Antifungal Antibiotics

Antifungal Antibiotics GAVIN HILDICK-SMITH, M.D., M.R.C.P.

Mycotic infections in children are caused by a wide variety of fungi capable of involving any organ and causing diseases having a variety of signs and symptoms. As most fungal infections are amenable to therapy, it is important that the practicing physician not only readily diagnose the more common forms of fungal infection encountered in the pediatric age group, but also be aware of the less common types of infections so that optimum effective therapeutic measures can be utilized. 16 The fungi responsible for infections in children can be separated into three main groups with the organisms having a tendency to proliferate in a specific histological site of the involved organs due to their specific metabolic requirements and environmental needs. One group, Candida species, can thrive in the normal vagina and gastrointestinal tract and on the skin surface and causes local infection with alteration in environmental conditions. As the superficial proliferation of Candida species in disease is generally a surface phenomenon, the infection can readily be controlled with topical medicaments. The group of fungi commonly referred to as dermatophytes is responsible for tinea capitis, tinea pedis, tinea corporis and tinea manuum. These organisms, unlike Candida species, proliferate not on the skin surface, but within the epidermis or hair shafts and, as such, are not generally influenced by topical medication. They respond only to systemic or topical therapy where the antifungal agent used can penetrate to the histological site of fungal proliferation. Finally, there is a group of fungi which, having gained access to the body, usually via the respiratory tract or gastrointestinal tract or following trauma, proliferate at the site of invasion and spread to involve other body organs or cause systemic disease. The infections caused by these organisms are commonly referred to as the deep mycoses and can be treated at this time only by parenteral administration of amphotericin B. Most fungal infections can be successfully treated with one of three antibiotics, amphotericin B, griseofulvin and nystatin. Other antiPediatric Clinics of North America-Vol. 15, No.1, February, 1968

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Table 1.

Fungal Injections in Which Amphotericin B, Grisecifulvin and Nystatin are Effective

TYPE OF INFECTING FUNGI

SITE OF INFECTION

ANTIBIOTIC

ROUTE OF ADMINISTRATION

Systemic

Nystatin or Amphotericin B Amphotericin B

Topical Intravenous

Dermatophytes

Intradermal Hair

Griseofulvin Griseofulvin

Oral Oral

Fungi causing deep mycoses

Systemic

Amphotericin B

Intravenous

Candida species

Superficial

biotics and compounds with antifungal activity are available for use in treating fungus diseases, but apart from some topical medicaments for use in dermatophyte infection, they offer little of therapeutic value over and above the three antibiotics mentioned. The antifungal action of these three antibiotics is specific, and it is important that the identity of the infecting fungi be established, so that the appropriate antibiotic may be used correctly to treat the disease process. An indication of the antibiotic to be used in each type of fungal infection is presented in Table 1. AMPHOTERICIN B

Chemical Structure. Amphotericin B is one of a group of antibiotics produced by Streptomyces and generally known as polyene antibiotics. These are macrolides which contain four to seven conjugated double bonds in the lactone ring. They are divided into groups depending on the number of double bonds present in the molecule, amphotericin B having seven double bonds and belonging to the heptaene group, and nystatin, with four double bonds, being a tetraene. 1 Mode of Action. The molecule of polyene antibiotics possesses various substituents, such as amino acids and carboxyl groups, which are not common to all, and it is considered that the macrolide ring, with its lipophilic and hydrophilic portions, is responsible for their antifungal activity. The selective toxic action of amphotericin B and also nystatin against a wide variety of fungal species is considered to be dependent on the binding of the antibiotic with the sterols in the protoplast membrane of sensitive microorganisms. The binding of the polyenes to the cell membrane alters the permeability of the cell wall, causing leakage of the potassium ions and sugar with resultant death of the cell. 20 . Resistance. The development of resistance of fungi to high levels of amphotericin B and nystatin is rare, and has been produced only under experimental in vitro conditions. 15 Although failure of an infection to respond to therapy has suggested the development of resistance by the pathogen to the antibiotic, critical study of infecting organisms has,

