APPLIED MICROBIOLOGY, Feb. 1971, p. 209-216Copyright ( 1971 American Society for Microbiology

Vol. 21, No. 2Printed in U.S.A.

Laminar Airflow Protection In Bone MarrowTransplantation

CLAUS 0. SOLBERG,' JOHN M. MATSEN, DONALD VESLEY, DONALD J. WHEELER,ROBERT A. GOOD, AND HILAIRE J. MEUWISSEN

School of Public Health, Department of Laboratory Medicine, and the Pediatric Research Laboratories of theVariety Club Heart Hospital, University of Minnesota, Minneapolis, Minnesota 55455

Received for publication 14 August 1970

A laminar airflow room was used to provide a low-pathogen environment fora child with lymphopenic immune deficiency transplanted with paternal bonemarrow. Comparison of flora from the patient, personnel, and the environmentindicated that no colonization with exogenous organisms occurred in the patientduring the 45-day period of study. The number of organisms recovered from thelaminar airflow room was exceedingly small. Conventional hospital isolation roomscontained more bacteria and fungi than the laminar airflow room, even when strictaseptic procedures were followed in the former. Patients with lymphopenic immunedeficiency and agranulocytosis admitted to conventional isolation rooms were

colonized with exogenous organisms within 1 week. Each developed infection withthese strains, and one patient died. Laminar airflow isolation seems at present thebest means to prevent exogenous infection during hospitalization of patients withlymphopenic and other severe immune-deficiency diseases and may be essentialwhen bone marrow transplantation is performed to treat their immunologicaldefect.

Patients who have undergone bone marrowtransplantation (BMTP) for lymphopenic im-mune deficiencies (LID) are exceptionally proneto severe infection (7). Lacking humoral andcellular immunities, they are often susceptible toinfection by organisms normally nonpathogenicto healthy individuals. After BMTP, they mayalso suffer from graft-versus-host (GVH) diseasewhich further depresses already impaired im-munity (2, 8).Laminar airflow (LAF) rooms have recently

been adapted for care of patients with a high riskof infection. When combined with strict aseptictechnique, they provide maximum protectionagainst microbial contamination from the en-vironment (3, 15). However, it has remained un-proven that these rooms can prevent colonizationof patients with environmental organisms.

In the present study, we describe a 1-year-oldboy with LID who received a paternal bonemarrow transplant and subsequently developedGVH disease. He was cared for in an LAF roomunder strict aseptic procedures. Extensive micro-biological monitoring of the patient, medicalpersonnel, and the LAF room was carried out toascertain whether the patient could be protected

lPresent address: University of Bergen, School of Medicine,Medical Department B, Bergen, Norway.

from acquisition of exogenous flora over a pro-tracted time period while undergoing critical-carenursing. Comparative results from the patientsand environment of six conventional isolationand hospital rooms are also presented.These observations establish that a patient with

LID can be cared for successfully in an LAF roomwithout detectable acquisition of environmentalpathogens. In contrast, patients cared for in con-ventional isolation and hospital rooms were con-tinually exposed to a large number of environ-mental pathogens.

In view of the extreme susceptibility of LIDpatients to infection, an effective protectiveenvironment should be used as an adjunct to theclinical management of these patients and otherpatients with immune deficiencies, particularlywhen BMTP is performed to treat their immuno-logical defect.

MATERIALS AND METHODSCase history. The patient was a 12-month-old boy

hospitalized to receive a BMTP for LID, i.e., sex-linked lymphopenic hypogammaglobulinemia, a con-sistently fatal disease. In the patient's family, over thelast three generations, 12 male infants less than 1 yearold had died from repeated infections. In the absenceof siblings, the father was selected as the bone marrowdonor. Three days before transplantation, the patient

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was admitted to the LAF room. Initial bacterialcultures were obtained and a single bath with 70%,0alcohol and a hexachlorophene detergent was given.At this time, pulmonary infection with Ptieumocystiscarinii was suspected, and the patient was treatedwith pentamidine isethionate. Prophylactic anti-microbial therapy was not instituted. To modify thecourse of the expected GVH disease, the bone marrowwas purified by albumin gradient centrifugation (5),antilymphocyte serum was administered before trans-plantation, and methotrexate was given at regularintervals after transplantation.Three weeks after transplantation, GVH disease

developed; 3 weeks later, the patient suddenly de-veloped gram-negative septicemia with severe shockand died within 3 hr. Autopsy revealed extensiveintestinal ulceration and scattered bilateral pneumonia.

