In Vitro Susceptibility Patterns of Methicillin-Resistant and Parenteral ...

ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, Mar. 1983, p. 450-457 0066-4804/83/030450-08$02.0O/0 Copyright 0 1983, American Society for Microbiology

Vol. 23, No. 3

In Vitro Susceptibility Patterns of Methicillin-Resistant and -Susceptible Staphylococcus aureus Strains in a Population of Parenteral Drug Abusers from 1972 to 1981 NORMAN MARKOWITZ, DONALD J. POHLOD, LOUIS D. SARAVOLATZ,* AND EDWARD L. QUINN Infectious Disease Research Laboratory, Henry Ford Hospital, Detroit, Michigan 48202 and the University of Michigan Medical School, Ann Arbor, Michigan 48109

Received 1 September 1982/Accepted 4 January 1983

Since 1980, infections caused by methicillin-resistant (MR) Staphylococcus aureus have been epidemic among Detroit-area parenteral drug abusers. Because of the increasing importance of this pathogen, in vitro susceptibilities were compared for 39 isolates of MR S. aureus from 1980 to 1981, and for 56 strains of methicillin-susceptible (MS) S. aureus from 1972 to 1981, recovered from drug abusers with community-acquired infections. Agar dilution studies were performed at 35°C, and minimal inhibitory concentrations were determined after incubation for 18 and 48 h. MR S. aureus exhibited cross-resistance to other betalactam antibiotics which frequently required 48 h for expression. MR S. aureus isolates were also resistant to tetracycline, clindamycin, tobramycin, and amikacin. All MR S. aureus isolates investigated synthesized an aminoglycoside 4'nucleotidyltransferase. Emergence of resistance to cefotaxime, tetracycline, and clindamycin was noted among current MS S. aureus isolates. Vancomycin, fusidic acid, trimethoprim/sulfamethoxazole, and rifampin were the most active agents against MR S. aureus and were equally effective against MS S. aureus. Strains of methicillin-resistant (MR) Staphylococcus aureus were first identified in British hospitals in 1960, and they became widespread in Europe and Canada by the middle of the same decade (11, 13, 16, 22, 24, 25, 33, 35, 44). Although there were initial reports of MR S. aureus from the United States in 1960 (18, 36), the recognition of its endemic importance in American hospitals has been recent, with increasing reports of infection caused by MR S. aureus appearing over the past 5 years (2, 8, 10, 20). Past reports of outbreaks involving MR S. aureus have emphasized its importance as a nosocomial pathogen (2, 5, 14, 15, 17, 27, 32, 34, 40). In 1980, this organism emerged among Detroit-area parenteral drug abusers in a community-acquired epidemic (9, 30, 38, 39). Because of the sudden appearance of this pathogen, the severe infections it caused, and the limited therapeutic options available, we compared susceptibility patterns of S. aureus isolated from drug abusers over a 10-year period to 22 antimicrobial agents. MATERAULS AND METHODS Strains. S. aureus isolates were identified on the basis of Gram stain, catalase, DNase, and the tube coagulase tests. All isolates were clinical specimens

