425

T E T R A C Y C L I N E : L A B O R A T O R Y A N D C L I N I C A L S T U D I E S . ~

By BaTAN McKINNEY.

T E T R A C Y C L I N E (Achromycin) is the most recent of the tetra- cycline group of antibiotics, the other members being chlortetra- cycline ('Aureomycin) and oxytetracycline (Terramycin). All

three are reported to have much the same range of activity against micro- organisms (English2), but it is said that tetracycline has a lower incidence of side-effects and fur thermore that a higher concentration can be obtained in the serum using a similar dosage. (Finland et al. 3)

There are many reports of its clinical use in non-pregnant patients (Milberg, Kamhi and Banowitch ~, McCorry and WeaverT), but few, so far as the author is aware, in puerperal infections.

The present paper is concerned with laboratory studies to determine (a) the serum levels of tetracycline obtainable with rout ine dosage and (b) the sensitivity to tetracycline of cez~ain common pathogens met with in this hospital. Clinical studies were also carried out on the response to tetracycline of fl-haemolytic streptococcal puerperal infection; the results obtained are compared with the response of a similar group of patients treated with penicillin or with penicillin and streptomycin.

Methods: (a) The assay of tetracycline in the serum was carried out by a simple agar diffusion technique with punch cups and the s tandard Oxford staphylococcus as the test organism. This method ~ves only approximate values for levels below 7 t~gs/ml. (b) Estimations of the minimal inhibi tory concentrations of antibiotics for different organisms were done by the same method as that used by McCorry and Weaver 7, but no s tandard organisms were used. (c) The method of Hilson and Elek 5 was used to determine whether tetracycline is bacteriostatic or bacteriocidal in action.

Results: Laboratory Studies. Serum levels ~n normal adults: Estimations of serum levels of tetra-

cycline were carried out in three normal adults. The dosage used was

TABLE 1.

Serum Levels of Tetracycline .follawing oral administration of five 0.50 gram doses a t six hourly intervals.

Pa t i en t 1 Hour

3 pgs./ml.

2/zgs./ml.

2 pgs./ml.

2 Hours

5 pgs./ml.

4 pgs./ml.

2 pgs./ml.

3 Hours

3 pgs./ml.

4 pgs./ml.

4 Hours 5 Hours

2 pgs./ml. 2 pgs./ml.

2 pgs./ml. 2 pgs./ml.

2 /zgs./ml. 5 pgs./ml.

6 Hours

2 ~gs./ml.

2 pgs./ml.

500 mgs. orally, at 6-hourly intervals, for 24 hours to give a total dosage of 2"5 grams. After tile fifth dose (i.e., at 24 hours) 6 specimens of

* From the Depar tment of Pathology, Rotunda Hospital, Dublin.

426 IRISH J O U R N A L OF MEDICAL SCIENCE

blood were taken at hourly intervals for assay. In all cases the values obtained were below 7 /~ gs /ml , and were of the order of 2 to 5 ~ gs/ml. (Table 1.) Serum levels of this order are in agreement with the figures published by Milberg, Kamhi and Banowitch ~ who, using a similar dos- age, found serum levels at 24 hours ranging from 1"25 / ,gs /ml . to 5 /xgs/ml, in 17 cases, However, in three other cases levels as high as 10 t~gs/ml, were found. Tunevall and Frisk TM using a dosage of 500 rags. every 8 hours obtained serum levels of 3 to 4 t~gs/ml, after 24 hours. H. owever, Gibson ~, using the same dosage scale and a serial dilution technique, could only estimate serum levels of any kind in 5 out of 9 patients to whom the drug was given orally. In these the serum levels ranged from 1"4 ~ gs. per ml. to 3 txgs/ml.

These levels are much the same as are obtained with the other tetra- cyclines; for both oxytetracycline and chlortetracycline levels of 3 to 4 /x gs. per ml. were obtained at 20 hours fol lowing four 500 mgs. oral doses at 6-hourly intervals. (WelcheS.) In this laboratory estimations of the serum levels of these antibiotics, using the same dosage and time schedule, gave values varying between 2 and 5 tLgs. per ml. which are in agreement with the above figures.