Laminar airflow room and conventional isolation andhospital rooms. The principles of LAF rooms, theiroperation, and efficiency have been described (3, 15,16). The present unit was a prototype of a commer-cially available model (kindly provided on loan by theApplied Science Division of the Litton Co., Minneapo-lis, Minn.) designed for ready installation in existinghospital rooms. Figure 1 illustrates the unit. One entirewall was made up of HEPA filters capable of removing99.97%0 of all particles at 0.3 Am. Virtually all bacteria,fungi, and protozoa are filtered out at this efficiency,and virus particles which may exist as individualparticles below 0.3,um are also removed by virtue ofBrownian movement.The unit was designed to operate at airflow rates of

30, 60, or 90 ft/min. The highest velocity resulted inapproximately 400 air changes per hour and was usedfor maximum activity situations. The crib was orientedperpendicular to the airflow at the filter wall of the

Buffer ZoneArea

FIG. 1. Laminar airflow room.

room so that all activities took place downstream fromthe patient.

All materials entering the facility, including food,were presterilized by steam autoclaving or ethyleneoxide gasclaving. Certain items, such as thermometerswhich could not be heated above 100 F (ca. 38 C),were sterilized by a 24-hr exposure to ethylene oxideat room temperature. All items were double wrappedbefore sterilization.

All personnel entering the facility performed a3-min hand scrub with 3(O hexachlorophene emulsionand dressed in special uniforms in a 3 by 3 ft entryway("buffer zone area") which was marked off by floortapes at the room opening. The uniforms consisted ofspecial presterilized hoods, laboratory coats, and calf-length boots, all made from Bar-Bac cloth (AngelicaUniform Co., St. Louis, Mo.); conventional pre-sterilized disposable masks and surgical gloves wereworn. Room furnishings and floor surfaces werecleaned thoroughly once daily by the nurses using 3 to4 sterile, single-use cleaning cloths and a phenolicdetergent-disinfectant solution (15). The whole pro-cedure lasted about 30 min.

For purposes of comparison, non-LAF hospitalrooms, with mechanical ventilation of less than fiveair changes per hour, were included in the study. Inthree hospital isolation rooms, the isolation and house-keeping procedures were the same as in the LAF roomexcept that the food and equipment were not sterilized.The patients occupying these rooms were 0.5, 3, and10 years old and suffered from LID, meningo-myelocele, and agranulocytosis, respectively. All werecompletely confined to bed during their stay in thehospital. None had open wounds.

Less rigid isolation procedures were followed inthree other single-bed hospital rooms. Persons en-tering these rooms used sterile gowns and masks.Bed clothes were changed every morning and therooms were cleaned with a phenolic detergent-disin-fectant. The patients occupying these rooms were 3,22, and 43 years old and suffered from purulentmeningitis, paraplegia, and cancer of the colon,respectively. They were all confined to bed duringtheir stay in the hospital.

Microbiological methods. Specimens from patientsand personnel were obtained, usually at 4- and 7-dayintervals, respectively, with cotton swabs moistenedwith sterile saline. The technique has previously beendescribed (17). Nose and throat swabs were streakedon 5% sheep blood-agar (SBA) plates, 5% rabbitblood-agar plates [both having Trypticase Soy Agarbase (BBL)], and on Levine's eosin-methylene blue(EMB) agar plates (BBL). Skin swabs were inoculatedon 5%O SBA and EMB plates and on plates containing5% sheep blood in Columbia selective agar (CSA,BBL).A quantitative platinum loop (0.001 ml) was used

for streaking urine specimens onto 5%o SBA and ontoEMB plates. Stools were inoculated on 5% SBA,EMB, and CSA plates; a 5% SBA plate and a platecontaining phenylethyl alcohol in 570 sheep blood-agar were incubated anaerobically.A Saboraud agar (Difco) plate with 2% added

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dextrose and 0.05% chloramphenicol was usedperiodically to detect the presence of yeast.