obtained from 84 parenteral drug abusers with community-acquired staphylococcal infections (38) who were admitted to Henry Ford Hospital. Studies were performed on 39 MR S. aureus isolates from the period 1980 to 1981 and on 56 strains of methicillin-susceptible (MS) S. aureus, 32 from 1980 to 1981 and 24 from 1972 to 1978. Of the 95 isolates studied, 64 (67.4%) were from blood cultures, 14 (14.7%) were respiratory, and 17 (17.9%) were from cerebrospinal fluid, bone, aortic valves, and abscesses. Eleven patients each had two isolates studied. In these instances, the organisms differed by phage type, susceptibility pattern, or both. Beta-latamase. The chromogenic cephalosporin nitrocefin (Glaxo Co., England) was dissolved in dimethyl sulfoxide, diluted in 0.1 M phosphate buffer, pH 7.0 (45), and pipetted over colonies of S. aureus grown overnight at 35°C on Mueller-Hinton agar (BBL Microbiology Systems, Cockeysville, Md.). S. aureus ATCC 25923 served as a negative control. Bacteriophage typing. Bacteriophage typing in routine test dilutions was performed by the Michigan Department of Public Health under the direction of R. Martin. The phages employed included: group 1, 29, 52, 52A, 79, 80; Group 2, 3A, 3C, 55, 71; Group 3, 6, 42E, 47, 53, 54, 75, 77, 83A, 84, 85; miscellaneous, 81, 94, 95, 187. Antimicrobial agents. Susceptibility testing was done with 22 antimicrobial agents. The suppliers and agents were as follows: Beecham Laboratories, Bristol, Tenn.: flucloxacillin, penicillin G; Bristol Laboratories, Syracuse, N.Y.: amikacin, cloxacillin, dicloxacillin, methicillin, oxacillin, tetracycline; Burroughs

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S. AUREUS IN DRUG ABUSERS

Wellcome Co., Los Angeles, Calif.: timethoprim/sulfamethoxazole; Dow Pharmaceuticals, Indianapolis, Ind.: rifampin; Hoechst-Roussel Pharmaceuticals, Inc., Somerville, N.J.: cefotaxime; Leo Laboratories LTD, Copenhagen, Denmark: sodium fusidate; Eli Lilly & Co., Indianapolis, Ind.: cefamandole, cefazolin, cephalothin, tobramycin, vancomycin; Merck, Sharp and Dohme, West Point, Pa.: N-formimidoyl thienamycin; Schering Corp., Kenilworth, N.J.: gentamicin, netilmicin; The Upjohn Co., La Porte, Ind.: clindamycin; Wyeth Laboratories, Philadelphia, Pa.: nafcillin. Media and susceptibility testing. After incubation for 18 h in tryptic soy broth (Difco), isolates of S. aureus were diluted in Mueller-Hinton broth (BBL) to a density equivalent to a 0.5 McFarland standard. Susceptibility testing was done by the agar dilution method (3), and Mueller-Hinton agar (BBL), pH 7.2 to 7.4, was employed for all determinations. Thymidine phosphorylase (Burroughs Wellcome) was used at 2 IU per ml of agar for trimethoprim/sulfamethoxazole testing. An inoculum of approximately 2 x 10' CFU/ml was delivered to the agar surface by a Steers-Foltz replicator (43). Isolates were incubated at 35°C (47), and the minimum inhibitory concentration (MIC) was read at 18 to 48 h (12, 21-23). S. aureus ATCC 25923 served as a control organism on every plate. Aminoglycoside modifying enzyme assay. Assay for the production of aminoglycoside adenylating, acetylating, and phosphorylating enzymes was done according to the method of Benveniste and Davies (4). Bacterial lysis was achieved with lysostaphin (Sigma Chemical Co., St. Louis, Mo.).

