Microbiol. Immunol. Vol. 22 (9), 565-568, 1978
Rifampicin Tuberculosis
Tokumitsu
Resistant Patients
TANAKA, Matsuhisa
Strains Treated
INouE,
of Bacteria with
the
and Susumu
from Drug
MITSUHASHI
Department of Microbiology,Schoolof Medicine, Gunma University,Maebashi (Received for publication, December 5, 1977)
According to the wide use of chemotherapeutic agents, drug resistant strains of bacteria appeared in many countries and have become one of the most important problems in practical medicine (6) . Chemotherapeutic agents that are used for the treatment of patients, cause appearance of resistant strains and select the resistant strains in parallel with the use of drug. Then, the practical use of chemotherapeutic agents and control of their use have become one of the urgent problems in practical medicine. Rifampicin has a wide antibacterial spectrum and is characteristic as antituberculosis drug. According to the fact that rifampicin (RFP) is used only for the treatment of tuberculosis patients in sanatoria for a long term, we investigated appearance of RFP-resistant strains of bacteria and their selection in RFP-treated patients. Tuberculosis patients at 5 sanatoria were randomly selected and they were all daily treated with oral administration of 400 mg of rifampicin (RFP) . Heart infusion agar (HIA, Eiken Chemical Co., Tokyo) was used for the isolation of Escherichiacoli strains from the faeces. HIA-Lac agar consisted of 1,000 ml of HIA, 15 g of lactose and 40 ml of 0.2% bromthymol blue. EMB-Lac agar consisted of 1,000 ml of EMB agar (Difco) and 15 g of lactose. HIA-salt medium was used for isolation of Staphylococcusaureus from sputa and consisted of 1,000 ml of HIA and 75 g of sodium chloride. Peptone water was used for liquid culture and consisted of 1,000 ml of distilled water, 5 g of NaCl and 10 g of peptone. E. coli strains were isolated from the faeces. A small volume of saline solution was added to faeces. The mixed material was streaked on HIA-Lac plates and incubated at 37 C. After 20 hr of incubation, 100-150 lactose-fermenting colonies were randomly picked and streaked on EMB-Lac agar plates. The biological and biochemical properties of these colonies were examined and E. coli strains were used for further studies. S. aureus strains were isolated from the sputa. Sputum was suspended in a small volume of saline solution and the mixed material was streaked on HIA-salt plates. The biological and biochemical properties of these colonies were examined and S. aureusstrains were used for further studies. The MIC (minimum inhibitory concentration) of RFP was examined using 100 E. coli and 100 S. aureus strains. They were all of clinical origin and the stock 565
566
T. TANAKA
(a)
ET AL
(b)
Fig. 1. MIC distributions of RFP against clinical isolates of E. coli and of S. aureus strains. One hundred strains of E. coli and S. aureus were all isolated from inpatients at various hospitals in Japan and not from tuberculosis patients. (a) E. coli strains; (b) S. aureusstrains.
cultures of this laboratory. According to the results shown in Fig. 1, HIA containing 50 ,ug/ml of RFP was used for the selection of RFP-resistant E. coli strains. HIA containing 0.4 ,ug/ml of RFP was used for the selection of RFP-resistant S. aureus strains. MIC was determined by inoculating a suspension of each colony in peptone water (about 106 cells per ml) on a series of HIA plates containing serial two fold dilutions of RFP. We selected randomly 89 tuberculosis patients at 5 sanatoria and treated them with rifampicin. One, 2, and 6 months after administration of rifampicin (RFP), we examined appearance of RFP-resistant strains of E. coli and S. aureus from the patients. One month after the treatment, we could demonstrate RFP-resistant E. coli strains from all patients examined and almost all of E. coli strains in faeces were found to be RFP-resistant. Two and 6 months after treatment with the drug, we could isolate highly RFP-resistant E. coli strains from the all patients. It should be noted that almost all of E. coli strains in the faeces are RFP-resistant (Table 1). By contrast, we could isolate RFP-resistant S. aureus strains from only 28 percent of the patients 1 month after treatment with RFP. But the isolation frequency of RFP-resistant S. aureusstrains increased gradually by the administration period of the drug from 28 to 43 and 50 percent. In contrast to the dense population of RFPresistant E. coli strains in the faeces, the population of RFP-resistant S. aureusstrains in the sputa varied from 100 to 1 percent even after 6 months' the treatment (Table 2). We examined the development of RFP-resistant strains of E. coli NIHJ and S. aureus 209P on HI agar plates containing 5 and 10 MIC concentrations of RFP for each strain, indicating no big difference in the development rate of RFP-resistant ones between both strains. RFP is well absorbed from the gastro-intestinal tract. Peak blood level usually occur between 2 and 4 hr following oral administration and therapeutically effective concentrations of the drug are maintained for 24 hr (1). The drug concentration of
567
NOTES Table
1.
a) Number
Table
Isolation
of RFP-resistant
of RFP-resistant
2.
