DIAGN MICROBIOL INFECT DIS 1 1 9 1987;8:119-122

NOTES

Susceptibi l i ty In Vitro of Nocardia Species to Antimicrobial Agents

P.M. Southern, Jr., A.E. Kutscher, R. Ragsdale, and B. Luttrell

Fifty-four clinical isolates of Nocardia spp. were tested in vitro for susceptibility to several antimicrobial agents. Of these, 89% were susceptible to ciprofloxacin, 86% to imipenem, 85% to fusidic acid, and 71% to cefotaxime. Some of the agents may be suitable alternative or adjunctive drugs to sulfonamides and aminoglycosides for chemotherapy of Nocardial infec- tions.

Infections due to Nocardia species have been seen more frequently in recent years, particularly as pathogens in immunocompromised individuals, irrespective of their underlying condition. The authors have seen Nocardial infection in organ transplant recipients, in patients with lymphoreticular malignancies, in persons receiving cy- totoxic agents and/or corticosteroids, and in persons with the acquired immunode- ficiency syndrome (AIDS). Similar reports have also appeared in recent literature (Bradsher et al., 1982; Curry, 1980; Frazier et al., 1975). For some time the standard of antimicrobial chemotherapy for Nocardial infections was with sulfonamides. More recently other agents have been shown to be effective in vitro, including kanamycin, amikacin, minocycline, doxycycline, and some of the newer beta-lactam agents (Acar et al., 1983; Cynamon and Palmer, 1981; Dewsnup and Wright, 1984; Garcia-Rod- riquez et al., 1982; Gombert, 1982; Gombert and Aulicino, 1983; Gombert et al., 1986; Gutmann et al., 1983; Kitzis et al., 1985; Sugar et al., 1983; Wallace et al., 1983). Many individuals cannot tolerate sulfonamides, and in others the aminoglycosides may be contraindicated because of compromised renal function. Furthermore, there have been reports of failure of therapy with sulfonamides and trimethoprim (Cockerill et al., 1984; Geiseler et al., 1979; Stamm et al., 1983). Smego, McKenzie, and McManus (1987) have recently pointed out the unavailability of parenteral sulfonamides in the United States, and in general emphasized the fact that few agents are available with demonstrated efficacy in vivo against infections due to Nocardia species. Thus it would be of potential clinical value to have additional alternative forms of therapy, particularly with oral agents. We elected to study the activity in vitro of several antimicrobial agents against a variety of clinical isolates of Nocardia species. Some of the agents studied are effective after oral administration.

The organisms used in this study were from three sources (all clinical isolates): 1) the Clinical Microbiology Laboratory of Parkland Memorial Hospital, 2) Dr. Richard

From the Department of Pathology, the University of Texas Health Science Center at Dallas (P.M.S., B.L.), and Parkland Memorial Hospital (P.M.S., A.E.K., R.R.), Dallas, Texas.

Address reprint requests to: Paul M. Southern, Jr., M.D., Department of Pathology, University of Texas Health Science Center at Dallas, 5323 Harry Hines Blvd., Dallas, TX 75235-9072.

Received January 7, 1987; revised and accepted July 2, 1987.

© 1987 Elsevier Science Publishing Co., Inc. 52 Vanderbilt Avenue, New York, NY 10017 0732-8893/87/$03.50

120 P .M. Southern, Jr., et al.

J. Wallace, Jr., of the Univers i ty of Texas Health Center at Tyler, and 3) Dr. Richard Sweet, Texas Depar tment of Health, Bureau of Laboratories, Austin. The collect ion of strains inc luded 54 isolates: Nocardia asteroides (40), Nocardia brasiliensis (9), Nocardia caviae (3), and an unident i f ied Nocardia sp. (2).