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in fact, failed to demonstrate the development of resistance under clinical conditions. Sensitive Organisms. Amphotericin B inhibits a wide variety of fungi, as well as protozoa, flatworms, snails and higher algae, but not bacteria. The in vitro concentrations of amphotericin B that inhibit patho. genic fungi are listed in Table 2. Toxicity. Amphotericin B has been given orally to adults, in daily doses up to 16 gm., without adverse effect. Intravenous administration of the antibiotic, however, is nearly always associated with side effects and can be accompanied by serious toxicity. Amphotericin B must, therefore, be given only under hospital conditions, with close attention to the instructions supplied by the manufacturer. Side effects commonly associated with intravenous administration of the antibiotic are fever, rigors, anorexia, nausea, vomiting and abdominal pain. As the binding of polyene antibiotics to cell membranes containing sterols is not confined to microorganisms and can occur in cardiac, renal and red blood cells,3 cardiac irregularities and arrest have occurred when the parenteral dose given exceeded the recommended dose or the antibiotic was infused at a rapid rate. 16 Nephrotoxicity is the most important limiting factor in amphotericin B therapy, as renal damage occurs in direct relation to the total amount of antibiotic given during a course of therapy.s In a course of treatment in adults the total intravenous administration of up to 5 gm. of the antibiotic seldom produces significant renal damage, but partial loss of renal function has been noted with total doses from 5 to 10 gm. and severe damage with total doses of more than 10 gm. 32 In addition to renal damage occurring in association with amphoTable 2.

Minimal Inhibitory Concentrations (/Lg. per mi.)

rif Amphotericin B for Various Fungi ORGANISM

Candida albicans Sporotrichum schenckii (yeast phase) Microsporum canis M. audouinii M. gypseum Trichophyton mentagrophytes T. tonsurans T. rubrum Cryptococcus neoformans Epidermophyton floccosum Blastomyces dermatitidis Leishmania brasiliensis Coccidioides immitis Mucoraceae spp. Blastomyces brasiliensis Histoplasma capsula tum

AMPHOTERICIN B

<

1.9 0.07

7.3 0.9

>40.0 2.4 4.9 7.3 0.2 0.2 0.1 0.01

0.5 0.03->2.5 0.2 0.04

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tericin B therapy, anemia due to bone marrow suppression and changes in blood electrolytes have been observed, with impaired liver function a rare finding. Distribution and Excretion. Available data show that in adults the intravenous administration of 1 mg. per kg. of the antibiotic will produce blood serum levels of 0.3 to 6.0 p,g. per ml. 24 to 48 hours after infusion. The antibiotic is excreted slowly via the kidneys, 40 per cent of a single dose being recovered from the urine in a 7-day period following infusion, and its presence can be detected in the urine 2 months after terminating therapy. The antibiotic can be assayed in the cerebrospinal fluid following intravenous administration, the cerebrospinal fluid concentration increasing to 1.5 p,g. per ml. when the antibiotic is given intrathecally with intravenous therapy.7 Administration. Amphotericin B is administered intravenously, as it is not absorbed in the gastrointestinal tract. Intravenous amphotericin B is administered in a concentration of 1 mg. in 10 ml. of 5 per cent dextrose injection U.S.P. (pH above 5.0), utilizing an initial intravenous dose of 0.25 mg. per kg. administered over a period of 6 hours. The dose is then gradually raised to the average optimal dose of 1 mg. per kg., the maximum daily dose never exceeding 1.5 mg. per kg. The initial doses are given daily, with alternate-day administration when the optimum dosage level is reached, and with all doses given over a 6-hour period. Rapid administration of the antibiotic must be avoided, as the drug may produce convulsions, anaphylaxis, hypotension or cardiac fibrillation or arrest. During intravenous therapy, amphotericin B solution should be gently agitated every 15 to 30 minutes to avoid precipitation of the drug, and the solution should be protected from light. As the intravenous needle may become obstructed, a Y tube or three-way stopcock should be used, so that the needle can be flushed or small amounts of medication administered. Heparin may be added to the infusion fluid in adults in dose of 5 mg. per 100 ml. to prevent local thrombophlebitis, and in adults 25 to 100 mg. of the sodium succinate ester of hydrocortisone may be added to the infusion fluid to reduce the fever, chills and rigor that may occur. If the blood urea nitrogen level rises to over 40 mg. per 100 ml., the drug should be discontinued for 2 to 7 days until the level falls to normal. A 50 per cent reduction in creatinine clearance indicates significant renal damage, and medication should be withheld, with treatment reinstated if the test returns to normal in 2 to 3 weeks' time. Temporary withdrawal of medication may be indicated in association with intractable nausea, vomiting and anorexia. If medication has been withdrawn for 7 days or more, the dose should be restarted at a level of 0.25 mg. per kg. per day to avoid the severe reactions which can arise when reinitiating medication after it has been withheld for 7 or more

