MORPHOLOGICAL MARKER OF GENTAMICIN RESISTANCE

1295

Letters

to

MORPHOLOGICAL MARKER OF GENTAMICIN RESISTANCE

the Editor

SiR,-We believe that we have found a previously unreported morphological marker of gentamicin resistance in Staphylococcus pyogenes. ENZYME ASSAY FOR SPERM MOTILITY

StR,—There is

no quantitative method of estimating sperm of artificial insemination or male infertility. viability Sperm motility, taken to be a better measure of fertility than sperm count, is usually estimated by eye, photographic record-

in

cases

ing, or expensive Doppler techniques. By enzymological

exam-

ination of crude and partially purified human semen, we found that the enzymes fumarase and diamine oxidase may be of considerable importance in the development of a quantitative assay for sperm motility and ultimately male fertility. Measurements of fumarase activity and of diamine-oxidase activity2 in whole semen gave the results in the accompanying table on 100 semen samples. Fumarase activity showed a highly significant correlation with the qualitative determinations of percentage motility, while diamine-oxidase activity rose significantly in samples with less than 30% motility. Similar correlations were shown between fumarase and sperm count, while diamine-oxidase activity levels were raised in samples with sperm counts less than 20 million ml-1. These relationships show that the assay of these enzymes in human semen could form the basis of quantitative estimations of the viability of human semen, both in male infertility and in the analysis of semen for artificial insemination. Measurements of sperm count and motility have until now been the only means of assessing seminal-fluid viability. These crude methods might be replaced by an index which relates directly to the metabolic state of the semen. Determination of fumarase, a key enzyme in the tricarboxylic-acid cycle, and diamine oxidase, which can convert both spermidine and

putrescine to their corresponding amino-aldehydes, would provide a fast, reliable, and accurate index of the fertilising potential of semen. Samples with a fumarase activity below 0.015 A A250 min-’ ml-l and a diamine-oxidase activity above 0.015 A A250 min-’ ml-l would have a low potential fertility. Large numbers of semen samples could be analysed routinely by a technician, each spectrophotometric assay using only one substrate, with a reproducibility better than that of the qualitative estimations; indeed, errors indicated in the enzyme estimations in the table are more likely to be due to errors in the determinations of motility. Current studies on a wide spectrum of enzymes in split ejaculates and purified preparations of human semen may help to elucidate the mechanisms of action of seminal enzymes and provide further useful relationships for fertility assessment. Department of Obstetrics and Gynæcology, St Mary’s Hospital, University of Manchester, Whitworth Park, Manchester M13 0JH

1 2

M.

JAMES C. CRABBE

Racker, E. Biochim. biophys. Acta, 1950, 4, 211. Bardsley, W. G., Crabbe, M. J. C., Shindler, J. S., Ashford, J. J 1972, 127, 875.

Three gentamicin-resistant strains, isolated at St. George’s Hospital from routine specimens (two wound swabs and a urine), were compared with three gentamicin-sensitive strains-namely, one resistant to penicillin, cloxacillin,

S. Biochem.

*":$’

1--Central dense line (DL) in septal walls of gentamicinresistant strains (x200 000).

Fig.

Fig. 2-Septal

walls of

gentamicin-sensitive

strains (x200

000).

and colistin, one sensitive to all of these antibiotics, and the Oxford strain. 4 h subcultures were centrifuged at 4000 r.p.m. for 20 min and the organisms were fixed in glutaraldehyde, post-fixed in osmium, dehydrated in ethanol, and embedded in ’Epon’. Thin sections were cut, stained with uranyl-acetate/Reynolds lead-citrate, and examined in a JEM-100C electron microscope. With all three gentamicin-resistant strains, we found on each side of the dividing septum a line of electron-lucent material (fig. 1) which was absent from the three sensitive strains

erythromycin, tetracycline, co-trimoxazole,

(fig. 2). The three resistant strains were subsequently found to have the same phage-typing pattern (47/83A/85) different from the three sensitive strains (83A; 79/80/84/85/81; Oxford). The additional electron-lucent material found in the resistant organisms may, therefore, be a morphological feature of the

FUMARASE AND DIAMINE-OXIDASE ACTIVITY IN HUMAN SEMEN

1296 than a marker of gentamicin think this unlikely. Further experidetermine its significance.

particular phage type, rather resistance, although ments are

planned to

we

Department of Medical Microbiology, St. George’s Hospital Medical School, London SW1

psoriasis at present, plans for these further studies are being discussed with the manufacturers and with the Committee on Safety of Medicines. in

J. E. BEESLEY

Department of Dermatology, Guy’s Hospital,

A. T. KLOUDA

London SE1 9RT

D. J. ATHERTON R. S. WELLS

Department of Cancer Chemotherapy, Imperial Cancer Research Fund,

RAZOXANE

(ICRF

159) IN PSORIASIS

SIR,-We wish to report our experience with razoxane, (±)-1,2-bis (3,5-dioxopiperazin-1-yl) propane (ICRF 159), in the treatment of psoriasis. This drug is an exceptionally well tolerated antimitotic agent which has so far proved free of toxic effects other than those predictable from its cytostatic action. The skin lesions of psoriasis are characterised histologically by abnormal epidermal proliferation and related changes in the dermal capillaries (dilatation, tortuosity, proliferation, and "tufting"). Similar microvascular phenomena are seen in tumours. Treatment with razoxane results in "normalisation" of the vascular architecture in several experimental tumours in animals,1 an effect which has not been observed with other antimitotic drugs. Judging from the efficacy of other antimitotic drugs in psoriasis and this more specific effect of razoxane, we believed that a pilot study in psoriasis was justified. 9 patients were treated with razoxane, of whom 7 (5 male, 2 female, age range 21-80 y) had severe and intractable psoriasis, and 2 relatively mild disease which they were unable to treat topically at home. The drug was given by mouth in courses of three days, either weekly or fortnightly, in doses varying from 375 to 750 mg/d. The dosage was determined initially by consideration of the patient’s age and surface area and the severity of the disease, and subsequently by response to therapy and the leucocyte count.

