ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, Aug. 1980, p. 338-345 Vol. 18, No. 2 0066-4804/80/08-0338/08$02.00/0 Comparison of In Vitro Activity of Sch 21420, a Gentamicin B Derivative, with Those of Amikacin, Gentamicin, Netilmicin, Sisomicin, and Tobramycin C. THORNSBERRY,l* A. L. BARRY,2 R. N. JONES,3 C. N. BAKER,' R. E. BADAL,2 AND R. R. PACKER3 Antimicrobics Investigations Section, Center for Disease Control, Atlanta, Georgia 303331; Clinical Microbiology Laboratories, University of California (Davis), Sacramento Medical Center, Sacramento, California 95817 2; Clinical Microbiology Division, Kaiser Foundation Hospital Laboratories, Portland, Oregon 97217 3 Sch 21420 is a new aminoglycoside synthesized from gentamicin B. Suscepti- bility tests with Sch 21420, amikacin, gentamicin, netilmicin, sisomicin, and tobramycin were performed on a variety of bacterial species including 44 with known mechanisms of resistance to aminoglycosides. Sch 21420 and amikacin had similar effects on all except Haemophilus influenzae and Neisseria species, which were more susceptible to amikacin. Except with some strains of Serratia marces- cens, the drugs used were bactericidal. Sch 21420 and amikacin were more stable than the other four aminoglycosides in the presence of the inactivating enzymes produced by some strains. Strains which were very resistant to Sch 21420 and amikacin either were permeability mutants or produced AAC (6')-I inactivating enzyme. The effect of cations on the susceptibilities of these strains to Sch 21420 and amikacin was seen mostly with Pseudomonas aeruginosa and to Sch 21420 with Acinetobacter. Cations did not affect the susceptibilities of other Pseudo- monas species, Enterobacteriaceae, Staphylococcus aureus, or Streptococcus faecalis to Sch 21420 or amikacin. Sch 21420 is an aminoglycoside which was laboratories participating in this study: 182 strains of synthesized by a method similar to that used in Enterobacteriaceae, 168 strains of gram-negative ba- the synthesis of amikacin, except that gentami- cilli other than Enterobacteriaceae (including 100 cin B was used in of The strains of P. aeruginosa), 108 strains of staphylococci place kanamycin (12). and streptococci, and 49 strains of Neisseria species. activity ofthis compound was found to be similar The various genera and species represented are shown to that of amikacin against the Enterobacteria- in Tables 1 and 2. In addition, 44 strains with known ceae and Pseudomonas aeruginosa (7, 9, 11, 13, aminoglycoside resistance mechanisms were also 15, 17). tested. In this study, we compared the inhibitory and Most of the isolates were tested in duplicate by two bactericidal activities of Sch 21420 to those of of the collaborating laboratories (Center for Disease amikacin, gentamicin, netilmicin, sisomicin, and Control and the Sacramento Medical Center) in a tobramycin against a wide variety of gram-pos- manner previously reported (1, 8). A third laboratory, itive and gram-negative organisms. The effect of the Kaiser Foundation, also tested a more limited in inoculum size and cation concen- number to study the effect of inoculum size on mini- differences mum inhibitory concentrations (MICs). Very similar tration on the activity of these six aminoglyco- MICs were obtained at the participating institutions. sides was assessed. The activities of these drugs Antibiotic susceptibility tests. MICs were deter- on organisms with various mechanisms ofresist- mined by the broth microdilution method with test ance (inactivating enzymes and permeability trays prepared commercially (Micro Media Systems, mutants) were also compared. San Jose, Calif.). Mueller-Hinton broth was dispensed into a single lot of trays and distributed to the partic- ipating laboratories. These trays were stored at -600C MATERIALS AND METHODS until inoculated. The trays were thawed at room tem- Antibiotics. Antibiotic powders suitable for sus- perature (approximately 20 to 30 min) and then were ceptibility tests were supplied as follows: Sch 21420, inoculated with disposable inoculators delivering 5 pd gentamicin, netilmicin, and sisomicin were from Scher- to each well. ing Corp., Bloomfield, N.J.; aninkacin was from Bristol At all three laboratories, an actively growing broth Laboratories, Syracuse, N.Y.; and tobramycin was culture was diluted to match the turbidity of a 0.5 from Eli Lilly & Co., Indianapolis, Ind. McFarland standard. The suspension was then diluted Organisms. A total of 507 bacterial isolates were 1:50 in sterile water containing 0.02% Tween 80 and collected in the following amounts from the various dispensed as described previously. The final inoculum 338 VOL. 18, 1930 Sch 21420 IN VITRO COMPARISON 339 U- o r-4 o- '6 L1- L6e1- c .4 eq ev ~~ .4 ;SA eq P-4 Aq-4 to I A o1 I oNu A o' ool o 0 equ .14 I--:, _i _ .