ANTIFUNGAL ANTIBIOTICS

111

days. Therapy should be continued for 1 to 3 months, and medication is withdrawn when the patient shows no signs of infection and is free of the invading organism or cannot tolerate the medication or shows signs of significant impairment of renal function. The information supplied by the manufacturer should be. carefully studied by the physician before initiating a course of intravenous amphotericin B. In seriously ill patients suffering from fungal meningitis or infection of the central nervous system, the intrathecal administration of amphotericin B is indicated, with a view to rapidly controlling the infection. In adults intrathecal therapy is initiated with a dose of 0.1 mg. and is increased to a dose of 0.5 mg. given every 48 to 72 hours, depending on the state of the patient and acceptance of medication. In adults 3 to 5 m!. of cerebrospinal fluid is withdrawn, and the antibiotic is dissolved in it prior to slow introduction of this solution into the intrathecal space. Transient urinary retention, hyporeflexia, paresthesia, paralysis and relaxation of sphincters can occur, as may local arachnoiditis and transient myelitis. These conditions are usually reversed on withdrawal of medication. The administration of amphotericin B intravenously in meningitis may be needed for 1 to 3 months to provide a cure. Clinical Usage. Amphotericin B is effective in the treatment of histoplasmosis, coccidioidomycosis, cryptococcosis, North and South American blastomycosis and disseminated candidiasis. In addition, it has merit in the treatment of sporotrichosis, mucormycosis and maduromycosis. The antibiotic is ineffective in the treatment of nocardiosis and actinomycosis. In infants and children systemic fungal disease is uncommon, histoplasmosis, coccidioidomycosis and systemic candidiasis being seen most often. Systemic fungal infection is encountered in patients with underlying diseases, such as leukemia and Hodgkin's disease, or undergoing cytotoxic drug therapy, when the inherent capability of the patient to resist infection is impaired. 22 Histoplasmosis. While histoplasmosis may not be a serious illness in older children, it frequently causes disseminated disease in children under 1 year and requires active therapy. A satisfactory therapeutic result was reported in eight children with disseminated histoplasmosis in which intravenous amphotericin B was given in optimum dose of 1 mg. per kg. per day for a period of 7 to 32 days.21 The available data indicate that an intravenous dose of 1 mg. per kg. should be given on alternate days in such cases for an optimum period of 4 weeks. Prompt diagnosis of the disease in children and adequate intravenous therapy with amphotericin B should achieve a satisfactory therapeutic result in most cases. In contrast, in a study of 67 cases of chronic histoplasmosis in adults treated with intravenous amphotericin B, 12 patients recovered, 33 were improved, 9 relapsed, 7 were unaltered, 4 died of other causes and 2 died of the disease. 12 Coccidioidomycosis. A report on nine children under 10 years of

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age who were suffering from disseminated coccidioidomycosis and were treated with intravenous amphotericin B showed that all patients recovered from infection. 32 A total dose of 105 to 3795 mg. of antibiotic was given to the children, the average total dose being 1266 mg. Management of disseminated coccidioidomycosis usually calls for more prolonged therapy than is utilized for primary coccidioidomycosis and continuing follow-up to rapidly detect relapses. Cryptococcosis. Cryptococcosis in children is an uncommon disease, only 42 cases being reported in the world literature as of 1944. Three patients with disseminated cryptococcosis were treated with amphotericin B, of whom one suffering from leukemia died. The two children who recovered received a total of 40 mg. of amphotericin B intravenously, utilizing a daily intravenous dose of 0.5 mg. per kg.28 A review of the data on amphotericin B treatment in adults suffering from cryptococcosis shows that in one series of 47 patients 50 per cent of those treated were free of infection 6 months after therapy. Systemic Candidiasis. Systemic candidiasis occurs commonly in children with leukemia or Hodgkin's disease or in those receiving cytotoxic drugs. Generalized cutaneous infection can occur in association with granuloma 26 and also with acrodermatitis enteropathica. In all cases of disseminated disease, intravenous amphotericin B should be given in recommended doses until the patient shows no signs of infection and the organism cannot be cultured from blood or urine. 29 Cutaneous Candidiasis. Amphotericin B is available as a topical medicament for the treatment of cutaneous candidiasis. The indications and method of use of topical amphotericin B in cutaneous candidiasis are similar to that for nystatin (see below).