All patients improved within four weeks (reduction of erythand scaling and relief of pruritus and soreness when these were prominent symptoms). In every case, further treatment is resulting in continued resolution of all lesions. At the time of writing, the first patient has completed sixteen weeks of treatment, and the last has completed five weeks. The skin of 2 patients has cleared altogether, 4 retain only fading dusky discolouration at the sites of previous lesions, and the remaining 3 patients, who have only received treatment for five weeks, are all improving progressively. The drug has been well tolerated, though 1 patient complained of intermittent transient sensations of "faintness" during her initial courses, which have since disappeared during continued treatment. Some neutropenia has been seen in all patients, though it has been generally mild, dose-related, and easily controlled by dose adjustment. 3 of the patients had previously had methotrexate, and the first 2 proved unexpectedly liable to neutropenia when they were given razoxane. In both ema

profound neutropenia developed (<1000 neutrophils/10-61), though in both cases the neutrophil count rapidly returned to normal when the drug was stopped and has since remained satisfactory during further courses of the drug in reduced dose. In the 3rd patient, it was possible to avoid this reaction by modifying the dose regimen. No other side-effects or adverse reactions have been detected. We believe there is reason to hope that razoxane may become a useful addition to present treatments for this distressing and disabling disease. We plan to follow this small un-, controlled pilot study with a controlled trial, and we are confident that hsematological reactions can be minimised by reduced dose schedules for patients previously treated with other antimitotic agents. Since razoxane is not specifically available for clinical trials 1. LeServe, A. W.,

Hellmann,

K. Br. med.

J. 1972,

i,

597.

Lincoln’s Inn Fields, London WC2A 3PX

K. HELLMANN

RESERPINE AND BREAST CANCER

SIR,-In our investigation’ of the controversial topic of rauwolfia alkaloids and breast cancer 2-9 we asked 488 women who had been admitted for breast biopsy about reserpine use. The interviews were carried out without knowledge of whether the eventual diagnosis would be breast cancer or a benign breast disorder. Most of the breast cancers were in women over the age of 50. Beyond that age, among cases and controls, the prevalence of reserpine use was more than 20%. We thus had ample data. From an age-stratified comparison of rates of reserpine use in cases and controls, we found no evidence to suggest an association with breast cancer. Moreover, it is unlikely that we would have missed a strong association by chance: the point estimate of relative risk was 0.6, and the upper 95% confidence limit was 1-1. al .7 suggested that long-term use of reserpine what would otherwise remain latent breast might cancer (promotion). We evaluated continuous use of reserpine for more than a year both when it took place in the past and when it had continued up to the time of admission. There was no evidence of a positive association.

Armstrong

et

uncover

Reserpine might cause benign breast disease as well as breast cancer, thus explaining our negative results, but we know of no evidence that reserpine has this effect. Nevertheless, as a precaution, we also compared our breast-cancer patients with a series of 101 women admitted for surgical treatment of various benign conditions. Because of small numbers, the confidence limits for this comparison were wide, but again we found no evidence of an association. In contrast to the U.S.A. and, more particularly, Britain, reserpine is used alone or in combination to treat over 80% of diagnosed hypertensive patients in Germany. If the rauwolfia

breast cancer, this would be a matter of However, whatever the reason for the disparmajor ate results in the various studies might be, we feel that our findings are reassuring; they argue rather persuasively against a causal relationship.

alkaloids do

cause

concern.

Department of Clinical Pharmacology, University of Berlin, Klinikum Steglitz, D-1000 Berlin 45, West Germany Free

H. KEWITZ P.-M. SCHRÖTER

Institute for Medical Statistics and Biomathematics,

University of Düsseldorf Central Institution for Data Free University of Berlin

H.-J. JESDINSKY Processing, E. LINDTNER

1. Kewitz, H., Jesdinsky, H.-J., Schröter, P-M., Lindtner, E. Eur. J clin Pharmac. (in the press). 2. Boston Collaborative Drug Surveillance Program. Lancet, 1974, ii, 669. 3. Armstrong, B., Stevens, N., Doll, R ibid. p. 272. 4. Hemonen, O. P., Shapiro, S., Tuominen, L., Turunen, M. I. ibid. p. 675. 5. O’Fallon, W. M., Labarthe, D., Kurland, L. T. ibid. 1975, ii, 292. 6. Laska, E. M., Siegel, S., Meisner, M., Fischer, S. ibid. p. 296. 7. Armstrong, B., Skegg, D., White, G., Doll, R. ibid 1976, ii, 8. 8. Mack, T M., Henderson, B. W., Gerkins, V. R., Arthaur, M., Babitiska. J. Pike, M. S. New Engl. J. Med. 1975, 292, 1366. 9. Lilienfeld, A. M., Chang, L., Thomas, D. B., Levin, M. L. Johns Hopkins med. J. 1976, 139, 41.