Sensitivity of Common Pathogens to Tetracyclines, Penicillin and Streptomycin. The minimal concentrations of tetracycline required to

TABLE 2.

Miminal Concentrations of Achromycin, Penicillin and Streptot~jcin required to inhibit in ~tro growth of 27 c~mmon p~thogens.

Organism Antibiotic

Staphy- Penicillin lococcus Streptomycin Aureus Achromycin

B a c t e r i u m Coli

fl-haemolytic Strepto- coccus

Streptomycin Achromycin

Penicillin Achromycin

. - q

2

6

Minimal Inhibitory Concentration (~g. or units per ml.)

m

_ I

1 1 5 6

1 1

Numerals indicate number of strains inhibited by that concentration and not by any lower concentration.

inhibit the in vitro growth of strains of staphylococcus aureus (eoagu- lase positive and mannitol fermenting), B. eoli and fl-haemolytic strep- loeoeei isolated in this hospital are shown in Table 2 ; the values obtained are somewhat higher than those found by McCorry and Weaver 7. None the less, all the strains of staphylococci and f~-haemolytie streptococci

T E T R A C Y C L I N E : LABORATORY AND CLINICAL S T U D I E S 427

examined were inhibited by a concentration of tetracycline (1"5 ~gs per ml. or less) which can be easily obtained in the serum by giving 500 rags. of tetracycline every 6. hours by mouth. On the other hand, only three of the six strains of Bact. coli are inhibited by 2"5 g gs per ml. or less.

I t should be noted that half of the staphyloccocal strains are sensitive to 1"5 units per ml. or less of penicillin, and can therefore be inhibited by the levels of penicillin obtained in the serum with normal dosage (crystalline penicillin 1,000,000 units initially, followed by 500,000 units 12-hourly), while 8 of the 15 strains of staphylococci tested against s t reptomycin are sensitive to 5 t~gs or less per ml., which concentration is within normal serum level range of this antibiotic using a dosage of 1 gram initially followed by 0"5 gram 12-hourly.

Mode of Action of Tetracycline. Tetracycline was found to be bae- teriostatic in action against staphylococcus aureus, sensitive strains of B. coll. and the fl-haemolytic streptococcus, at the levels normally obtained in the blood. I t has a bacteriocidal action in very high con- centrations (100 to 1,000 /~gs/ml.) against most of these organisms. I t is therefore comparable in action with the other tetracyclines, as well as with Chloromycetin and Ery throc in ; these, to(), are bacteriostatic in action except in very high concentrations when their action is bacterio- cidal. Penicillin on the other hand is bacteriocidal against the fl-haemolytic streptococcus and sensitive strains of staphylococcus aureus at the usual serum concentrations (2 to 4 units/ml.) , and strep- tomycin is similarly found t~ be bacteriocidal in action against sensitive strains of B. coli and staphylococcus aureus at the concentrations (10 to 20 tt gs/ml.) usually obtainable in the blood.

These findings agree with those of Jawetz e,t al2 and of Hilson and Elek 5, who state that penicillin and streptomycin are bacteriocidal in normal concentration whereas chloromycetin, chlortetracycline and oxytetracycline are bacteriostatic in action at normal blood levels and only bacteriocidal at high concentrations. Neither of these groups of workers carried out any studies on tetracycline.

Cl~nica~ Studies. Tetracycline was used routinely for the t reatment of all puerperal

cases with pyrexia who were thought to require chemotherapy, that is, any patient who had a temperature of 99 G F. or above on three successive bi-daily readings or whose temperature rose to '100 ° F. on any one occasion.

All these patients were examined clinically and a catheter specimen of ur ine and a cervical swab were takerr before tetracycline was given.

A diagnosis of genital t ract infection was made in these cases if careful clinical examination failed to show any cause for the pyrexia arising out- side the genital t ract , if the catheter specimen of urine was sterile and contained no pus cells, or if there were clinical signs of infection localised in the genital tract.