Organisms from sources other than urine wereevaluated by using a semiquantitative rating of lessthen 10 colonies (+), 10 to 30 colonies (++),greater than 30 colonies but no growth in the thirdsegment of streaking (+++), and greater than 30colonies and growth in all streaking segments

Rodac plates containing 15.5 ml of medium wereused for surface sampling (4) and Casella slit-samplers,for air sampling. The sampling medium was TrypticaseSoy Agar. For the Rodac plates, a neutralizer system(0.5% Tween 80 and 0.07% soy lecithin) was in-corporated in the medium.

Methods of organism identification follow guide-lines described elsewhere (11). Sensitivity determina-tions were done by using either the twofold serial tubedilution method [employing Mueller-Hinton broth(BBL) and an inoculum of approximately 105 to 106organisms per ml] or the single high-potency disctechnique (1). Serological typing of Escherichia coilwas done through the courtesy of Albert Balows atthe Center for Disease Control, Atlanta. Pyocintyping of Pseudomonas aeruginosa was done byShirley Parker of the University of Manitoba, Winni-peg. Klebsiella strains were serotyped in the laboratoryof one of the authors (JMM; 13). Phage typing ofStaphylococcus aureus was performed as previouslydescribed (17). Identity of bacterial strains was docu-mented by antibacterial spectra and biochemicaltesting in addition to routine cultural methods.Serological, pyocin, and phage typing were performedas indicated above.

RESULTSPatients and medical personnel. On admission

to the hospital, the bone marrow transplant pa-tient was directly transferred to the LAF roomwhere he stayed for 45 days. Samples from hisnose, throat, axillae, groin, perineum, stool, andurine were obtained immediately before he enteredthe LAF room and later usually at 4-day intervals.Table 1 shows the organisms isolated from thepatient's nose, throat, perineum, and stool duringhis stay in this room. Before admission to theLAF room, he was heavily contaminated withgram-negative rods which showed a tendency tobuild up during hospitalization, not only in thenose and throat, but also in the perineum andstool. The more benign organisms, such asdiphtheroids and Neisseria sp., disappeared earlyduring hospitalization. P. aeruginosa was isolatedfrom the nose and throat before the patiententered the isolation room; later, identical strainswere isolated not only from these sites, but alsofrom the perineum and stool. E. coil and Klebsiellasp., identical to the organisms in the stool andperineum, invaded the nose and throat. S.epidermidis was cultured from the axillae before

the patient was placed in the isolation facility.Identical strains were later recovered from thenose, throat, and perineum. All staphylococcalstrains were chai acteristic in being coagulase-negative and exhibited the identical antibiogram,being resistant to both penicillin and methicillin.The urine was sterile. Organisms isolated fromthe groin were identical to those in the perineum.E. coli and P. aeruginosa identical to the organismsisolated from the patient before and during hisentire stay in the LAF room were cultured fromthe blood, before and immediately after his death,and from lung specimens obtained at autopsy.P. carinii organisms were also identified in lungspecimens.

Cultures from the nose and throat of the per-sonnel entering the LAF room were obtained with1-week intervals. During the first 5 weeks, identi-cal strains were not isolated from these individualsand the patient. But later, P. aeruginosa identicalto the patient's strain was isolated from the nasalsamples of one of the nurses. No organisms fromthe environment or from the personnel enteringthe room were cultured from the patient duringhis stay in the LAF room.

