Potato dextrose agar antifungal susceptibility testing for yeasts and molds: evaluation of phosphate effect on antifungal activity of CMT-3

The broth macrodilution method (BMM) for antifungal susceptibility testing, approved by the National Committee for Clinical Laboratory Standards (NCCLS), was found to have deficiencies in testing of the antifungal activity of a new type of antifungal agent, a nonantibacterial chemically modified tetracycline (CMT-3). The high content of phosphate in the medium was found to greatly increase the MICs of CMT-3. To avoid the interference of phosphate in the test, a new method using potato dextrose agar (PDA) as a culture medium was developed. Eight strains of fungi, including five American Type Culture Collection strains and three clinical isolates, were used to determine the MICs of amphotericin B and itraconazole with both the BMM and the PDA methods. The MICs of the two antifungal agents determined with the PDA method showed 99% agreement with those determined with the BMM method within 1 log(2) dilution. Similarly, the overall reproducibility of the MICs with the PDA method was above 97%. Three other antifungal agents, fluconazole, ketoconazole, and CMT-3, were also tested in parallel against yeasts and molds with both the BMM and the PDA methods. The MICs of fluconazole and ketoconazole determined with the PDA method showed 100% agreement within 1 log(2) dilution of those obtained with the BMM method. However, the MICs of CMT-3 determined with the BMM method were as high as 128 times those determined with the PDA method. The effect of phosphate on the antifungal activity of CMT-3 was evaluated by adding Na2HPO4 to PDA in the new method. It was found that the MIC of CMT-3 against a Penicillium sp. increased from 0.5 microg/ml (control) to 2.0 microg/ml when the added phosphate was used at a concentration of 0.8 mg/ml, indicating a strong interference of Na2HPO4 with the antifungal activity of CMT-3. Except for fluconazole, all the other antifungal agents demonstrated clear end points among the yeasts and molds tested. Nevertheless, with its high reproducibility, good agreement with NCCLS proposed MIC ranges, and lack of interference of phosphate, the PDA method shows promise as a useful assay for antifungal susceptibility testing and screening for new antifungal agents, especially for drugs that may be affected by high (supraphysiologic) phosphate concentrations.     

Telephone subsidy: an effective incentive for successful participation in home memory monitor study

CONTEXT: The Collaborative Home Infant Monitoring Evaluation (CHIME) study enrolled healthy term infants and 3 groups of infants considered to be at increased risk for sudden infant death syndrome to evaluate apnea and bradycardia events in the home. Mother-infant pairs without a telephone were ineligible for enrollment. OBJECTIVE: To determine whether mother-infant pairs who were offered a telephone subsidy would agree to enroll in CHIME and achieve protocol compliance rates comparable with those of matched subjects able to afford telephones. DESIGN: Thirty-one telephone subsidy subjects were retrospectively compared with 55 control subjects matched for study group, site, birth weight, and maternal race, age, and education. SETTING: Collaborative Home Infant Monitoring Evaluation clinical research centers in Honolulu, Hawaii, and Toledo, Ohio. INTERVENTION: Provision of telephone subsidy to otherwise eligible enrollees for CHIME. MAIN OUTCOME MEASURES: Frequency of compliance with protocol requirements for follow-up evaluations and for extent of home monitoring. RESULTS: Subsidy subjects achieved protocol completion rates that were comparable with those of control subjects, for developmental assessments at 56 and 92 weeks postconceptional age (PCA), and for the polysomnogram. Unexpectedly, however, subsidy subjects were more likely to have a developmental assessment at 44 weeks PCA (P =.02), as well as a cry analysis (P =.04). They were also more likely to use the CHIME home monitor for more hours during weeks 2 through 5 (P =.004), have a higher percentage using the monitor for 10 or more hours per week during weeks 2 through 5 (P =.009), and have a higher total number of days of monitor use throughout 6 months (P <.001). Mean cost of the subsidy was $3.25 per day of monitor use, and monitor use per day was directly related to total cost of the subsidy (P =.02). CONCLUSIONS: Telephone subsidy is an effective financial incentive. At least within the context of the CHIME study, telephone subsidy enhanced access to health care, and in some categories it resulted in enhanced protocol compliance.     