451

patterns varied considerably. An organism des-

ignated

as an MR S. aureus isolate required an MIC of -12.5 ,ug of at least one of the following agents per ml: methicillin, nafcillin, or oxacillin. After incubation for 48 h, every MR S. aureus isolate had an MIC of .12.5 ,ug of all three of these agents per ml. These organisms also exhibited cross-resistance to other beta-lactam antibiotics. This resistance, however, was not uniformly expressed after incubation for 18 h. For cephalothin, 40o of the isolates required MICs -6.2 FLg/ml at 18 h, whereas at 48 h, 87% had MICs in the 25 to 100 ,ug/ml range. Cefamandole displayed a similar pattern. When MICs were read at 18 h, all MR S. aureus isolates appeared susceptible to dicloxacillin and flucloxacillin, and nearly all appeared susceptible to cloxacillin and N-formimidoyl thienamycin. Figure 1 compares 18- versus 48-h MICs for dicloxacillin. At 18 h, all isolates were inhibited by 3.1 ,ug/ml or less, but at 48 h, 74% had MICs .12.5 ,ug/ml. This phenomenon was more dramatically observed with N-formimidoyl thienamycin (Fig. 2). Among the 11 beta-lactam stable agents studied, 100% resistance after 18 h of incubation was noted only with oxacilin and cefazolin. However, 48 h of incubation favored the expression of cross-resistance, referred to as hetero-resistance to beta-lactam antibiotics rather than simply methicillin resistance by Seligman and Hewitt (41). RESULTS All MR S. aureus isolates were resistant to All MR S. aureus isolates and 93% (51 of 55) tetracycline, and 79% (31 of 39) were resistant to of MS S. aureus isolates produced beta-lacta- clindamycin. Among MR S. aureus isolates, 7 of mase. Among the four MS S. aureus strains 8 clindamycin-susceptible isolates required negative for beta-lactamase, three were from MICs c0.2 ,g/ml, whereas 31 of 31 clindamy1972 to 1978, and one was from 1980. These cin-resistant MR S. aureus isolates required organisms were susceptible to penicillin. MICs .50 ,ug/ml. A single phage type (29/52/80) accounted for The aminoglycoside susceptibility profile-was 95% (37 of 39) of MR S. aureus isolates (Table striking in the MR S. aureus strains infecting 1). Of the remaining two strains, one was phage drug abusers. Every isolate was susceptible to type 52 and the other was untypable. Approxi- gentamicin and netilmicin but was resistant to mately 18% of MS S. aureus, regardless of the tobramycin and amikacin. period isolated, were phage type (29/52/80). Regardless of the duration of incubation, MS The results of antimicrobial susceptibility test- S. aureus required MICs -3.1 ,ug of methicillin, ing after 18 h of incubation are shown in Tables 2 nafcillin, and oxacillin per ml (except for one and 3. Although nearly all MR S. aureus isolates isolate with an MIC of 6.2 ,ug of methicillin per were of one phage type, their susceptibility ml at 18 and 48 h). Unlike MR S. aureus isolates,

TABLE 1. Bacteriophage types from S. aureus strains isolated from drug abusers Category of isolates'

MS MR a MS,

No. of isolates

56 39

Methicillin-susceptible

isolated during 1980 to 1981.

29/52/80

No. (%) of each phage type 94/96 thyes

Untypable

10 (17.9) 9 (16.1) 20 (35.7) 17 (30.3) 37 (95.0) 0 1 (2.5) 1 (2.5) S. aureus isolated during 1972 to 1981; MR, methicillin-resistant S. aureus

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ANTIMICROB. AGENTS CHEMOTHER.

TABLE 2. MICs, after 18 h of incubation at 35°C, of 18 antimicrobial agents against S. aureus