Organisms were tested in a manner s imilar to that reported by Wallace et al. (1983). Briefly, isolates were incubated for 3 days in brain heart infusion (BHI) broth at 35°C in ambien t air. Once dai ly the tubes were mixed thoroughly with a Vortex mixer for approx ima te ly 30 sec, wi th sterile, 0.5-mm glass beads in the tubes to reduce the t endency for c lumping of colonies. The inoculum for suscept ibi l i ty testing was p repared by adjus t ing the dens i ty of each isolate to match that of a McFar land No. 0.5 s tandard. The inocu lum was app l ied to the surface of Mue l l e r -Hin ton agar wi th a Steers inocu lum-rep l i ca t ion device. The inoculum was quantitated, and con- ta ined app rox ima te ly i x 104 CFU per spot of repl icator prong. Plates were incubated for 48 hr at 35°C in air. M i n i m u m inhibi tory concentrat ion (MIC) was interpreted as the lowest concent ra t ion of an t imicrobia l agent that resulted in no visible growth on the plates. Qual i ty controls inc luded a plate wi thout added antibiotic, and two stan- dard strains of bacteria: Escherichia coli ATCC 25922, and Staphylococcus aureus ATCC 29213. The fol lowing concentra t ions of ant imicrobial agents were used: ami- kac in - -0 .25 , 0.5, 1.0, 2.0, 4.0, 8.0, 16.0, 32.0 Ixg/ml; ceftazidime and i m i p e n e m - - 0.25, 1.0, 4.0, 8.0, 16.0, 32.0 p,g/ml; cefamandole, cefotaxime, ceftizoxime, fusidic acid, and novob ioc in - -0 .5 , 2.0, 4.0, 8.0, 16.0, 32.0 Ixg/ml; c iprofloxacin--0.25, 1.0, 2.0, 4.0, 8.0, 16.0, 32.0 Ixg/ml. Amikac in was inc luded for comparison and as an add i t iona l in ternal control.

Breakpoints for suscept ib i l i ty (see Table 1) are those recommended by the National Commit tee for Clinical Laboratory Standards (1985). In the case of fusidic acid and novobiocin, the recommenda t ions of Acar and Goldstein (1986) were used, and for c iprof loxacin the recommenda t ions of Miles Pharmaceuticals ("Handling of Cipro- floxacin [Bay 0 9867] in The Microbiological Laboratory," Revised July 1984) and those of Barry et al., (1984) were followed. In calculating the percent of strains suscept ib le in vitro, the categories of suscept ible plus moderately susceptible were used to denote suscept ibi l i ty .

The E. coli control strain was regularly inhibi ted by all agents except fusidic acid and novobiocin. The S. aureus control strain was inhibi ted by all agents; however, med ian MICs for cefotaxime, ceftazidime, and ceftizoxime were 8.0 jxg/ml. Suscep- t ib i l i ty of the Nocardia isolates to test agents is shown in Table 1. With the exception

TABLE 1. In Vitro Suscept ib i l i ty of Nocardia Species to Ant imicrobial Agents

MIC (ixg/ml) ~ Percent

Agent Range 25% 50% 75% 90% Susceptible b

Amikacin 40.25-32.0 0.5 1.0 2.0 4.0 95 Cefamando|e 40.5->32.0 4.0 8.0 32.0 >32.0 56 Cefotaxime ~<0.5->32.0 2.0 2.0 32.0 >32,0 71 Ceftazidime ~<0,25->32.0 4.0 >32.0 >32.0 >32,0 29 Ceftizoxime ~<0.5->32.0 4.0 8.0 32.0 >32,0 63 Ciprofloxacin ~<0,25-32.0 <0.25 ~<0.25 2.0 8.0 89 Fusidic acid ~<0.5->32.0 2.0 8.0 16.0 32.0 85 Imipenem ~<0.25->32.0 1.0 1.0 8.0 8.0 86 Novobiocin <0.5-~32.0 8.0 32.0 >32.0 >32.0 35

°Minimum inhibitory concentration; inhibition of 25, 50, 75 and 90% of strains. bBreakpoints for susceptibility (~.g/ml): ciprofloxacin 42.0; imipenem, fusidic acid, novobiocin 48.0;

amikacin, cefamandole, cefotaxime, ceftazidime, ceftizoxime 416.0.

Notes 121

of cefamandole , when results were segregated into species, values were not signifi- cant ly different from those of the entire group of Nocardia strains. Consequently, the data are shown only for the entire group. Most N. asteroides were suscept ible to cefamandole , whi le most other Nocardia species were resistant to that agent.