-, I-- I-- U . L Lo~ C 5 - cc Z Ci eq -I- iA A C" " -1 -- .9 A Lo co C'l eq cl I 6 6Ci c6 0o 0 C6 _. A A I.-, .- 4.R o a - -- - eq~~C eq- S I Q o, 1- 9 Z A A A 0 0 0 0 A > .o u oq oq M 2o- 66C ci 6 C; 4. aCD 0 .U -- - Co co * I-, I-,I- I- I., - o0 C0 6 __ r eDq ~o eq eq,V- A Lo 6 A o o 6 cc 0A 6oi05 *o0 4 _ S. _ Q o _ _ eq I" l " . I. I.- I-- e9I-- I--Cq _I_-l.e A cc .Q5 U - Do co Z5 acc cc 0 c6 0 A _-P- A . l~e u I LO cc Lo 0 o Ci o- o o C5 r0U2: o 0 CD eqN -4 Cq r- _-/ cq I" 0 V-4 IS _ _ _ A '- _ 0 sr eq _0 I A§N X rn A A3 r 4 A o o eq o LO P-4 "- lO L 6 6 6 6 6 65 6 6 a eqi.I_UM 340 THORNSBERRY ET AL. ANTIMICROB. AGENTS CREMOTHER. was approximately 105 colony-forming units (CFU) 101 4' 04> CO_ per ml. For testing the fastidious streptococci, includ- ing Streptococcus pyogenes and Streptococcuw pneu- .9cc *E 04 CO a0440N moniae, the inoculum was standardized in Mueller- 0, Hinton broth containing 5% lysed rabbit blood, and 0.1 ml of this adjusted cell suspension was added to each microdilution well, giving a final concentration of o es1 es e ut CFU/ml. In tests with Haemophilus influenzae, cc .2 II enough Filde reagent was added to the inoculum to VI yield a final concentration of 1%. 1. The MIC was recorded as the lowest concentration O19C?o C o?' totally inhibiting bacterial growth (clear well) after 101 CO' ' COaQ0 > approximately 18 h of incubation at 35°C in a forced- A air incubator. co .0 N e- c:- q _ Minimum lethal concentrations (MLCs) were de- termined for 56 strains from nine genera by subcultur- ng 5-pd portions from each microdilution well onTryp- _ _ ticase (BBL Microbiology Systems, Cockeysville, Md.) .0 Vi VI soy agar with 5% sheep blood. The subculture was *tt 1 made with a multiple inoculum replicator onto a plate Ul LO W." IV 0 _O0 (150 by 100 mm). After 48 h of incubation, the end- a .2 points were read as the lowest concentration yielding .2 no more than 0.1% survivors (99.9% killed). I U N1 Nq C#3 Cf 7 p p The effect of varying the inoculum concentrations A ObC4NI" ^ on MIC and MLC endpoints was studied with 56 rapidly growing faculative anaerobes. Trays were in- I48 VI VI oculated to achieve final concentrations of 103, 105, 16. 4) and 107 CFU/ml. MICs and MLCs were interpreted as %9z. described above. Comparisons of MICs and MLCs were made with results obtained with an inoculum density of 1lO CFU/ml. t CIO 66 C0C0a- Neisseria gonorrhoeae and Neisseria meninWitidis _1 were tested by the agar dilution method. Proteose - - peptone agar with 1% hemoglobin and 1% Kellogg 5.. VI VI I-, _l, supplement was prepared by incorporating appropri- ate antibiotic concentrations. The inoculum was made by suspending colonies in Mueller-Hinton broth di- -^ 0- luted to a concentration of 1(9 CFU/mL Plates were 90 .2 then inoculated by a Steers replicator (16). The MICs (NO U were determined after 24 h of incubation in 5% CO2 at es 35°C. 04. 6 'C RESULTS 0 S N The ranges and modes of the MICs of Sch iSZs C 0 21420 and five other aminoglycosides for 325 enteric and nonfermentative .1.Go gram-negative ba- eq *# cilli are shown in Table 1. Based on MICs, the I to most active aminoglycoside was sisomicin, fol- O lowed in decreasing order by tobramycin, gen- .E tamicin, netilmicin, Sch 21420, and amikacin. 'VI 4 44 |' However, most of these organisms were suscep- O4 -to tible to all the drugs. Some strains ofthe Pseudomonas and d1 0.2 t species etlo nearly all strains of Providencia stuartii were N- 00. very resistant to the six drugs as indicated by a MICs of >64 ug/ml. Some strains of Serratia COh marcescens were very resistant to Sch 21420, amikacin, and tobramycin at the >64-pg/ml 'C level, as were some strains of Acinetobacter 1004j f ju calcoaceticus subsp. anitratus to gentamicin, I netilmicin, sisomicin, and tobramycin. Most of B . these species, however, had a modal MIC within a susceptible or moderately susceptible range or VOL. 18, 1980 Sch 21420 IN VITRO COMPARISON 341 bimodal MICs with susceptible and very resist- ant groups, as seen with most of the results with t. qa 'vc eqeq cq eqc Pseudomonas species other than P. aeruginosa. .5 Similar results for H. influenzae, Neisseria LO eq eq _-4 - - 00X - q species, staphylococci, and streptococci are 0i shown in Table 2.