Category of isolatesa

No. of

isolates

MIC (>Lg/nI)b 90%

50%

Range

Methicillin

MS 12 3.1 3.1 1.5-6.2 6.2->50 MR 24 25 >50 NafcilUin MS 55 0.4 0.8 0.2-1.5 MR 39 12.5 3.1-50 50 55 MS 0.1-3.1 Oxacillin 0.4 0.8 MR 38 25 12.5->50 50 51 MS Cloxacillin 0.4 0.8 0.2-0.8 MR 0.4-12.5 38 1.5 3.1 MS 0.2-0.8 51 0.4 0.4 Dicloxacillin MR 0.2-3.1 38 0.8 1.5 MS 0.2-0.8 51 0.4 0.8 Flucloxacillin MR 38 0.4-3.1 0.8 1.5 MS 53 50 >50 Penicillin G 50 MR 39 >50 25->50 >50 51 0.4 MS 0.8 0.1-0.8 Cephalothin MR 38 12.5 25 0.8-50 51 0.8 1.5 MS Cefazolin 0.2-3.1 MR 38 50 >50 25->50 51 0.8 1.5 MS 0.2-1.5 Cefamandole 12.5 MR 38 12.5 1.5-12.5 MS 51 3.1 12.5 Cefotaxime 0.8-25 MR 38 50 6.2->50 >50 51 0.05 MS s0.025 s0.025-0.05 N-formimidoyl thienamycin MR 38 0.4 0.8 S0.025-25 6.2 53 >50 MS 0.8->50 Tetracycline MR 39 >50 >50 >50 56 0.1 6.2 MS s0.025->50 Clindamycin MR 39 >50 >50 0.1->50 0.4 56 0.8 sO.025-12.5 MS Gentamicin 0.4 0.4 MR 39 0.05-0.8 38 0.2 0.4 0.05-0.8 MS Netilmicin 34 MR 0.2 0.4 0.05-0.8 56 0.2 0.8 MS s0.025->50 Tobramycin MR 39 >50 >50 >50 56 3.1 6.2 0.2-25 MS Amikacin 12.5 39 50 6.2-50 MR a MS, Methicilliti-sensitive S. aureus isolated during 1972 to 1981; MR, methicillin-resistant S. aureus isolated during 1980 to 1981. b 509o MIC required for 50% inhibition; 90%o, MIC required for 90% inhibition.

which tended to appear as distinct colonies at subinhibitory concentrations, MS S. aureus isolates grew confluently (11, 16, 22, 33, 41, 44). There were no substantial differences between

18- and 48-h readings for MS S. aureus with these agents. For the majority of antibiotics tested, there were no differences in susceptibility when MS S.

TABLE 3. In vitro activity' of vancomycin, fusidic acid, trimethoprim/sulfamethoxazole, and rifampin against MS and MR S. aureus combined, 1972 to 1981 No. of isolates

Antimicrobial agent Antimicrobial agent

50%

MIC

90%

(p.g/ml)b

Range

1.5 0.4-3.1 93 0.8 0.8 0.8-1.5 35 0.8 0.08/1.52 0.08/1.52 0.04/0.76-0.08/1.52 89 Trimethoprim/sulfamethoxazole 0.004->50c 0.004 0.004 93 Rifampin a Results reported are at 35C after 18 h of incubation. MICs at 48 h were unchanged. b MIC for 901% inhibition. MIC for inhibition; c Two isolates of MS S. aureus had MICs >50 All other isolates were inhibited by 50.031 Fg of rifampin per ml.

Vancomycin Fusidic acid

50o%o

50%o

90%o,

p.g/ml.

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1001

A

80" '0 0) ._

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60' 40-

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H I

20-

80-

60 40-

-

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453

II

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,

20

T

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MIC (pg/mi) FIG. 1. Activity of dicloxacillin against methicillin-resistant S. aureus. In vitro susceptibility at 35°C after 18 h of incubation (B) is compared with MICs after 48 h of incubation (A). Solid black bars represent the percentage of isolates inhibited at a given MIC. Stippled bars added to the solid bars represent the cumulative percentage of isolates inhibited at a given MIC.

aureus isolates from 1972 to 1978 were compared with MS S. aureus isolates from 1980 to 1981. However, with cefazolin, cefotaxime, tetracycline, and clindamycin, a broader distribution of MICs was observed among current MS S. aureus isolates compared with those from 1972

100' 80' 60' .0 ._

'._

to 1978. Whereas all organisms from the earlier period required MICs of cefazolin of 0.2 to 0.8 ,ug/ml, 30% of MS S. aureus isolates from 1980 to 1981 required MICs of 1.5 to 3.1 ,ug/ml. With the third generation cephalosporin cefotaxime, more important differences were noted. Al-

A

40' 20'

o 0.

