Michael E. Klepser

Influence of Test Conditions on Antifungal Time-Kill Curve Results: Proposal for Standardized Methods

ABSTRACT This study was designed to examine the effects of antifungal carryover, agitation, and starting inoculum on the results of time-kill tests conducted with various Candida species. Two isolates each of Candida albicans, Candida tropicalis, and Candida glabrata were utilized. Test antifungal agents included fluconazole, amphotericin B, and LY303366. Time-kill tests were conducted in RPMI 1640 medium buffered with morpholinepropanesulfonic acid (MOPS) to a pH of 7.0 and incubated at 35°C. Prior to testing, the existence of antifungal carryover was evaluated at antifungal concentrations ranging from 1× to 16× MIC by four plating methods: direct plating of 10, 30, and 100 μl of test suspension and filtration of 30 μl of test suspension through a 0.45-μm-pore-size filter. Time-kill curves were performed with each isolate at drug concentrations equal to 2× MIC, using a starting inoculum of approximately 105 CFU/ml, and incubated with or without agitation. Last, inoculum experiments were conducted over three ranges of starting inocula: 5 × 102 to 1 × 104, >1 × 104 to 1 × 106, and >1 × 106 to 1 × 108 CFU/ml. Significant antifungal carryover (>25% reduction in CFU/milliliter from the control value) was observed with amphotericin B and fluconazole; however, carryover was eliminated with filtration. Agitation did not appreciably affect results. The starting inoculum did not significantly affect the activity of fluconazole or amphotericin B; however, the activity of LY303366 may be influenced by the starting inoculum. Before antifungal time-kill curve methods are routinely employed by investigators, methodology should be scrutinized and standardized procedures should be developed.

Assessment of Patients' Perceptions and Beliefs Regarding Herbal Therapies

We evaluated the demographics and beliefs regarding safety and efficacy of herbal therapy among individuals in Iowa and assessed the willingness to discuss the use of these products with health care providers. We distributed 1300 surveys to two random samples: patients attending eight clinics, and residents of the state (mailing). Data were categorized according to herb use and compared between users and nonusers. The response rate was 61% (794 people), with 41.6% of respondents reporting herb use. They were predominately white women and were likely to have had education beyond high school (p<0.05). Their use of prescription drugs was high (p<0.05). Although users rated safety and efficacy of herbs higher than nonusers (p<0.05), both groups believed that health care providers should be aware of use and would provide this information.

Novel triazole antifungal agents

The risk of opportunistic infections is greatly increased in patients who are immunocompromised due to AIDS, cancer chemotherapy and organ or bone marrow transplantation. Candida albicans is often associated with serious systemic fungal infections, however other Candida species such as Candida krusei, Candida tropicalis and Candida glabrata, as well as Cryptococcus neoformans and filamentous fungi such as Aspergillus, have also emerged as clinically significant fungal pathogens. Two triazole antifungal agents, fluconazole and itraconazole, were introduced over a decade ago and since then have been used extensively for the prophylaxis and treatment of a variety of fungal infections. Although both drugs are effective and have their place in therapy, limitations regarding the utility of these agents do exist. For example, fluconazole is not effective for the prophylaxis or treatment of Aspergillus species and has limited activity against C. krusei and C. glabrata. The use of itraconazole has been limited secondary to concerns regarding unpredictable bioavailability. The rising incidence of fungal infections and the reported increase of non-albicans candidal infections noted over the past two decades highlight the need for new antifungal agents with improved spectra of activity. Several new triazole agents are in various phases of preclinical and clinical trials and may be available for human use in the near future. Three such agents voriconazole, posaconazole and ravuconazole are reviewed and compared with existing agents.

Antifungal dynamics of LY 303366, an investigational echinocandin B analog, against Candida ssp.

Two isolates each of Candida albicans, Candida tropicalis, and Candida glabrata were selected for time-kill curve testing against LY 303366 at concentrations ranging from 0.125 x MIC to 16 x MIC. RPMI 1640 buffered morpholinepropanesulfonic acid (MOPS) was utilized as growth medium. Samples were obtained at predetermined time points over 24 hours and streaked for colony count determination. Against C. albicans (one strain) and C. glabrata isolates, LY 303366 exhibited fungicidal (> or = three log10 reduction in CFU) activity. In contrast, fungistatic activity was observed with LY 303366 against C. albicans (one strain) and C. tropicalis isolates at all of the multiples of the MIC tested. With the exception of one C. glabrata strain, the rate and extent of activity against test isolates was not enhanced with concentrations exceeding the MIC. Our data indicate that maximal antifungal activity with LY 303366 may be achieved by optimizing the time of fungal exposure to the drug. Additionally, these data suggest that use of the current interpretive endpoint for MICs in RPMI may underestimate the antifungal activity of LY 303366. Thus, the MIC endpoint may need to be re-evaluated, or perhaps an alternative media, such as antibiotic medium #3, should be utilized for determination of LY 303366 MICs.

