that donor are pre-treated with 15 kGy (k 3 kGy) gamma-irradiation before being processed (cleaned and rinsed with water, antibiotics and ethanol) and released for transplantation purposes.
3.
4. 5.
6.
7.
8.
9.
10.
11.
References 1. Lord CF, Gebhardt Mankin HJ. Infection
2.
MC, in
Tomford WW, bone allografts.
et ol.
Incidence, nature, and treatment. J Bone Joint Surg Am 1988; 70: 369-376. Mankin HJ, Gebhardt MC, Jennings LC, Springfield DS, Tomford WW. Long-term results of allograft replacement in the management of bone tumors. Clin Orthop 1996; 324: 86-97. Tomford WW, Thongphasuk J, Mankin HJ, Ferraro MJ. Frozen musculoskeletal allografts. A study of the clinical incidence and causes of infection associated with their use. r Bone Joint Surg Am 1990; 72: 1137-l 143. AATB. Standards for tissue bunking: AATB, 1996. EAMST, EATB. Common standards for musculo skeletal tissue banking. Vienna: European Association for Musculo Skeletal Transplantation and European Association of Tissue Banks, 1997. Farrington M, Matthews I, Foreman J, Richardson KM, Caffrey E. Microbiological monitoring of bone grafts: two years’ experience at a tissue bank. J Hasp Infect 1998; 38: 261-271. Veen MR. Bone allografts: a study into bacterial contamination, sensitivity of cultures, decontamination, and contribution to postoperative infection (Thesis). University of Leiden, 1994. Bettin D, Harms C, Polster J, Niemeyer T. High incidence of pathogenic microorganisms in bone allografts explanted in the morgue. Acta Orthop &and 1998; 69: 311-314. Deijkers RLM, Bloem RM, Petit PLC, Brand R, Vehmeyer SBW, Veen MR. Contamination of bone allografts: analysis of incidence and predisposing factors. J Bone Joint Surg Br 1997; 79: 161-l 66. Martinez OV, Malinin TI, Valla PH, Flores A. Postmortem bacteriology of cadaver tissue donors: an evaluation of blood cultures as an index of tissue sterility. Diagn Microbial Infect Dis 1985; 3: 193-200. Veen MR, Bloem RM, Petit PLC. Sensitivity and negative predictive value of swab cultures in musculoskeletal allograft procurement. Clin Orthop 1994; 300: 259-263.
journal of Hospital Infection ( 1999) 43: 305-308
A comparative study of blood and bone marrow cultures in cadaveric bone donation S. B. W. Vehmeyeflt, F?L. C. Petits
R. M. Bloem$$, R. L. M. DeijkerQ,
M. R.Veen$$,
*B/S Foundation, Plesmanlaan I 00,230 I CH Leiden, The Netherlands, f Department of Surgery, Westeinde Hospital, Lijnbaan 32, ‘s-Gravenhage, The Netherlands, # Department of Orthopaedic Surgery, Leiden University Medical Center,Albinusdreef2,2333 Z4 Leiden,The Netherlands, §Netherlands Bone Bank Foundation, Plesmanlaan I 00,230 I CH Leiden, The Netherlands, Department of Microbiology, She/and Hospital, Burg. Knappertlaan 253 I I6 B/3 Schiedam, The Netherlands.
Summary: The results of blood cultures taken from cadaveric bone donors were compared with bone marrow and also swab cultures of the procured grafts. In eight of the 95 donors evaluated, pathogenic micro-organisms were detected in the blood sample. In two, identical micro-organisms were cultured from the blood and bone marrow sample whilst the swab cultures were negative. Considering the low sensitivity of the swab culture, the organisms detected in the blood culture were likely to have spread haematogenously and considered to be present in the explanted grafts. Bacteriological screening of bone donors is extremely important since the transmission of microorganisms via an allograft can lead to serious complications in the recipient. Positive blood cultures provide important information on the presence of pathogenic micro-organisms in grafts obtained from cadaveric bone donors and are therefore essential in deciding whether a graft is bacteriologically acceptable for transplantation. 0 I999The
Keywords: Bone donation;
cadaver;
blood cultures;
most serious complication assouse of bone allografts.lW3 Careful aseptic donor procurement and screening of donors are essential these complications. Although
Received 6 January 1999; revised manuscript accepted 19 August 1999 Address for correspondence: S.B.W. Vehmeyer, BIS Foundation, PO. Box 2304, 2301 CH Leiden, The Netherlands, Tel.: 31-71-5795710 Fax: 31-71-5790903 E-mail: [email protected]
0195-6701/99/120305+04$12.00
Infection
Society
microbiology.