0

0

80' 60' 40'

20' 50

MIC (pg/mi) FIG. 2. Activity of N-formimidoyl thienamycin against methicillin-resistant S. aureus. In vitro susceptibility at 35C after 18 h of incubation (B) is compared with MICs after 48 h of incubation (A). Solid black bars represent the percentage of isolates inhibited at a given MIC. Stippled bars added to the solid bars represent the cumulative percentage of isolates inhibited at a given MIC.

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though all MS S. aureus isolates from 1972 to 1978 were inhibited by 3.1 ,ug or less of cefotaxime per ml, 25% of current isolates were relatively resistant (6 of 30 required MICs of 12.5 ,ug/ml, and 2 of 30 required MICs of 25 jig/ml). Before 1978, 85% of strains required MICs s3.1 p,g of tetracycline per ml; MICs -6.2 ,ug/ml were found in 75% of MS S. aureus isolates from 1980 to 1981. Likewise, clindamycin resistance emerged among current MS S. aureus strains, with 15% requiring >50 ,ug/ml to inhibit growth. For vancomycin, fusidic acid, trimethoprim/ sulfamethoxazole, and rifampin (Table 3), in vitro susceptibilities were similar for MR and MS S. aureus. Since MICs were essentially unchanged after incubation for 48 h, the results shown represent 18-h readings. All strains were inhibited over a narrow range of dilutions and were susceptible to all four agents, except for two MR S. aureus isolates that were resistant to rifampin. The latter were recovered from two patients with rifampin-susceptible MR S. aureus isolated from blood cultures obtained before antimicrobial therapy; however, rifampin resistance emerged after 3 and 4 weeks of combination therapy with vancomycin and rifampin, respectively. Every MR S. aureus isolate studied and one isolate of MS S. aureus from 1980 to 1981, phage type (29/52/80), showed gentamicin-netilmicin susceptibility and tobramycin-amikacin resistance. An assay for aminoglycoside modifying enzymes was performed on 13 MR and the one MS S. aureus isolate displaying this aminoglycoside profile. All 14 organisms produced an aminoglycoside 4'-nucleotidyltransferase (29). Acetylating and phosphorylating enzymes were not detected. DISCUSSION The first parenteral drug abuser with an MR S. aureus infection was admitted to Henry Ford Hopsital in March 1980. This patient had tricuspid valve endocarditis with the epidemic phage type (29/52/80). Currently, 50% of all staphylococcal infections in this population are caused by MR S. aureus. A review of epidemiology records and isolates of S. aureus obtained before March 1980 failed to disclose organisms resistant to oxacillin among drug abusers. However, when these MS S. aureus strains isolated before 1980 were compared with recent ones, resistance to agents other than the semisynthetic penicillins emerged. Formerly, all S. aureus isolates from this population were susceptible to clindamycin, and in vivo experience was quite favorable (6). Currently, 20% of MS S. aureus strains are resistant to this agent. Moreover, one patient with osteo-

ANTIMICROB. AGENTS CHEMOTHER.