Samples from the nose, throat, perineum, stool,and urine of the patients occupying the conven-tional isolation and hospital rooms were ob-tained at 3- to 6-day, usually 4-day, intervals.The patients with LID and agranulocytosis werecolonized with environmental organisms duringthe first week in the hospital and the other pa-tients within less than 3 weeks, the organismsmost frequently isolated being gram-negativerods and Candida albicans. The patients withLID and agranulocyto,.is developed infectionwith these organisms, and the patient withagranulocytosis died of the infection.Laminar airflow room and conventional isolation

and hospital rooms. Table 2 shows the microbialcontamination of surfaces and furnishings in theLAF room and the other hospital rooms 4 to 6 hrafter the beds were made and the rooms cleaned.The blankets and pillows in the conventionalisolation and hospital rooms had 5 to 10 timesmore bacterial and fungal contaminants, and thefloors, walls, and furniture usually 100 to 300times more bacterial and fungal contaminantsthan did the LAF room.The microbial air contamination was measured

before, during, and after bedmaking (Fig. 2).Four experiments were performed in the LAFroom and two experiments in each of the otherhospital rooms. In the conventional hospitalrooms, the mean air contamination increasedfrom 9 colonies/ft3 before bedmaking to 33colonies/ft3 during bedmaking, and then de-

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SOLBERG ET AL. APPL. MICROBIOL.

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LAMINAR AIRFLOW PROTECTION IN HOSPITALS

TABLE 2. Microbial contaminatioln on surfaces and furnishings in laminlar airflow room anzd coniventionalisolation anid hospital rooms

Item

Pillows andblankets ....

Floors.........Walls..........Tables and

shelves......Chair..........Lamp..........

No. ofRodacplates

per expt

10154

1544

Colonies per Rodac plate

Laminar airflow room Conventional isolation Conventional hospitalrooms rooms

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Time rmnutes,FIG. 2. Air contamination in laminar airflow room

and conventional isolation and hospital rooms measuredby slit-samplers placed 3 ft inside the room entrances.Curve for laminar airflow room is meant offour experi-ments, othe-s are mean of two experiments.

creased to about 15 colonies/ft3 10 min lter. Themean air contamination in the conventionalisolation rooms was about half that in the con-ventional hospital rooms. In the LAF room, very

few organisms were isolated from the air duringthe entire period.

Table 3 gives the mean values of the microbialair contamination for all experiments in the threedifferent types of rooms. The air contained 170and 370 times more bacterial and fungal or-ganisms per cubic foot in the conventional isola-tion and hospital rooms, respectively, than in theLAF room.

In the LAF room, all of the 118 colonies in theair samples, the samples from floors, walls, andfurniture, and 50 randomly chosen colonies in thesamples from the bed clothing were identified.For each experiment in the other six rooms, 25,15, and 10 randomly chosen colonies in the airsamples, samples from the floors, walls, andfurniture, and from the bed clothing, respectively,were identified (Table 4). Far more total andpathogenic organisms (Tables 2 and 3), especiallyenteric and fungal strains, were isolated from theconventional isolation and hospital rooms thanfrom the LAF room. Strains isolated were com-pared with organisms isolated simultaneouslyfrom the patients in each of these rooms. Themajority of organisms isolated from blankets andpillows were identical to organisms isolated fromthe patient inhabitant of the bed (Table 5). Thiswas true to a lesser extent when floors, walls,tables, etc., and the room air were sampled. Inthe LAF room, 48 or 50 colonies in the samplesfrom the bed clothing were identical with thepatient's organisms. The frequency of patientidentical strains in air samples and in samplesfrom the floors, walls, and furniture was alsohigher in the LAF room than in the other rooms.