GRISEOFULVIN

Griseofulvin was isolated in 1939 and its structure described, but its therapeutic value was not realized until 1958. 13 Chemical Structure and Mode of Action. Griseofulvin is 7-chloro-2', 4, 6-trimethoxy-6'-methylspiro [benzofuran-2 ( 3H ) ,1'- ( 2) cyclohexene] -3, 4'dione.2 In spite of the dennition of griseofulvin's structure, only limited research has been done on its mode of action. Exposure of mycelia of sensitive fungi to griseofulvin shows that existing hyphae are unaltered by the antibiotic, but that newly produced hyphae are affected and show a characteristic deformation of growth, the cytoplasm shrinking and lipid granules becoming more pronounced. Griseofulvin is not translocated within fungal hyphae and only involves hyphae in direct contact with the antibiotic. Griseofulvin appears to interfere with fungal mitosis, impairs synthesis of protein and nucleic acid and causes the breakdown of intracellular organelle membranes. 6

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Table 3.

Minimal Inhibitory Concentrations oj GriseoJulvinjor Dermatophytes CONCENTRATION AT ORGANISM

Microsporum canis M. gypseum M. audouinii Epidermophyton floccosum Trichophyton mentagrophytes T. rubrum T. tonsurans T. megninii T. gallinae T. schoenleinii T. violaceum T. verrucosum T. concentricum

72

HOURS

(lLg. per mI.)

0.20-0.24 0.42-0.46 0.40-0.46 0.38-0.42 0.38-0.42 0.14-0.18 0.28-0.32 0.30-0.34 0.40-0.44 0.34-0.38 0.36-0.40 0.28-0.30 0.26-0.30

Resistance. Fungi resistant to griseofulvin have been produced in vitro, but are rarely seen in therapy.6. 14 Although a wide range of fungi are sensitive to griseofulvin, its value lies in its inhibition of dermatophytic fungi. It is ineffective against bacteria, Candida species and fungi causing deep mycoses. The dermatophytes sensitive to griseofulvin are listed in Table 3. Toxicity. The oral administration of griseofulvin in total daily doses up to 5 gm. for prolonged periods in innumerable patients has produced a remarkably low incidence of significant adverse effects. Griseofulvin, however, interferes with the activity of coumarin congeners by impairing their effect in lowering blood coagulation. 1o Griseofulvin therapy has been reported to increase blood and urinary porphyrin levels and should therefore not be used in patients suffering from porphyria. l l A wide variety of minor side effects have been reported, but in most instances have been relieved on withdrawal of the antibiotic. They include skin lesions, such as photosensitivity and urticaria, gastrointestinal disturbances and symptoms such as fatigue, nausea and fainting. 2 Distribution and Excretion. Orally administered griseofulvin is partially absorbed from the gastrointestinal tract, studies in rats showing that only 5.4 per cent of a given dose was absorbed. 5 The amount of griseofulvin absorbed from the bowel is increased when a diet high in fat is given. The microcrystalline form is more readily absorbed than the regular form and produces blood levels twice as great as those of the regular form when given in comparable doses. 9 Although in vitro studies of rat tissues show that the antibiotic is incorporated in the major organs, it appears to be metabolized only by the liver.30 Blood level studies in man show that following a single oral dose of 1 gm. of the regular form of the antibiotic, peak serum levels are noted 4 hours after ingestion, with trace levels noted 72 hours after