myelitis and clindamycin-susceptible MS S. aureus isolated by bone biopsy failed to improve after 2 weeks of therapy with this agent. Another patient with endocarditis caused by a methicillin-resistant organism had multiple blood isolates of S. aureus differing only in susceptibility to clindamycin. One strain required a MIC of 0.1 ,ug of clindamycin per ml; the other strain had a MIC of >50 p.g/ml. Otherwise, these strains were identical with respect to antibiotic profile, production of an aminoglycoside 4'-nucleotidyltransferase, and bacteriophage type. Such experiences have made us reluctant to use clindamycin as an anti-staphylococcal therapy in drug abusers. Isolates of MR S. aureus expressed resistance to multiple agents, including other beta-lactams, tetracycline, clindamycin, tobramycin, and amikacin. In agar dilution studies, all MR S. aureus isolates were resistant to oxacillin and cefazolin after 18 h of incubation at 35°C, and all MS S. aureus isolates were susceptible, suggesting that these antibiotics are the most reliable in screening tests. In vitro susceptibility testing failed to demonstrate nafcillin resistance in 12 of 39 oxacillinresistant isolates after 18 h of incubation. The MICs of nafcillin against these organisms tended to be about eightfold higher than those seen for MS S. aureus, but resistance was only expressed when incubation was extended to 48 h. Although the clinical significance of such strains is not precisely known (26), caution must be exercised in classifying organisms with borderline susceptibilities as MS S. aureus (22), and in vitro susceptibilities should be performed under conditions favorable to the phenotypic expression of methicillin resistance. Techniques employed by other workers include high inoculum, temperatures of 30 to 35°C, prolonged incubation time, and media containing 5% NaCl (11, 12, 16, 21, 22, 25, 26, 37, 41, 44, 47). MIC determinations for cephalothin against MR S. aureus were unreliable. Even after 48 h of incubation, 5 of 38 MR S. aureus isolates were cephalothin susceptible. The use of cephalosporins in this setting has been associated with a high incidence of bacteriological and clinical failure (1). Canawati et al. (7) have recently shown that cephalosporin resistance is more clearly evident with incubation at 30°C. As observed by Hewitt et al. (22), both low temperature and prolonged incubation time may be required to detect resistance to cloxacillin. This is consistent with our observations for cloxacillin, dicloxacillin, and flucloxacillin. We have also noted this for N-formimidoyl thienamycin. Wise et al. (50) reported five strains of MR S. aureus to be susceptible to this agent by agar dilution testing. However, these studies

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performed at 37°C and were read after 18 h of incubation. Likewise, Verbist and Verhaegen (48) reported 11 strains of oxacillin-resistant S. aureus inhibited by 0.12 to 2.0 jig of N-formimidoyl thienamycin per ml after 18 h at 36°C. Kropp et al. (28) described one strain of MR S. aureus resistant to this agent (MIC = 20.0 ,jg/ml) under routine test conditions (37°C, 24 h of incubation). Only one of the MR S. aureus isolates that we studied had MICs :12.5 ,ug/ml at 18 h. Recently, Thompson et al. (46) found that 90% of 43 MR S. aureus isolates were inhibited by c8 ,g of N-formimidoyl thienamycin per ml after 18 h of incubation at 35°C. Similar results were also obtained by Witte et al. (51) for 82 isolates of MR S. aureus, even at a temperature of 30°C. Both reports note a disparity between MICs for MS and MR S. aureus, the latter being somewhat higher. This is in agreement with our data. Perhaps prolonged incubation is a more sensitive method for detecting resistance to this agent. We did not determine the concentration of Nformimidoyl thienamycin in agar after 48 h. However, neither the MS S. aureus strains nor the control organisms grew after the longer incubation time. Furthermore, MR S. aureus appeared as isolated colonies at subinhibitory concentrations, suggesting subpopulations of staphylococci heterogeneous in their resistance to this agent (11). The use of thienamycin for the therapy of MR S. aureus infections must be approached with the same caution reserved for other beta-lactam antibiotics. All MR S. aureus isolates investigated produced an aminoglycoside 4'-nucleotidyltransferase capable of adenylating the 4'-hydroxyl group of tobramycin, amikacin, and kanamycin. Gentamicin and netilmicin lack this group and showed in vitro activity against all MR S. aureus isolates. Plasmid-mediated synthesis of this enzyme has been previously described by Le Goffic et al. (29). It is noteworthy that one methicillin-susceptible strain of the epidemic phage type, resistant to tobramycin, amikacin, and kanamycin, also produced this enzyme. Thus, methicillin resistance was not uniformly linked with the production of an aminoglycoside adenylating enzyme. Perhaps this represents an undetected subpopulation of MR S. aureus among a large majority of methicillin-sensitive organisms, the dissemination of a plasmid among strains of staphylococci, or a loss of expression of methicillin resistance. Vancomycin, fusidic acid, trimethoprim/sulfamethoxazole, and rifampin were the most active antimicrobial agents in vitro against methicillinresistant isolates and were equally active against methicillin-sensitive strains. In two patients with MR S. aureus infections, rifampin resistance