DISCUSSION

Patients with LID are remarkably susceptibleto infection because they lack immunological

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defense of cellular and humoral types (7). Oftenthey become chronically infected with organismsnormally nonpathogenic to healthy individuals,e.g., C. albicans (7), and may die from systemicvaricella, rubeola, BCG, and vaccinia infections(7, 9). The majority of LID patients die duringtheir first year of life, and few, if any, reach theage of 2 years. Our experience with numerousLID patients indicates that hospitalization, evenunder strict conventional isolation procedures,

TABLE 3. Air conitaminiationi in laminiar airflowroom and coniventioiial isolation

and hospital rooms

Colonies/cubic foot

Laminar airflow roomaExpt (downstream) at Conventional Conventional

airflow velocity of isolation hospitalroomsb roomsb

90 ft/min 60 ft/min

1 0.047 0.050 8.16 15.902 0.039 0.068 9.61 21.56

Mean 0.043 0.059 8.89 18.73

a Total of 660 ft3 sampled.b Mean of three experiments, one in each of

three different rooms. Total of 696 ft3 sampled.

frequently results in colonization with resistanthospital organisms, especially gram-negativebacilli (P. aeruginosa, Klebsiella sp., E. coli),fungi (Candida sp.) and P. carinii which oftencause sepsis and death. In addition, if GVHdisease develops after BMTP as in our patient,LID patients become even more susceptible toinfection, not only with endogenous flora, butalso with hospital organisms. Therefore, thecapability of LAF rooms to prevent colonization

TABLE 5. Percenitage of patientt idenitical strainis insamples from environimenit of lamintar airflow

room and conivenitionial isolatioln anldhospital roomsa

Floors,Blankets walls,

Room and tables, Airpillows lamp,

chair

Laminar airflow room 96 (50) 52 (61 ) 46 (57)Conventional isola-

tion rooms......... 78 (60) 36 (90) 33 (150)Conventional hospitalrooms.............. 67 (60) 20 (90) 29 (150)

a Values in parentheses express numbers ofcolonies examined.

TABLE 4. Identification of enivirontmenital contaminiants from laminiarflow room anid conventiontal isolationand hospital rooms

No. of colonies

Laminar airflow room Conventional isolation Conventional hospitalOrganisms rooms rooms

Blankets Floors, walls, Blankets Floors, walls,' Blankets Floors, wallsBandketllos Ffurniture, and pillows furniture, and pillows furniture,adplos and air ananaan and air

Bacillus sp....................... 16 14 81 10 63Candida sp ......................... 2 4 1 5Clostridium sp1......................Diphtheroids ....................... 2 14 3 28 4 12Enterobacter ........................ 2 2 1Enterococcus....................... 3 3 2 2 3 3E.coli............................. 4 2 1 2 4 6Herellea sp I 1 1 1Klebsiella sp........................ 4 1 2 2 9Lactobacillus sp . 1 2 1Neisseria sp ..9 2 16 2 21Proteus mirabilis.1 1 2 4 1 3Pseudomonas aeruginiosa............. 3 2 1 8 3 2Saccharomyces. 1 2Staphylococcus aureus 2 9 4 11S. epidermidis ...................... 33 68 20 61 22 79Viridans group streptococcus 3 2Unidentified molds 2 4 16 3 20

Totals 50 118 60 240 60 240

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and infection with environmental organisms wasput to a severe test by our patient.LAF systems have been described previously

(3, 15, 16). Air entering the LAF room is virtuallysterile: not only fungi, bacteria, and protozoa(P. carinii) but also viruses are effectively elimi-nated by the filtration technique used. Further-more, if the patient is kept upstream, close to thefilter wall, infectious particles originating frommedical personnel will be carried downstreamaway from the patient by the airflow. Penlandand Perry (16) described the mechanism of theLAF room in detail but reported no microbio-logical data. Bodey et al. (3) treated patients withleukemia in LAF rooms, but in their study noattempts were made to associate the micro-organisms isolated in the LAF room with thosefrom the patients. Therefore, colonization ofpatients by exogenous organisms could not beruled out. Michaelsen et al. (15) performed themost extensive microbiological studies on LAFrooms reported to date. In their investigation,they used normal human volunteers and studiedthe effectiveness of the air purification system.They did not study the bacterial flora of thehuman subjects and did not identify the or-ganisms isolated. Their volunteers stayed in theLAF room for 12 days only and did not need theextensive and complex care that our patient re-ceived.Our patient is the first human to be studied in