Isolation
E. coli strains
of RFP-resistant
a) See the footnote of tested
E. coli strains
of Table
1. No.
per
S. aureus
total
from
number
strains
of RFP-resistant
from
patients
treated
of tested
patients
S. aureus strains
with
RFP
E. coli strains.
treated
per
with
total
RFP
number
S. aureus strains.
RFP in the liver is the highest and followed by these in the lung and kidney (2, 3) . Elimination of RFP into bile is much higher than that in the urine (2, 4) and the sustained blood levels have been attributed to the existence of an entero-hepatic recirculation of RFP (1, 5) . But the concentration of RFP in sputum is rather low even when a high level of RFP is maintained in blood (3) . These data reflect the rapid appearance and a high population of RFP-resistant E. coli strains in faeces compared with those of S. aureusstrains in sputum. We are greatlyindebtedto H. Baba (NakanoChestHospital,Tokyo),H. Yoshida(OharaSanatorium,Saitama),T. Oshima(NationalSanatoriumOhinata-So,Gunma)and S. Ogiwara(Gunma PrefecturalMaebashiHospital,Gunma),for the isolationof the bacterialstrainsfrom the tuberculous patients.
568
T. TANAKA
ET AL
REFERENCES 1) 2) 3) 4) 5) 6)
Curci, G., Ninni, A., and Di Mezza, F. 1967. Indagini sperimentali sulla farmaco-cinetica della rifampicina. Arch. Tisiol. 23: 293-314. Furesz, S., Scotti, R., Pallanza, R., and Mapelli, E. 1967. Rifampicin: a new rifampicin. III. Absorption, distribution and elimination in man. Arzneimittel-Forsch. 17: 534-543. Kawamori, Y., and Nishizawa, N. 1970. Antistaphylococcal activity of rifampicin. Shinryo 23: 996-1002. Keberle, H. Meyer-Brunot, H.G., and Schmidt, K. 1966. Pharmacokinetic and metabolic studies with labeled rifamycin antibiotics. Antimicrob. Ag. Chemoth. p. 365. Mashimo, K. 1974. Pharmacokinetics of drugs, p. 78-105. In Mashimo, K., and Mitsuhashi, S. (eds), Drug resistance and chemotherapy, Igakushoin, Tokyo. Mitsuhashi, S. 1977. Epidemiology of bacterial drug resistance, p. 3-37. In Mitsuhashi , S. (ed), R factor, drug resistance plasmid, University of Tokyo Press, Tokyo.
Requests for reprints should be addressed to Dr. Susumu Mitsuhashi, Department of Microbiolgy , School of Medicine, Gunma University, Showa-machi, Maebashi, Japan.
Rifampicin Tuberculosis
Tokumitsu
Resistant Patients
TANAKA, Matsuhisa
Strains Treated
INouE,
of Bacteria with
the
and Susumu
from Drug
MITSUHASHI
Department of Microbiology,Schoolof Medicine, Gunma University,Maebashi (Received for publication, December 5, 1977)
According to the wide use of chemotherapeutic agents, drug resistant strains of bacteria appeared in many countries and have become one of the most important problems in practical medicine (6) . Chemotherapeutic agents that are used for the treatment of patients, cause appearance of resistant strains and select the resistant strains in parallel with the use of drug. Then, the practical use of chemotherapeutic agents and control of their use have become one of the urgent problems in practical medicine. Rifampicin has a wide antibacterial spectrum and is characteristic as antituberculosis drug. According to the fact that rifampicin (RFP) is used only for the treatment of tuberculosis patients in sanatoria for a long term, we investigated appearance of RFP-resistant strains of bacteria and their selection in RFP-treated patients. Tuberculosis patients at 5 sanatoria were randomly selected and they were all daily treated with oral administration of 400 mg of rifampicin (RFP) . Heart infusion agar (HIA, Eiken Chemical Co., Tokyo) was used for the isolation of Escherichiacoli strains from the faeces. HIA-Lac agar consisted of 1,000 ml of HIA, 15 g of lactose and 40 ml of 0.2% bromthymol blue. EMB-Lac agar consisted of 1,000 ml of EMB agar (Difco) and 15 g of lactose. HIA-salt medium was used for isolation of Staphylococcusaureus from sputa and consisted of 1,000 ml of HIA and 75 g of sodium chloride. Peptone water was used for liquid culture and consisted of 1,000 ml of distilled water, 5 g of NaCl and 10 g of peptone. E. coli strains were isolated from the faeces. A small volume of saline solution was added to faeces. The mixed material was streaked on HIA-Lac plates and incubated at 37 C. After 20 hr of incubation, 100-150 lactose-fermenting colonies were randomly picked and streaked on EMB-Lac agar plates. The biological and biochemical properties of these colonies were examined and E. coli strains were used for further studies. S. aureus strains were isolated from the sputa. Sputum was suspended in a small volume of saline solution and the mixed material was streaked on HIA-salt plates. The biological and biochemical properties of these colonies were examined and S. aureusstrains were used for further studies. The MIC (minimum inhibitory concentration) of RFP was examined using 100 E. coli and 100 S. aureus strains. They were all of clinical origin and the stock 565
566
T. TANAKA
(a)
ET AL
(b)
Fig. 1. MIC distributions of RFP against clinical isolates of E. coli and of S. aureus strains. One hundred strains of E. coli and S. aureus were all isolated from inpatients at various hospitals in Japan and not from tuberculosis patients. (a) E. coli strains; (b) S. aureusstrains.