were


455

emerged while the patients were receiving combination therapy with vancomycin and rifampin. The initial blood isolates before therapy required MICs (,ug/ml) of rifampin of 0.001 and 0.004, respectively. Minimal bactericidal concentrations were 0.004 and 0.125 ,ug/ml, respectively. The resistant organisms were recovered from bone and aortic valve vegetations. It is disconcerting that resistance to rifampin may emerge even when this agent is used with a second effective antimicrobial agent. Another area of concern for the clinical application of the combination of vancomycin and rifampin is the antagonism that Watanakunakorn and Guerriero observed in many isolates (49). With the persistence of MR S. aureus as a community-acquired pathogen in drug abusers and the dissemination amnong non-abusers in the Detroit area (38, 39), alternative therapies need to be identified. Although vancomycin is the treatment of choice (31, 42), clinical failure with this agent has been reported (19, 42). Based on the in vitro studies reported in this paper, fusidic acid, trimethoprim/sulfamethoxazole, and rifampin, in combination with other agents, may warrant clinical evaluation (42). ACKNOWLEDGMENTS We thank Kenneth Price and David Bobey of Bristol Laboratories, who performed the aminoglycoside modifying enzyme assay, and Margaret Somerville, R. Martin, Larry Ross, Eugene Mezger, Patricia Cornett, and Geri Sanders. LITERATURE CITED 1. Acar, J. F., P. Courvaln, and Y. A. Chabbert. 1971. Methicillin-resistant staphylococcemia: bacteriological failure of treatment with cephalosporins, p. 280-285. Antimicrob. Agents Chemother. 1970. 2. Barrett, F. F., R. F. McGhee, and F. Finland. 1968. Methicillin-resistant Staphylococcus aureus at Boston City Hospital. Bacteriologic and epidemiologic observations. N. Engl. J. Med. 279:441-448. 3. Barry, A. L. 1976. The antimicrobic susceptibility test: principles and practice, p. 236. Lea & Febiger, Philadelphia, Pa. 4. Benveniste, R., and J. Davies. 1971. R-factor mediated gentamicin resistance: a new enzyme which modifies aminoglycoside antibiotics. FEBS Lett. 14:293-296. 5. Boyce, J. M., M. Landry, T. R. Deetz, and H. L. Dupont. 1981. Epidemiologic studies of an outbreak of nosocomial methicillin-resistant Staphylococcus aureus infections. Infect. Control 2:40-46. 6. Burch, K. H., E. L. Quinn, F. Cox, T. Madhavan, E. Fisher, and D. Roinlg. 1976. Intramuscular clindamycin for therapy of infective endocarditis. Report of 23 cases and review of the literature. Am. J. Cardiol. 38:929-933. 7. Canawati, H. N., J. L. Witte, and F. L. Sapico. 1982. Temperature effect on the susceptibility of methicillinresistant Staphylococcus aureus to four different cephalosporins. Antimicrob. Agents Chemother. 21:173-175. 8. Centers for Disease Control. 1981. Methicillin-resistant Staphylococcus-United States. Morbid. Mortal. Weekly Rep. 30:140, 145-147. 9. Centers for Disease Control. 1981. Community-acquired methicillin-resistant Staphylcoccus aureus infectionsMichigan. Morbid. Mortal. Weekly Rep. 30:185-187. 10. Centers for Disease Control. 1981. Methicillin-resistant Staphylococcus aureus-United States. Morbid. Mortal.

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doyl-thienamycin, a novel 3-lactam: an in vitro comparison with other P-lactam antibiotics. J. Antimicrob. Chemother. 7:521-529. 51. Wltte, J. L., F. L. Sapico, and H. N. Canawatl. 1982. In


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