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Pseudomonas aeruginosa S-827 Fig. 1. Method of biofilm formation. Table 1. Susceptibilities of antimicrobial agents on agar plate and in broth against Pseudomonas aeruginosa S-827

JULY CHEMOTHERAPY 888 Fig.2. Scanning electron microscopy 1992 of Pseudomonas aerugi- nosa S 827. (A) Fig. 4. Floating Bactericidal activity ceftazidime Fig. 3. Outer membrane profile of sessile and floating cells of Pseudornonas aeruginosa S827. Lanes: A, S-827; B, S-827 sessile cell (8 days); C, S-827 floating cell (8 days); D, molecular size standards were Cytochrome c hexamer (74.4 KDa), Cytochrome c tetramer (49.6 KDa), Cytochrome c trimer (37.2 KDa), and Cytochrome c dimer (24.4 KDa); E, S-827 sessile cell (3 days); F, S-827 floating cell (3 days). (A) in (-), cellお よ びsessile cellに 対 す る薬 剤 の殺 菌作 用 Floating TFLX,CPFXお ( ), and glucose. tosufloxacin 0.2ƒÊg/ml; 度 と し た Floating and (B) sessile Symbols: 0.2ƒÊg/ml; (ƒ ), ceft- cellに 対 し て キ ノロ ン は共 に 優 れ た殺 菌 力 を 示 し,薬 剤 接 触 後3時 で,両 薬 剤 共 生 菌 数 を1,000分 し め た そ れ に対 しCAZは せ え な か っ た ま た,sessile 剤2剤 quinolones aeruginosa cells 0.2% cells 6.25ƒÊg/ml. の 殺 菌 効 果 は,floating も の の,90%以 4.Floating and ciprofloxacin azidime 剤2剤 floating saline control: ( œ), 1MIC濃 of against Pseudomonas S-827 cells (B)Sessllc cells の1以 間 下 に まで減少 せ ま っ た く生 菌 数 を減 少 さ cellに 対 し て キ ノ ロ ン cellに 対 し て よ り劣 る 上 の 生 菌 数 の 減 少 が 認 め られ た しか し,CAZはfloating cellに 対 し て と同 様 ま っ た く効 果 を示 さ な か っ た cellあ る い はsessile よ びCAZの 示 し た 殺 菌 作 用 濃 度 は,3薬 cellに 対 す る 殺 菌 作 用 を,Fig.4に 剤 と も液 体 培 地 中 で の 5.TFLXと 多 糖 類 あ る い は タ ン パ ク分 解 酵 素 と の併 用効 果 0.2μg/mlのTFLXと,0.125mg/mlのlysozy-

Table 2. Activity of tosufloxacin combined with mucolytic enzymes against Pseudomonas aeruginosa S-827 sessile cells (A) Tosufloxacin (B) Ciprofloxacin (C) Ceftazidime Fig. 5. Bactericidal activity of (A) tosufloxacin, (B) ciprofloxacin, and (C) ceftazidime combined with erythromycin against Pseudomonas aeruginosa S-827 sessile cells in saline and 0.2% glucose. Symbols: (A): (-), control; ( ), tosufloxacin 0.2ƒÊg/ml;(ƒ ), erythromycin 200 ( œ), tosufloxacin 0.2 ƒêg/ml and erythromycin 200 (B): (-), control; ( ), ciprofloxacin 0.2ƒÊg/ml;

CHEMOTHERAPY JULY 1992 (A) Erythromycin (B) Clarithromycin Fig. 6. Bactericidal activity of tosufloxacin and (A) erythromycin or (B) clarithromycin against Pscudomonas acru,ginosa S 827 sessile cells in saline and 0.2% glucose. Symbols: (A): (-), control; ( œ), tosufloxacin 0.2ƒÊg/ ml, ( ), erythromycin 1ƒÊg/ml; ( œ), tosufloxacin 0.2 pg/m1 and erythromycin 1 g/ml. (B): (-), control; ( Z), tosufloxacin 0.2 pg/ml; (A), clarithromycin 1ƒÊg/ml; ( œ), tosufloxacin 0.2 pg/ml and clarithromycin 1 pg/nil. Fig. 7. Combined effect of tosufloxacin an erythromycin against Pseudomonas aeruginosa S-827 in brain heart infusion broth. Symbols: ( ), control; ( ), tosufloxacin 0.2 g/ml; (A), erythromycin 200 pg/ml; ( œ), ƒê tosufloxacin 0.2 pg/ml and erythromycin 200 g/ml. ƒê ƒê

VOL.40 NO.7

CHEMOTHERAPY JULY 1992 5) Buxton T B, Horner J, IIinton A, Rissing J P: In vivo glycocalyx expression by Staphylococcus aurcus phage type 52/52 A/80 in S. aurcus osteomyelitis. J Infect Dis 156: 942 `946, 1987 7) Da11 L, Barnes W G, Lane J W, Mills J: Enzymatic modification of glycocalyx in the treatment of experimental endocarditis due to viridans streptococci. J Infect Dis 156: 736 `740, 1987 8) Dix B A, Cohen P S, Laux D C, Cleeland R: Radiochemical method for evaluating the effect of antibiotics on Escherichia coli biofilms. Antimicrob Agents Chemother 32: 770 `772, 1988 activities of T 3262, a new fluoroquinolone. Antimicrob Agents Chemother 32: 827 `833, 1988 20) Goswami S K, Kivity S. Marom Z: Erythromycin inhibits respiratory glycoconjugate secretion 9) Anwar H, Costerton W: Enhanced activity of combination of tobramycin and piperacillin for eradication of sessile biofilm cells of Pseudonionas aeruginosa. Antimicrob Agents Chemother 34: 1666 `1671, 1990 10) Widmer A F, Wiestner A, Frei R, Zimmerli W: Killing of non-growing and adherent Escherichia coli determines drug efficacy in device-related infections. Antimicrob Agents Chemother 35: 741 746, 1991 ` 11) Fujimaki K, Noumi T, Saikawa I, Inoue M, Mitsuhashi S: In vitro and in vivo antibacterial from human airways in vitro, Am Rev Respir Dis 141: 72 `78 1990

VOL.40 NO.7 Characteristics of biofilm formed by Pseudornonas aeruginosa in vilro Kazuo Fujimaki, Yasushi Ikeda, Masahiro Takahata and Takashi Yasuda Research Laboratories, Toyama Chemical Co., Ltd., 2 4-1 Shimookui, Toyama, Japan Biofilm formation by Pseudornonas aeruginosa was investigated by using medical teflon sheet as an artificial foreign device in vitro. Biofilm formed by the glycocalyx and cells was observed by scanning electron microscopy. Ceftazidime (CAZ) at 1 MIC did not show any bactericidal activity against sessile or floating cells. Tosufloxacin (TFLX) and ciprofloxacin (CPFX) had excellent bactericidal effects at 1 MIC against floating cells and a >90% decrease was observed in the number of viable sessile cells. Enhanced activities were observed against sessile cells not only with a combination of TFLX and CPFX at 1 MIC and erythromycin (EM) at 1/4 MIC to when the hourly changes of viable cells were observed but also for TFLX at 1 MIC and EM or clarithromycin at 1ƒÊg/ ml when the changes were observed over days. However, synergism was not observed when CAZ and EM were combined. The activity of TFLX was not enhanced when it was combined with lysozyme, hyaluronidase, protease or streptokinase.

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