cultures of this laboratory. According to the results shown in Fig. 1, HIA containing 50 ,ug/ml of RFP was used for the selection of RFP-resistant E. coli strains. HIA containing 0.4 ,ug/ml of RFP was used for the selection of RFP-resistant S. aureus strains. MIC was determined by inoculating a suspension of each colony in peptone water (about 106 cells per ml) on a series of HIA plates containing serial two fold dilutions of RFP. We selected randomly 89 tuberculosis patients at 5 sanatoria and treated them with rifampicin. One, 2, and 6 months after administration of rifampicin (RFP), we examined appearance of RFP-resistant strains of E. coli and S. aureus from the patients. One month after the treatment, we could demonstrate RFP-resistant E. coli strains from all patients examined and almost all of E. coli strains in faeces were found to be RFP-resistant. Two and 6 months after treatment with the drug, we could isolate highly RFP-resistant E. coli strains from the all patients. It should be noted that almost all of E. coli strains in the faeces are RFP-resistant (Table 1). By contrast, we could isolate RFP-resistant S. aureus strains from only 28 percent of the patients 1 month after treatment with RFP. But the isolation frequency of RFP-resistant S. aureusstrains increased gradually by the administration period of the drug from 28 to 43 and 50 percent. In contrast to the dense population of RFPresistant E. coli strains in the faeces, the population of RFP-resistant S. aureusstrains in the sputa varied from 100 to 1 percent even after 6 months' the treatment (Table 2). We examined the development of RFP-resistant strains of E. coli NIHJ and S. aureus 209P on HI agar plates containing 5 and 10 MIC concentrations of RFP for each strain, indicating no big difference in the development rate of RFP-resistant ones between both strains. RFP is well absorbed from the gastro-intestinal tract. Peak blood level usually occur between 2 and 4 hr following oral administration and therapeutically effective concentrations of the drug are maintained for 24 hr (1). The drug concentration of
567
NOTES Table
1.
a) Number
Table
Isolation
of RFP-resistant
of RFP-resistant
2.
Isolation
E. coli strains
of RFP-resistant
a) See the footnote of tested
E. coli strains
of Table
1. No.
per
S. aureus
total
from
number
strains
of RFP-resistant
from
patients
treated
of tested
patients
S. aureus strains
with
RFP
E. coli strains.
treated
per
with
total
RFP
number
S. aureus strains.
RFP in the liver is the highest and followed by these in the lung and kidney (2, 3) . Elimination of RFP into bile is much higher than that in the urine (2, 4) and the sustained blood levels have been attributed to the existence of an entero-hepatic recirculation of RFP (1, 5) . But the concentration of RFP in sputum is rather low even when a high level of RFP is maintained in blood (3) . These data reflect the rapid appearance and a high population of RFP-resistant E. coli strains in faeces compared with those of S. aureusstrains in sputum. We are greatlyindebtedto H. Baba (NakanoChestHospital,Tokyo),H. Yoshida(OharaSanatorium,Saitama),T. Oshima(NationalSanatoriumOhinata-So,Gunma)and S. Ogiwara(Gunma PrefecturalMaebashiHospital,Gunma),for the isolationof the bacterialstrainsfrom the tuberculous patients.
568
T. TANAKA
ET AL
REFERENCES 1) 2) 3) 4) 5) 6)
Curci, G., Ninni, A., and Di Mezza, F. 1967. Indagini sperimentali sulla farmaco-cinetica della rifampicina. Arch. Tisiol. 23: 293-314. Furesz, S., Scotti, R., Pallanza, R., and Mapelli, E. 1967. Rifampicin: a new rifampicin. III. Absorption, distribution and elimination in man. Arzneimittel-Forsch. 17: 534-543. Kawamori, Y., and Nishizawa, N. 1970. Antistaphylococcal activity of rifampicin. Shinryo 23: 996-1002. Keberle, H. Meyer-Brunot, H.G., and Schmidt, K. 1966. Pharmacokinetic and metabolic studies with labeled rifamycin antibiotics. Antimicrob. Ag. Chemoth. p. 365. Mashimo, K. 1974. Pharmacokinetics of drugs, p. 78-105. In Mashimo, K., and Mitsuhashi, S. (eds), Drug resistance and chemotherapy, Igakushoin, Tokyo. Mitsuhashi, S. 1977. Epidemiology of bacterial drug resistance, p. 3-37. In Mitsuhashi , S. (ed), R factor, drug resistance plasmid, University of Tokyo Press, Tokyo.
Requests for reprints should be addressed to Dr. Susumu Mitsuhashi, Department of Microbiolgy , School of Medicine, Gunma University, Showa-machi, Maebashi, Japan.