As noted above, other investigators have evaluated some of the same ant imicrobial agents as used in the present s tudy in vitro versus Nocardia sp. Acar et al. (1983) and Kitzis et al. (1985) found results s imilar to ours with cefamandole, cefotaxime, cef tazidime, and i m i p e n e m (12 strains). There was no enhancement of activity when c lavulanic acid, sulbactam, or BLP2013 (Bristol) were added. The latter agent is a penic i l lan ic acid sulfone derivative. Cynamon and Palmer (1981) evaluated nine strains of N. asteroides in vitro and found that all were inhibi ted by imipenem at

6.25 ~g/ml (eight were inhibi ted by ~< 1.56 ~,g/ml). Cefamandole inhibi ted five strains at ~ 12.5 ~.g/ml. Dewsnup and Wright (1984) evaluated 52 isolates of N. asteroides against 24 ant imicrobia l agents in vitro. They found that cefamandole, cefotaxime, ceft izoxime, ceftazidime, and imipenem had MICs0 values of 4, 4, 64, 256, and 0.5 p.g/ml, respect ively. These results are probably not significantly different from our s, wi th the possible except ion of ceftizoxime, whose MICs0 in our s tudy was 8.0 g.g/ml. The overal l range of MIC for that agent was probably not greatly different in our group of organisms, however. Garcia-Rodriquez et al. (1982) evaluated 10 strains of Nocardia sp. in vitro against a variety of agents, inc luding cefamandole, cefotaxime, cef tazidime, and imipenem. They found nine, eight, four, and 10 strains to be suscept ible , respect ively, according to the breakpoints used in our study. These results are not mater ia l ly different from our own (35 of 40 isolates of N. asteroides were suscept ib le to cefamandole at MIC <~ 16 ~g/ml). Gombert (1982) s tudied 26 isolates of N. asteroides in vitro against a variety of agents. Their results wi th ami- kacin, cefotaxime, and imipenem were vi r tual ly identical to ours. Gombert and Au- l icino (1983) also s tud ied imipenem and amikacin in vitro in combinat ion with cefotaxime versus N. asteroides and found either synergy or an addi t ive effect wi th i m i p e n e m - c e f o t a x i m e (13 strains), and with 14 of 15 strains with amikac in-cefo- taxime. Amikac in plus imipenem gave addi t ive results in 75% of strains. Gutmann, in coopera t ion with the Acar group (Gutmann et al. 1983), s tudied suscept ibi l i ty in vitro of 12 strains of N. asteroides to 46 antibiotics. Their results wi th amikacin, cefamandole , cefotaxime, ceftazidime, fusidic acid, and imipenem were similar to ours. It wou ld thus appear that our data are essential ly similar to those of other groups s tudying Nocardia spp. in vitro. Our s tudy supports the others, it contained more isolates than any previous study, and it evaluated three oral agents (ciproflox- acin, fusidic acid, and novobiocin) not previous ly well documented in this area.

In summary , we found that 89%, 86%, and 85% of Nocardia species tested were suscept ib le to c l in ica l ly re levant serum concentrat ions of ciprofloxacin, imipenem, and fusidic acid, respect ively. For reference, 95% of isolates were susceptible to amikacin. Seventy-one percent of isolates were suscept ible to cefotaxime and 63% to ceft izoxime. Relat ively few were suscept ible to ceftazidime and novobiocin. Be- cause of in vitro suscept ib i l i ty we suggest that ciprofloxacin, imipenem, and fusidic acid, and, to a lesser extent, cefotaxime and possibly ceftizoxime, may be suitable al ternat ive or adjunct ive ant imicrobia l agents in treating infections due to Nocardia species, par t icu la r ly in persons who cannot tolerate sulfonamides or aminoglyco- sides. Ciprofloxacin and fusidic acid have the added advantage of being efficacious after oral adminis t ra t ion .

The authors thank Drs. Richard Sweet and Richard J. Wallace, Jr., for supplying some of the Nocardia strains studied, and Pattie Pipes for expert secretarial assistance.

122 P . M . Southern , Jr., et al.

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