an LAF room by extensive microbiological moni-toring for a prolonged period of time (45 days)while under intensive medical care. Our micro-biological studies involved not only repeatedsampling of the air and surfaces of the LAF roombut also extensive monitoring of the patient andmedical personnel entering the room. To deter-mine exogenous colonization of the patient,bacteria were characterized by phage typing,biochemical and serological methods, and anti-biotic-sensitivity determinations.Our observations show that no demonstrable

colonization by new organisms occurred duringthe period of study. In addition, the microbialcontamination of the air (per cubic foot), walls,floor, and furniture of the LAF room was low,and the majority of the organisms were identicalto the bacteria isolated from the patient. The veryfew organisms which could not with certainty betraced back to the patient's flora were usuallyfound far downstream in the LAF room and,therefore, did not constitute a major threat to thepatient. By contrast, a great number of pathogenicbacteria and fungi were isolated from the environ-ment of conventional isolation and hospitalrooms, and, as documented in our study, pa-tients occupying these rooms were readily in-

fected by these org?nisms. Therefore, theserooms appeared unsuited for treatment of LIDpatients and other patients with markedly reducedresistance to infection.

Although the concentration of airborne micro-organisms in terms of colonies per cubic foot was170 to 370 times-greater in the conventional roomsthan in the LAF room in this study (Table 3), thedifferences in total nwnbers of airborne micro-organisms were small because of the greaterventilation used in the LAF room. Thus, at aventilation rate 80 times as great (400 versus 5 airchanges per hour) in the LAF than in the con-ventional rooms, the total number, rather thanthe total concentration, of airborne micro-organisms was only about two to four timesgreater in the conventional rooms.BMTP now offers a chance of cure for patients

with LID, particularly if a histocompatibilitylocus-A (HL-A) identical sibling is available as adonor (6, 14). However, after transplantation themajority of patients undergo GVH reactionswhich markedly enhance susceptibility to infec-tion by depression of the marrow, production oflymphoid atrophy, and initiation of intestinalulcerations. In our patient, measures were takento modify the severity of the GVH reactions,including albumin gradient centrifugation of themarrow (5) and administration of antilympho-cyte serum and methotrexate. Nevertheless,severe GVH disease with multiple intestinalulcerations developed. Disruption of the intestinalbarrier most probably facilitated seeding of gram-negative organisms into the blood and resultedin terminal septicemia. Our experience with bonemarrow transplant patients (Brit. Med. J., inpress) and recent studies of GVH disease inexperimental animals (10, 12) indicate that LIDpatients considered for BMTP should undergosuppression of oropharyngeal and intestinal floraif severe GVH disease is expected. Under thesecircumstances, environmental protection is evenmore imperative since decontamination pro-cedures in the absence of effective isolationfacilities are regularly disastrous. Our patient wasclosely observed by skilled personal during hisentire stay in the LAF room. His microbial florawas monitored continuously and found to befairly susceptible to antibiotics, and intestinalulcerations were not observed while the patientwas alive. Therefore, prophylactic treatment withnonabsorbable oral antibiotics was not considerednecessary. Our policy was immediate institutionof parenteral and oral antibiotic therapy at thefirst signs of infection. However, the terminalepisode of septicemia was so rapid in its onset andso fulminant in its course that antibiotic therapywas of no help. In retrospect, it is felt that pro-

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216 SOLBERG ET AL.

phylactic antibiotic therapy might have preventedthis terminal septic episode. Therefore, in futureBMTP, hospitalization of the patients in LAFrooms under strict isolation procedures and de-contamination of the intestinal flora may beimportant steps in securing survival of the pa-tients.

It should be emphasized that LAF rooms pro-vide no protection against sepsis caused byendogenous organisms. Neither can LAF roomsbe expected to function optimally unless a strictand meticulous aseptic technique is maintained.However, if good isolation procedures are fol-lowed, the LAF rooms constitute a significantadvance in environmental microbial control.Nursing of patients with LID and other immuno-logical deficiency diseases in a LAF environmentseems at present the best means to prevent hospitalinfections in these patients and may be essentialif transplantation with HL-A nonidentical bonemarrow is considered.