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medication. all The antibiotic can be detected in lower layers of the stratum corneum 2 to 3 days after medication35 and halfway through the stratum corneum after 2 to 3 weeks of therapy.27 Clinical Application. In clinical usage griseofulvin is effective in treatment of infections of hair, skin and nails caused by dermatophytes, but is ineffective in the treatment of candidal and bacterial infection. Tinea Capitis. Orally administered griseofulvin is the treatment of choice in tinea capitis. Daily dose of 250 mg. of microcrystalline griseofulvin given to children under 10 years and 500 mg. for children over 10 years will cure most patients.25 Maintaining closely cut hair and daily application of Whitfield's ointment, tolnaftate cream or solution or Desenex Ointment will minimize the risk of cross infection during therapy. If patients fail to respond to therapy, consideration must be given to failure to take the medication, poor absorption or distribution in tissues and, lastly, resistance of the pathogen to the antibiotic. Effectiveness of oral griseofulvin in management of tinea capitis is demonstrated by the cure rate of 96.2 per cent in 766 patients in whom the microcrystalline form of the antibiotic was given in a dose of 12.5 mg. per kg. and regular material in 25 mg. per kg. per day.2.34 Under certain circumstances where repeated daily doses of griseofulvin are impractical, the use of a single large dose of the antibiotic is of value. A single 3 gm. dose of regular griseofulvin in conjunction with a daily application of Whitfield's ointment cured 81 per cent of cases of tinea capitis in 4 weeks. aa Tinea Corporis. Glabrous skin lesions caused by dermatophytic fungi can be effectively treated by daily or twice daily topical application of tolnaftate cream or solution, Desenex Ointment or Whitfield's ointment. Daily oral administration of 250 mg. of microcrystalline griseofulvin in children under 10 years and 500 mg. in older children will eradicate the infecting organisms in 2 to 6 weeks. As relapse is liable to occur, close surveillance of patients is indicated. A review of pooled data on 113 cases of dermatophyte skin infection (excluding Trichophyton concentricum) confined to trunk and extremities showed a cure rate of 98 per cent following oral griseofulvin therapy.2 Tinea Pedis, Tinea Cruris and Tinea Manuum. Microcrystalline griseofulvin is effective in treating tinea pedis, tinea cruris and tinea manuum, utilizing an oral daily dose of 250 mg. for children under 10 years and 500 mg. for older children. Medication is maintained until symptomatic relief is procured and the pathogenic organism eradicated. In acute lesions the additional topical application of tolnaftate or Desenex lotions two or three times a day will help alleviate the symptoms. The topical use of tolnaftate or Desenex Ointment alone two or three times a day will usually control infection in 5 to 10 weeks. Dermatophytic lesions of the groin and feet are most prone to recur, and medication should be reinstated when relapse or reinfection occurs. The daily prophylactic use of talc dusting powders alone or dusting powders con-

115

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taining antifungal agents will help to minimize local cutaneous trauma and thereby minimize the risk of infection. 4 A review of 304 cases of infection of palms or soles by T. rubrum or T. mentagrophytes treated with oral griseofulvin showed a 53.3 per cent cure rate, failures being associated with nail involvement. 2 Onychomycosis. Oral administratIon of griseofulvin is effective in the management of nail infection due to dermatophytes, but the medication must be maintained for a significant period of time to procure an optimum therapeutic result. The dosage of griseofulvin is that indicated for other dermatophytic infections, except that treatment of fingernails is maintained for a minimum of 4 months, with 6 to 12 months of medication indicated in the treatment of the toenails. Regular surveillance of the treated lesion is indicated, with procurement of cultures from the site of infection to determine when to terminate therapy. In those patients failing to respond to therapy, a twice daily application of tolnaftate cream or solution should be tried in association with griseofulvin therapy.25 In a series of 158 cases of fingernail infection 56.9 per cent were cured, while only 16.7 per cent of cases of toenail involvement were cured in 299 cases. 2

NYSTATIN

Nystatin was isolated in 1949 from a Streptomyces obtained from a dairy farm in Virginia and has an antifungal spectrum of action similar to that of amphotericin B. Chemical Structure and Mode of Action. Nystatin is a tetraene polyene antibiotic, as it has four conjugated double bonds in its molecular structure. 16 Studies on the mode of action of the antibiotic against sensitive fungi show that, like amphotericin B, it binds to the sterol component of the protoplastic cell membrane, causing loss of small cellular ions and sugars. Resistance of Candida species to the antibiotic has been produced in vitro,16 but extensive therapeutic use of the antibiotic in the management of superficial candidal lesions has, as yet, failed to produce resistant organisms. The likelihood of fungi developing resistance to polyene antibiotics in clinical medicine is small, and no data are available to indicate that such resistance is likely to be a problem in the therapeutic use of these antibiotics. Nystatin, like other polyene antibiotics, inhibits a wide range of fungi, which include Candida species as well as fungi responsible for dermatophytic and deep mycotic infections. Although nystatin can be administered orally in significant doses without adverse effect and has been combined with antibacterial antibiotics to suppress intestinal candidal growth, the antibiotic is not absorbed from the gastrointestinal tract, and therefore has no therapeutic value in the treatment of systemic