Update on antifungal resistance

Overuse of antifungal agents has resulted in the selection of naturally resistant Candida species, as well as expression of resistance from previously susceptible species resulting from genetic mutations and/or selection of resistant subpopulations. Strategies for the appropriate use of antifungal agents need to be developed to prevent further development of resistance.

Drug Treatment of HIV-Related Opportunistic Infections

SummaryThe AIDS epidemic has led to the emergence of several disease entities which in the pre-AIDS era were rare or seemingly innocuous. Experience of treating these diseases varies. In some instances, such as Pneumocystis carinii pneumonia, there is an abundance of published literature to direct our course of action. However, for many of these newly recognised diseases our treatment experience is limited. Furthermore, in many instances, well controlled trials evaluating treatment modalities in the AIDS population are lacking. We have identified 13 disease entities (P carinii pneumonia, toxoplasmosis, cryptococcosis, histoplasmosis, Mycobacterium tuberculosis, Mycobacterium avium complex, cytomegalovirus, coccidioidomycosis, isosporiasis, candidosis, Kaposi’s sarcoma, herpes simplex virus, and varicella zoster virus) and have reviewed the current literature with regard to their treatment.

Comparative Bactericidal Activities of Ciprofloxacin, Clinafloxacin, Grepafloxacin, Levofloxacin, Moxifloxacin, and Trovafloxacin against Streptococcus pneumoniae in a Dynamic In Vitro Model

ABSTRACT Several new quinolones that exhibit enhanced in vitro activity against Streptococcus pneumoniae have been developed. Using a dynamic in vitro model, we generated time-kill data for ciprofloxacin, clinafloxacin, grepafloxacin, levofloxacin, moxifloxacin, and trovafloxacin against three isolates of quinolone-susceptible S. pneumoniae. Three pharmacokinetic profiles were simulated for each of the study agents (0.1, 1, and 10 times the area under the concentration-time curve [AUC]). Target 24-h AUCs were based upon human pharmacokinetic data resulting from the maximal daily doses of each agent. Ciprofloxacin was the least active agent against all three isolates. With regimens that simulated the human 24-h AUC, ciprofloxacin resulted in an initial, modest decline in the numbers of CFU per milliliter; however, by 48 h the numbers of CFU per milliliter returned to or exceeded the starting inoculum. At the AUC, levofloxacin resulted in variable bacteriostatic and bactericidal activities against the isolates. The remaining agents yielded bactericidal (99.9% reduction) activity by 48 h with regimens that simulated the AUC. At 0.1 time the AUC ciprofloxacin and levofloxacin produced no inhibitory effect, grepafloxacin exhibited bacteriostatic activity, trovafloxacin had mixed static and cidal activities, and clinafloxacin and moxifloxacin caused significant reductions in the numbers of CFU per milliliter by 48 h. All six agents produced cidal activity at 10 times the AUC. In this dynamic in vitro model of infection, the quinolones demonstrated various degrees of activity against S. pneumoniae. The rank order of activity, with respect to bactericidal effect, was ciprofloxacin (least active) ≪ levofloxacin < grepafloxacin, trovafloxacin < clinafloxacin and moxifloxacin (most active). The rank order of the agents with respect to the selection of resistance was ciprofloxacin (most likely) > grepafloxacin, moxifloxacin, and trovafloxacin > levofloxacin > clinafloxacin.

Comparison of bactericidal activities of intermittent and continuous infusion dosing of vancomycin against methicillin-resistant Staphylococcus aureus and Enterococcus faecalis.

Study Objective. To describe the pharmacokinetic profiles of vancomycin administered by continuous infusion and intermittent dosing and compare the duration of activity of the regimens.

In vitro pharmacodynamic characteristics of flucytosine determined by time-kill methods☆

Two Candida albicans isolates, three non-albicans Candida isolates (Candida glabrata, Candida krusei, and Candida tropicalis), and one Cryptococcus neoformans isolate were evaluated by time-kill methods to characterize the relationship of flucytosine concentrations to antifungal activity and the duration of the post-antifungal effect (PAE). Against Candida and Cryptococcusisolates, flucytosine at concentrations > 1 x MIC exhibited fungistatic (</=99% reduction in CFU) activity over a 24-h time-period. The rate and extent of fungistatic activity of flucytosine against all isolates was generally not increased when 5-FC concentrations exceeded 4 x MIC. A notable PAE was detected for flucytosine against both Candida and Cryptococcus species that persisted 2 to 4 h. These in vitro data suggest that flucytosine is predominately a concentration-independent fungistatic agent at clinically achieved serum concentrations. This pharmacodynamic characteristic coupled with the persistent PAE and the relatively long half-life of flucytosine in humans (> 5 h), suggests lower daily dosing may possible without loss of antifungal efficacy.

Pharmacoeconomic Impact of Once‐Daily Aminoglycoside Administration

A retrospective cost analysis compared hospital costs of standard gentamicin dosing and once‐daily regimens in 1127 patients. Hospital costs compared were drug/supply/preparation/administration (DSPA;