Introduction Infection is the ciated with the donor selection, bacteriological in preventing
Hospital
standards issued by the American Association of Tissue Banks (AATB),4 the European Association of Tissue Banks (EATB) and the European Association for MusculoSkeletal Transplantation (EAMST)S offer guidelines for tissue banks on most of these aspects, guidelines on the interpretation of the microbiological test results of a donor are not included. Such interpretation is often difficult. It is unclear whether positive cultures are caused by contamination during procurement, or whether they are the result of a haematogenous spread of micro-organisms preor post-mortem. 0 I999 The Hospital
Infection
Society
S. B.W. Vehmeyer
306
In a period of 26 months, bone marrow samples were aseptically obtained from the iliac crest prior to every procurement procedure. The samples were cultured for aerobic and anaerobic micro-organisms for five days in blood culture media. The results of the blood and bone marrow cultures, if positive, were compared with the results of the swab cultures. For the interpretation of the culture results a differentiation was made between micro-organisms of low pathogenicity including skin contaminants such as coagulase negative staphylococci and Propionibacterium acnes and micro-organisms generally considered of higher potential pathogenicity, including Staphylococcus aureus, Streptococcus spp and Candida spp.’
Although other culture methods have been described,” most tissue banks rely on swab cultures taken from the external surface of every graft. The aim of this study was to show that swab cultures alone provide insufficient information for the microbiological assessment of bone grafts, and that blood culture results should always be taken into account when interpreting the microbiological test results of a donor.
Materials
and methods
Bone and soft tissue were retrieved from cadaveric donors that met the standard selection criteria issued by the EAMST and the EATB.’ Exclusion criteria included all signs of clinical infections. Procurement was performed in operating rooms using aseptic techniques. Prior to the procedure, blood samples for blood cultures were taken under aseptic conditions through a cardiac puncture. The blood cultures (Bactec, Becton Dickinson Microbiological systems, Cockeysville, Maryland USA) were cultured aerobically and anaerobically for five days. During the procurement procedure every obtained graft was rinsed with a sterile saline solution. Subsequently the entire bone surface was carefully swabbed using a 15 cm polyestertipped applicator (Becton Dickinson), which was then put in transport medium (Port-A-Cul, Becton Dickinson). The swabs were inoculated within 24 h on to blood agar and chocolate plates and cultured under aerobic and anaerobic conditions for 48 h. The swab sticks themselves were cultured in a Brain-heart infusion culture broth (Oxoid Unipath Ltd., Basingstoke, Hampshire, UK) which was subcultured after 48 h. Table
I
Results Samples from 95 donors were used in this study, and the results of the blood cultures versus the bone marrow cultures are listed in Table I. In 65 donors (68%) the blood and bone marrow cultures were negative. Organisms of “low” pathogenicity were detected in the cultures of 19 donors (20%). In only one of these donors, the blood and bone marrow culture yielded an identical isolate, P acnes. Pathogenic micro-organisms were detected in the cultures of 11 donors (12%). In three of these, both the blood and bone marrow cultures were positive (Table I I). In nine donors, identical micro-organisms were retrieved from the swab culture of one of the grafts as well as from the blood or the bone marrow sample. In eight of these, coagulase negative staphylococci were found. In these donors, the bone marrow culture was negative.
Results ofblood cultures versus bone marrow cultures
Negative
Bone marrow Low pathogenicity
cultures High pathogenicity
Total
Blood cultures Negative ‘Low’ pathogenicity ‘High’ pathogenicity Total
65 13 5 83
et al.
5 I 0 6
3 0 3 6
73 I4 8 95
Culture
of cadaveric
Table
307
bone
II
Pathogenic micro-organisms in blood and bone marrow cultures
Donor
Blood cultures
Bone marrow
I 2 3 4 5
Negative Negative Negative K. oxyloca Viridaus streptococci, E cloacae 5. sanguis Viridans streptococci Viridans streptococci 5. haemolyticus, Neisseria sp Viridans streptococci S. agaloctiae Viridans streptococci, S. aureus Gram negative cocci*
C. albicans E. fiecalis S. oureus Negative
6 7 8 9 IO II
*= could not be cultured
cultures
Negative Negative Negative Negative Viridans streptococci S. agalactiae Gram negative rods*
for determination
In one donor, S. aureus was detected in the bone marrow sample as well as in the swab culture of the proximal humerus, whilst the blood culture was negative.