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GAVIN HILDICK-SMITH

fungal diseases. Nystatin's intravenous toxicity is such that its therapeutic use in man cannot be seriously entertained. Accordingly, nystatin's therapeutic value lies in the treatment of superficial candidal infection, whether it be of the skin or mucous surfaces. The antibiotic is ineffective in the topical treatment of cutaneous dermatophyte infections. The extensive uses of nystatin on the skin or mucous surfaces have shown the antibiotic to be well tolerated and significantly free of local irritation or discomfort; it rarely produces sensitization. Clinical Use. Cutaneous Candidiasis. The topical application of an anticandidal preparation to candidal lesions23 effectively controls the infection, as the organisms proliferate essentially on the skin surface and the antifungal compounds can reach them in inhibitory concentration. 31 The topical application of nystatin, in cream, ointment or lotion, alone or in combination with a glucocorticosteroid, applied three or four times a day, will readily control candidal infection of the skin. Therapy must be continued for at least 7 days, as premature cessation of medication is commonly associated with relapse of infection. 1s Elimination of the pathogenic Candida species from the lesion under therapy is an indication for withdrawal of medication. When candidal infection recurs, it is important to determine whether the patient is suffering from an underlying disease which might predispose to the infection. In intertrigenous lesions the utilization of talc containing nystatin applied several times a day to the skin site may help minimize the risk of infection. Candidal Diaper Dermatitis. Candida species can be isolated from cases of diaper dermatitis in which they are merely contaminants; however, where skin scrapings show that candidal hyphae are present, the organism is responsible for the clinical lesion. Candidal diaper dermatitis is frequently seen in newborn infants in association with oral candidiasis, which in tum is related to the presence of Candida species in the birth canal of the mother.11l, 24 In older children the pathogen is usually present in the stools and contaminates the perineum, where under optimum conditions it will proliferate to cause a lesion. The topical application of nystatin ointment two or three times a day for 7 days will usually control the candidal infection. In acute or severe cases, concomitant application of preparations containing nystatin combined with glucocorticosteroids, such as Mycolog Cream or Lotion, will hasten resolution of the lesion. The prevention of recurrence of lesions requires careful attention to local hygiene and changing of diapers as soon as they are soiled, the local application of talc containing nystatin being expected to help minimize the risk of reinfection. Oral Candidiasis. Oral candidiasis is frequently seen in infants; the pathogen is present in the mother's vaginal tract. 16 Oral candidiasis is readily treated by instillation of 1 to 2 ml. of nystatin for suspension applied to the lesion three to four times a day for 7 to 14 days. The infection usually responds rapidly to this therapy. Relapse is likely if

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ANTIFUNGAL ANTIBIOTICS

medication is not continued for an adequate period, and when it occurs calls for further treatment. Intestinal Candidiasis. Intestinal candidiasis is uncommon, but can be responsible for gastrointestinal signs and symptoms in infants and small children. The lesion can be associated with oral candidiasis and responds readily to the oral administration of 1 to 2 ml. of nystatin for suspension given three to four times a day for 7 to 14 days,17