F i f t y cases of puerperal pyrexia have so far been treated and of these, 33 are regarded on clinical grounds as cases of genital t ract infection; the pyrexia in the remainder was due to pyelitis (8 cases), respi ra tory diseases (4 cases), mastitis (2 cases), and in 3 cases to other extra-genital causes. The bacteriology of the genital t rac t in the 33 cases of genital

428 IRISH JOURNAL OF MEDICAL SCIENCE

tract infection is detailed in Table 3. The incidence of the various organisms agrees closely with that found by Calman and Gibson' in an analysis of 43 high vaginal swabs taken from similar pyrexia cases with genital tract infections.

TABLE 3.

Results obtained on culture of cervical swabs taken from 33 cases of genital tract infection.

Predominan t Organism

Mixed Flora . . . . . . . . Sterile . . . . Staphylococcus" Aureu's" k~-haemolytie Streptococci (ali Groups) Streptococcus Viridans . . . . Anaerobic Streptococcus . . . . Non-haemolyt ie Streptococcus . . Bac ter ium Coli (alone) . . . .

N u m b e r of Cases

II (33%) I (3%) 1 (3%)

II (33%) 2 (6%) 3 (9%) 2 (6%) 2 (6%)

The dosage of tetracycline was 500 mgs. administered orally at 6- hourly intervals for 5 days, giving a total dosage of 10 grams. In some cases it was not possible to complete the course as the pyrexia settled rapidly and the patient wished to go home.

This dosage is twice that recommended by the manufacturers, but is the same as that used by Gibson ~ and slightly higher than that used by Tunevall '°. This dosage was decided on as other workers (Milberg, Kamhi and BanowitchS), have shown that 250 mgs. given 6-hourly will not always give a therapeutic concentration in the blood.

Of the 50 patients treated with tetracycline side-effects were noted in 4 cases. These took the form of disturbances of the gastro-intestinal tract and necessitated stopping the drug on three occasions, but the patients had clinically improved so much in these cases that no further treatment was needed.

These side-effects are listed below :

Patient. Side-Effects. Diagnosis and Course.

1 Diarrhoea on third day.

Bact. coli pyelitis. Temperature fell to normal in 12 hours. Urine sterile on fifth day. Total dosage, 10 grams.

Anorexia, vomiting and diarrhoea on third day.

Bronchopneumonia. Sputum - - pre- dominant growth of pneumococci. Apyrexial in 48 hours. Tetracycline stopped after 72 hours due to side- effects. Total dosage, 6 grams.

Anorexia, vomiting and diarrhoea on fourth day.

Abscess in thigh. Temperature did not resolve until after surgery. Tetra- cycline stopped on 7th day. Total dosage, 13"5 grams.

~TETRACYCLINE : L A B O R A T O R Y AND C L I N I C A L S T U D I E S 429

4 Anorexia, nausea and Puerpera l infection due to anaerobic diarrhoea on four th streptococcus. Apyrexia l in 36 hours. day. Tetracycline stopped on fifth day.

Total dosage, 8"5 grams.

In these four pat ients the gastro-intestinal disturbances settled down within 24 hours of cessation of the rapy and there was no recurrence.

The incidence of side reactions in the whole series is S per cent. McCorry and Weaver 7 found a similar incidence for diarrhoea, but also noted 3 per cent. of pat ients suffering f rom nausea alone. No cases of nausea alone were noted in this series. Milberg, Kamhi and Banowitch ~ had an incidence of 14-5 per cent. of gastro-intestinal side reactions in a series of 77 out-patients receiving 2 grams of tetracycline daily, but Palazzolo et al2 noted only 4"4 per cent. of ga~tro-intestinal disturbances in t rea t ing pat ients with pneumococcal pneumonia with a similar dosage. These workers account for their low incidence of side-reactions by the fact tha t most of their patients come f rom the lower socio-economic groups. No such relationship was seen in this series.