ACKNOWLEDGMENTS

This investigation was supported by Public Health Servicegrants lF05-TW-01494-01 from the Fogarty International Center,1-ROI-CA11505-01 from the National Cancer Institute, AI-08677 from the National Institute of Allergy and InfectiousDiseases, and HE-06314 from the National Heart and LungInstitute and grants from the National Foundation-March ofDimes and the American Heart Association.

LITERATURE CITED

1. Bauer, A. W., W. M. M. Kirby, J. C. Sherris, and M.Turck. 1966. Antibiotic susceptibility testing by a standard-ized single-disk method. Amer. J. Clin. Pathol. 45:493-496.

2. Blaese, R. M., C. Martinez, and R. A. Good. 1964. Immuno-logic incompetence of immunologically runted animals.J. Exp. Med. 119:211-225.

3. Bodey, G. P., E. J. Freireich, and E. Frei. 1969. Studies ofpatients in a laminar air flow unit. Cancer 24:972-980.

4. Bond, R. G., M. M. Halbert, K. M. Keenan, H. D. Putnam,0. R. Ruschmeyer, and D. Vesley. 1963. Development of

APPL. MICROBIOL.

a method for microbial sampling of surfaces with specialreference to reliability. Contract PH-86-62-182, Divisionof Hospital and Medical Facilities Bureau of State Services,U.S. Department of Health, Education and Welfare.

5. Dicke, K. A., J. van Hooft, and D. W. van Bekkum. 1968.The selective elimination of immunologically competentcells from bone marrow and lymphatic cell mixtures.Transplantation 6:562-570.

6. Gatti, R. A., H. J. Meuwissen, H. D. Allen, R. Hong, andR. A. Good. 1968. Immunologic reconstitution of sex-linked lymphopenic immunological deficiency. Lancet2:1366-1369.

7. Good, R. A., and D. Bergsmna. 1968. Immunologic deficiencydiseases in man, vol. 4, no. 1. The National Foundation.

8. Howard, J. G., and M. F. A. Woodruff. 1961. Effect of graftversus host reaction on the immunological responsive-ness of the mouse. Proc. Roy. Soc. London Ser. B.154:532-539.

9. Hoyer, J. R., M. D. Cooper, A. E. Gabrielsen, and R. A.Good. 1968. Lymphopenic forms of congenital immuno-logic deficiency diseases. Medicine 47:201-226.

10. Keast, D. 1968. A simple index for the measurement of therunting syndrome and its use in the study of the influenceof the gut flora in its production. Immunology 15:237-245.

11. Laboratory Procedures in Diagnostic Microbiology. 1969.Department of Laboratory Medicine, University of Minne-sota, Minneapolis.

12. Ledney, G. D. 1969. Antilymiiphocyte serum in the therapy andprevention of acute secondary disease in mice. Transplanta-tion 8:127-140.

13. Matsen, J. M., and D. J. Blazevic. 1969. Characterization ofornithine decarboxylase-positive, nonmotile strains of theKlebsiella-Enterobacter group. Appl. Microbiol. 18:566-569.

14. Meuwissen, H. J., R. A. Gatti, P. I. Terasaki, R. Hong, andR. A. Good. 1969. Treatment of lymphopenic hypogam-maglobulinemia and bone marrow aplasia by transplanta-tion of allogeneic marrow. N. Engl. J. Med. 281:691-697.

15. Michaelsen, G. S., M. M. Halbert, S. Sorenson, and D. Vesley.1968. Development of an open isolation system for the careof low resistance patients. Final report under contractPH-43-65-999 from the National Cancer Institute, NationalInstitutes of Health, U.S. Public Health Service.

16. Penland, W. Z., Jr., and S. Perry. 1970. Portable laminar-air-flow isolator. Lancet I:174-176.

17. Solberg, C. 0. 1965. A study of carriers of Staphylococcusaureus with special regard to quantitative bacterial estima-tions. Acta Medica Scand. 178 (Suppl. 436):28-29.

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