REFERENCES 1. Abraham, E. P.: The chemistry of new antibiotics. Am. J. Med., 39:692, 1965. 2. Anderson, D. W.: Griseofulvin: Biology and clinical usefulness. Ann. Allergy, 23:103,1965. 3. Arora, H. R.: Effects of polyene antibiotics on the heart. A study with nystatin and lagasin. Med. Pharmacal Exp. (Basel), 13:155, 1965. 4. Baer, R. L., and Rosenthal, S. A.: The biology of fungous infections of the feet. J.A.M.A., 197:1017, 1966. 5. Bedford, C., et al.: Studies on the biological disposition of griseofulvin, an oral antifungal agent; A.M.A. Arch. Dermat., 81:735, 1960. 6. Bent, K. J., and Moore, R. H.: The mode of action of griseofulvin. In Newton, B. A., and Reynolds, P. E., eds.: Biochemical Studies of Antimicrobial Drugs. Cambridge, Cambridge University Press, 1966, p. 82. 7. Butler, W. T.: Pharmacology, toxicity and therapeutic usefulness of amphotericin B. J.A.M.A., 195:127, 1966. 8. Butler, W. T., et al.: Nephrotoxicity of amphotericin B. Early and late effects in 81 patients. Ann. Intern. Med., 61:175, 1964. 9. Crounse, R. G.: Human pharmacology of griseofulvin. The effect of fat intake on gastrointestinal absorption. J. Invest. Derm., 37:529, 1961. 10. Cullen, S. I., and Catalano, P. M.: Griseofulvin-warfarin antagonism. J.A.M.A., 199:582, 1967. 11. De Matteis, F., and Rimington, C.: Disturbance of porphyrin metabolism caused by griseofulvin in mice. Brit. J. Derm., 75:91, 1963. 12. Furcolow, M. L.: The use of amphotericin B in blastomycosis, cryptococcosis and histoplasmosis. Med. Clin. N. Amer., 47:1119, 1963. 13. Gentles, J. C.: Experimental ringworm in guinea pigs: Oral treatment with griseofulvin. Nature, 182:476, 1958. 14. Grin, E. I.: Investigations on the adaptivity of dermatophytes to griseofulvin. Mycopathologia, 30:31, 1966. 15. Habeka, E. K., and Solotorovsky, M.: Development of resistance to polyene antibiotics in Candida albicans. J. Bact., 89:1533, 1965. 16. Hildick-Smith, G., Blank, H., and Sarkany, I.: Fungus Diseases and Their Treatment. Boston, Little, Brown & Co., 1964. 17. Hildick-Smith, G.: Candidal enteritis. elin. Ped., 8:28, 1964. 18. Kozinn, P. J.: Cutaneous candidiasis in early infancy and childhood. Pediatrics, 20:827, 1957. 19. Kozinn, P. J., et al.: Incidence and pathogenesis of neonatal candidiasis. Pediatrics, 21:421,1958. 20. Lampen, J. 0.: Interference by polyenic antifungal antibiotics (especially nystatin and fllipin) with specific membrane function. In Biochemical Studies of Antimicrobial Drugs. Cambridge, Cambridge Univ. Press, 1966, p. 111. 21. Little, J. A.: Intravenous amphotericin therapy in children with histoplasmosis. J. Kentucky Med. Assoc., 60:965, 1962. 22. Louria, D. B., Blevins, A., Armstrong, D., et al.: Fungemia caused by "nonpathogenic" yeasts. Arch. Intern. Med., 119:247, 1967. 23. Maibach, H. I., and Kligman, A. M.: The biology of experimental human cutaneous moniliasis. Arch. Derm., 85:233, 1962.

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24. Rohatiner, J. J.: Relationship of Candida albicans in the genital and anorectal tracts. Brit. J. Vener. Dis., 42:197,1966. 25. Samman, P. D.: Management of disorders of the nails. Brit. Med. J., 11:1122,1966. 26. Saul, A.: Candidosis granulomatosa. Rev. Med. Hosp. Gen. (Mex.), 29:213, 1966. 27. Shimazaki, J.: Studies on griseofulvin. II. Skin griseofulvin levels after oral administration. J. Med. Mycol., 6: 186, 1965. 28. Siewers, C. M. F., and Cramblett, H. G.: Cryptococcosis (torulosis) in children. Pediatrics, 34:393, 1964. 29. Simon, V. C.: tJber eine durch Amphotericin B geheilte Candida sepsis eines Friihgeborenen. Annal. Paed. (Basel), 204:406, 1965. 30. Symchowicz, S., and Wong, K. K.: Metabolism of griseofulvin-HC studies in vitro. Biochem. Pharmacol., 15: 1601, 1966. 31. Warin, R. P.: Fungal infections of the skin. Brit. Med. J., 2:1307, 1966. 32. Winn, W. A.: Coccidioidomycosis and amphotericin B. Med. CHn. N. Amer., 47:1131, 1963. 33. Zaias, N., et al.: Evaluation of microcrystalHn griseofulvin therapy in tinea capitis. J.A.M.A., 198:805, 1966. 34. Ziprkowski, L., et al.: Tinea capitis eradication campaign in Israel. Hare-Fuah, 70:418, 1966. 35. Ziprkowski, L., et al.: The effect of griseofulvin in hereditary porphyria cutanea tarda. Arch. Derm., 93:21, 1966. Johnson & Johnson New Brunswick, N. J. 08903