Tetracycline in fl-haemolytic streptococcal infections. The eleven cases in this group responded well to tetracycline in that the t empera ture fell to normal within 48 hours and no other antibiotic had to be iised to control the infection. However, in one case the tempera ture rose again to 100"2 ° F. on the four th day of therapy, although it fell to normal aga in on the following day. In three other cases the t empera ture continued to rise, or rose the following day to a higher t empera tu re than was noted when t rea tment first started. Six patients became apyrexial in 12 hours, one in 24 hours, one in 36 hours, two in 48 hours, while the remaining pat ient became apyrexia in: 108 hours. The average t ime for the pyrexia to settle was 30 hours.

These finding are compared with those of nine cases of fl-haemolytic streptococcal infection t rea ted in the last year in this hospital by other antibiotics. These cases were t reated with penicillin alone if only ~-haemolytic streptococci were found (7 cases), or penicillin and strep- tomycin if B. coli was also present in the cervical smear (2 cases). The dosage of penicillin used was 1,000,000 uni ts init ially followed by 500,000 units 12-hourly. St reptomycin was given as one gram initially and then 0"5 gram ]2-hourly. All these cases responded well to therapy, the pat ients becoming apyrexial , and no other antibiotic had to be used to control the infection. In only one case did the t empera ture continue to rise af ter therapy had been started. Three pat ients were apyrexial in 12 hours, two in 36 hours, three in 24 hours and one in 48 hours. The average time for the pyrexia to settle was 26 hours in this group. No side-effects were noted.

I t is quite clear tha t the clinical response in the group of tetracycline t reated cases is very little different f rom tha t of the penicillin t rea ted group.

Conclusions. The serum levels of tetracyclin~ obtainable a f te r 24 hours on a dose of

500 mgm. 6-hourly are of the order of 2 to 5 ~ g / m l . This level is above tha t required to inhibit the in vitro growth of all s trains of ~-haemolyt ie streptococcus and staphylococcus aureus, but only half of the s trains of

430 IRISH JOURNAL OF MEDICAL SCIENCE

B. colt. The actual serum values are in agreement with those found by other workers.

On clinical grounds tetracycline is as effective as, but not superior to~ penicill in in the treatment of fl-haemolytic streptococcal puerperal infection. Because of the bacteriocidal action of penicillin at normal serum levels it would seem to be the drug of choice in the seriously ill patient with this infection.

From in vitro data it is likely that tetracycline will be effective in the treatment of staphylococcal and some coliform infections. It i~ emphasised, however, that half the strains of staphylococci isolated in this hospital are sensitive to penicillin,

Acknowledgments . I am grateful to Dr. I-[. C. Moore, Director of Pathology, for his advice and criticism,

to the Mas~er, Dr. E. W. L. Thompson, for providing access to the post-natal wards: and giving me the fullest co-operation, and lastly to Lederle Laboratories for providing the " Achromycin " used in this investigation and contributing a grant towarsd expenses.

References. 1. Cahnan, l~., and Gibson, J. (1954). J . Obst. Gyn. Brit . E m p . , 61, 623. 2. English, A., P 'An et al. (1954). Antibiot . & Chemother., 4, 411. 3. Finland, M., Purcell, E., et al. (1954). J . A m . Med . Assn . , 154, 561. 4. Gibson, R. M. (1954). Personal communication. 5. Hilson, G., and Elek, S. (1954). J . Lab. Clin. Med. , 44, 589. 6. Jawetz, E., Gannison, J . , et al. (1952). J . Bact. , i, 29. 7. McCorry, R., and Weaver, J. (1955). Lancet, i, ]102. 8. Milberg, Kamhi and Banoviteh (1954). Antibiot . & Chemother., 4, 10, 1086. 9. Pallazzolo, Eisenberg, et al. (1954). Ibid. , 4, 10, 1075.

10. Tuneva]l, G., and Frisk, A. (1955). t ty f fc ia (Sweden). 11. Welch, I t . (1950). A n n . N . Y . Acad. Sci . , 53, 253.