Excessive matrix metalloproteinase activity in diabetes: inhibition by tetracycline analogues with zinc reactivity

Diabetes mellitus in rats is characterized by excessive activity of several matrix metalloproteinases (MMPs), notably collagenase(s) and gelatinase(s), in skin, gingiva, and other tissues. A number of tetracyclines (TCs), both antimicrobial compounds as well as chemically modified non-antimicrobial TC analogues (CMTs) are known to possess potent inhibitory activity against these enzymes. Three conventional antimicrobial TCs and six CMTs were used in this study. In vitro, doxycycline was shown to possess higher inhibitory capacity (i.e. lower IC(max)) against diabetic rat skin collagenase than either minocycline or tetracycline HCl. Addition of excess zinc partially reversed the proteinase inhibition by TCs. In vivo, using rats made diabetic with streptozotocin (STZ), oral administration of various TCs led to decreased weight loss and substantial reductions in the activity of both skin collagenase and skin gelatinase (primarily MMP-9, 92 kDa) without affecting blood glucose. Using an in vitro spectrophotometric technique, the Zn(++) reactivity of several CMTs was assessed and found to be positively related to the potency of these compounds as MMP inhibitors. One particular CMT (CMT-5, pyrazole analogue), which is neither antimicrobial nor capable of binding metal cations, did not inhibit the MMPs. TCs have potential utility in management of diabetic complications mediated by excessive activity of MMPs.     

Is the U.S. experiencing an incipient epidemic of hallucinogen use?

NHSDA and MTF survey data indicate "epidemic"-like growth in hallucinogen use from 1992-1996 and associated increases in cocaine, crack, heroin and amphetamine use. These trends might have resulted from a proliferation of raves and dance clubs in the U.S. as occurred in Europe and elsewhere, although in contrast to evidence regarding European experiences the American epidemic involves primarily teens as opposed to persons in their twenties and involves primarily use of LSD as opposed to MDMA. This analysis highlights the need for further research into the context, significance, and consequences of these recently popular American drug use practices.     

The adrenal and the metabolic syndrome

This review discusses the possible interrelationships between adrenal steroid hormones and the metabolic syndrome. Abnormal regulation of the hypothalamic-pituitary-adrenal axis has been proposed. Studies in the United Kingdom associated the metabolic syndrome with low birth weight and hyperactivity of the entire axis. In Italy, increased pituitary responsiveness to stimulation with vasopressin and corticotrophin-releasing hormone was demonstrated in women with central obesity. Swedish researchers have reported that increased stress responses of the axis correlated with a less variable but decreased cortisol level. An allele of the glucocorticoid receptor was also associated with various components of the metabolic syndrome. Evidence also suggests that central obesity is associated with an increased peripheral conversion of cortisol to cortisone and subsequent feedback stimulation of the axis. On the other hand, central fat may have an increased local metabolism in the direction of cortisol. Roles for dehydroepiandrosterone and aldosterone in the syndrome have also been proposed.     

Nuclear foci of mammalian recombination proteins are located at single-stranded DNA regions formed after DNA damage

A sensitive and rapid in situ method was developed to visualize sites of single-stranded (ss) DNA in cultured cells and in experimental test animals. Anti-bromodeoxyuridine antibody recognizes the halogenated base analog incorporated into chromosomal DNA only when substituted DNA is in the single strand form. After treatment of cells with DNA-damaging agents or gamma irradiation, ssDNA molecules form nuclear foci in a dose-dependent manner within 60 min. The mammalian recombination protein Rad51 and the replication protein A then accumulate at sites of ssDNA and form foci, suggesting that these are sites of recombinational DNA repair.     

Pet sensitivities in asthmatic children

A history of pet contact and/or apparent clinical sensitivity was obtained in 65 (55%) of 118 unselected asthmatic children. These 65 children were skin tested and their sera examined for specific IgE using the radioallergosorbent test. Those children who had apparent clinical sensitivities had larger skin test reactions and were more likely to have positive specific IgE results than those without apparent sensitivities. Positive skin tests were very common (80%), but the larger the skin test reaction (weal diameter greater than 4 mm diameter) the more likely was there to be a positive history or a positive specific IgE result. Hence a large skin test reaction can provide a helpful pointer to animal allergy of clinical importance. Commercially available animal extracts have limitations for diagnostic tests. A questionnaire survey of 150 day schools emphasized the potential opportunities for contact with animal allergens at school.