Discussion Micro-organisms, of either “low” or “high” pathogenicity, can be detected in the tissue cultures of bone allografts.6j8-‘0 Moreover, Veen et al. found a contamination rate of 92%, mainly ” Organisms originate of low pathogenicity. from (i) the procurement personnel; (ii) the skin of the donor; (iii) the environment or (iv) an endogenous source. The presence of organisms of low pathogenicity is mainly associated with external factors.” The only predisposing factor for blood cultures with micro-organisms of high pathogenicity found by Deijkers et al., was a traumatic cause of death.’ Martinez found a similar correlation with gunshot injuries.” Other tissue banks have reported high rates of pathogenic micro-organisms of associated with procurement of a donor in a mortuary.6,sa’0 But as all donors in the study of Deijkers et al. and this study were procured in operating theatres using standard aseptic techsource for these niques, the most probable organisms is endogenous. The absence of signs
of clinical infection in the donors with positive cultures seems to imply a haematogenous spread of such organisms, pre- or post-mortem. In eight donors (8%), the blood culture was micro-organisms, positive with pathogenic comparable to the study of Martinez et a1.l’ No comparable organisms were cultured from any of the grafts obtained from these donors. In two donors, identical micro-organisms, viridans steptococci and S. agalactiae, were cultured and in a third donor, mixed flora were detected in the blood sample and Gram-negative rods were cultured from the bone marrow sample (Table I I). Although we can not exclude the possibility of contamination of the samples of these three donors, an endogenous source seems most likely. Of these three donors, only one had a traumatic cause of death, which could explain the mixed flora detected in the blood and bone marrow samples, indicating mucosal damage. In the second donor, from which S. agalactiae were cultured, a non-heart-beating nephrectomy was performed, prior to the bone procureIn this case, a catheter was ment procedure. placed in the right femoral artery shortly after circulatory arrest, and could have acted as a possible source. In the third donor, from which viridans streptococci were cultured from the blood and bone marrow, no predisposing factors could be found.
S. B.W. Vehmeyer
Pathogenic organisms were detected in the bone marrow culture of six donors in three of whom the blood culture was negative, whilst all swab cultures were negative. The organisms isolated from bone marrow alone were C. albicaus, E. faecalis and S. aureus Whilst there results might be indicative of the importance of bone marrow cultures, we feel, however, they are not convincing enough to justify a recommendation to routinely perform bone marrow cultures. In two donors, identical pathogenic organisms were found in the blood sample as well as in the bone marrow sample, whilst the swab cultures, from the external surface, were negative. This confirms the limited sensitivity of swab and also cultures as found by Veen et al.,” stresses the importance of blood cultures should a tissue bank rely on swab cultures alone, For some surgical procedures it would be preferable to use large unprocessed bone allografts of which the essential characteristics are still intact. To ensure the safety of these grafts, careful donor selection, vigorous virological screening and microbiological surveillance are of extreme importance. The Netherlands BoneBank Foundation provides unprocessed allografts for limited indications, of which the swab cultures and, most importantly, the blood cultures should always be negative. If the swab culture of a graft or the blood culture of a donor is positive with pathogenic micro-organisms, all grafts of that donor are pre-treated with 15 kGy (k 3 kGy) gamma-irradiation before being processed (cleaned and rinsed with water, antibiotics and ethanol) and released for transplantation purposes.
3.
4. 5.
6.
7.
8.
9.
10.
11.
References 1. Lord CF, Gebhardt Mankin HJ. Infection
2.
MC, in
Tomford WW, bone allografts.
et ol.
Incidence, nature, and treatment. J Bone Joint Surg Am 1988; 70: 369-376. Mankin HJ, Gebhardt MC, Jennings LC, Springfield DS, Tomford WW. Long-term results of allograft replacement in the management of bone tumors. Clin Orthop 1996; 324: 86-97. Tomford WW, Thongphasuk J, Mankin HJ, Ferraro MJ. Frozen musculoskeletal allografts. A study of the clinical incidence and causes of infection associated with their use. r Bone Joint Surg Am 1990; 72: 1137-l 143. AATB. Standards for tissue bunking: AATB, 1996. EAMST, EATB. Common standards for musculo skeletal tissue banking. Vienna: European Association for Musculo Skeletal Transplantation and European Association of Tissue Banks, 1997. Farrington M, Matthews I, Foreman J, Richardson KM, Caffrey E. Microbiological monitoring of bone grafts: two years’ experience at a tissue bank. J Hasp Infect 1998; 38: 261-271. Veen MR. Bone allografts: a study into bacterial contamination, sensitivity of cultures, decontamination, and contribution to postoperative infection (Thesis). University of Leiden, 1994. Bettin D, Harms C, Polster J, Niemeyer T. High incidence of pathogenic microorganisms in bone allografts explanted in the morgue. Acta Orthop &and 1998; 69: 311-314. Deijkers RLM, Bloem RM, Petit PLC, Brand R, Vehmeyer SBW, Veen MR. Contamination of bone allografts: analysis of incidence and predisposing factors. J Bone Joint Surg Br 1997; 79: 161-l 66. Martinez OV, Malinin TI, Valla PH, Flores A. Postmortem bacteriology of cadaver tissue donors: an evaluation of blood cultures as an index of tissue sterility. Diagn Microbial Infect Dis 1985; 3: 193-200. Veen MR, Bloem RM, Petit PLC. Sensitivity and negative predictive value of swab cultures in musculoskeletal allograft procurement. Clin Orthop 1994; 300: 259-263.