Measurement of serum D-arabinitol/creatinine ratios for initial diagnosis and for predicting outcome in an unselected, population-based sample of patients with Candida fungemia.

D-Arabinitol (DA) is a useful diagnostic marker for candidiasis in patients with neutropenia and other high-risk groups, but its use in unselected patients with a broad range of underlying diseases and conditions has not been studied. We used an automated enzymatic fluorometric assay to measure serum DA/creatinine ratios (DA/cr's) in 30 healthy adults, 100 hospitalized controls without Candida fungemia, and 83 patients from a study of all Candida fungemias in Connecticut between October 1998 and September 1999. Sixty-three of 83 (76%) fungemic patients and 11 of 100 (11%) nonfungemic controls had serum DA/cr's >or=3.9 microM/mg/dl (mean + 3 standard deviations for 30 healthy adults). High serum DA/cr's were less frequent in patients with cancer or fungemia caused by the DA nonproducer Candida glabrata than in patients with cancer or fungemia caused by a DA producer, C. albicans, C. tropicalis, or C. parapsilosis. The serum DA/cr was first >or=3.9 microM/mg/dl before, on the same day as, or after the first positive blood culture was drawn for 30 (36%), 22 (27%), and 11 (13%) fungemia patients, respectively. Mortality did not differ significantly among the patients with high or normal initial or peak serum DA/cr's, but mortality was higher if any serum DA/cr value was >or=3.9 microM/mg/dl 3 or more days after the onset of fungemia (18/27 versus 4/24 patients, respectively; P < 0.001). We conclude that serum DA/cr's are useful both for the initial diagnosis of Candida fungemia and for prognostic purposes for unselected patients with a broad range of underlying diseases and conditions.

Development and application of a rapid diagnostic method for invasive Candidiasis by the detection of D-/L-arabinitol using gas chromatography/mass spectrometry.

A rapid non-culture-based diagnostic method utilizing d-/l-arabinitol (DA/LA) ratios as a chemical marker of invasive candidiasis was developed and explored. The enantiomers-ratios detection was made possible by the use of gas chromatography coupled with mass spectrometry (GC/MS). The mean DA/LA ratios +/- standard deviation (range) in urine (n = 40) and serum (n = 20) were 2.08 +/- 0.78 (0.57 to 3.55) and 1.79 +/- 0.75 (0.74 to 3.54), respectively, from patients without evidence of fungal infection or colonization; in patients (n = 7) with culture-proven invasive candida infections, the figures were 9.91 +/- 3.04 (7.24 to 16.27) and 13.58 +/- 7.31 (5.57 to 25.88) in urine and serum, respectively. The differences in DA/LA ratios between the candidemic patients and the non-candidemic patients were statistically significant (p < 0.01) in both serum and urine samples. The DA/LA ratios were not significantly affected in patients with oral or vaginal candidiasis and candiduria.

[Gas chromatography in express diagnosis of candidiasis and monitoring of antifungal therapy efficacy by D-arabinitol and mannose levels in pediatric patients]. / Gazokhromatograficheskaia ékspress diagnostika kandidoza i monitoring éffektivnosti protivogribkovoi terapii po urovniu D-arabinitola i mannozy u detei.

Seventy three children (40 blood and 43 liquor specimens) were examined with the use of gas chromatography (GC) to detect background concentrations of Candida metabolites. The criterium of the children enrollment to the control group was the absence of the clinical and laboratory signs of the fungal infection. The normal contents of the fungus metabolites were considered to be 0.51 +/- 0.28 microgram/ml for D-arabinitol and 17.7 +/- 10.4 micrograms/ml for mannose in the serum and 7.24 +/- 3.04 micrograms/ml for D-arabinitol and 67.1 +/- 47.4 micrograms/ml for mannose in the liquor. Fifty four children at the age of 1 month to 12 years with the signs of the fungal infection requiring systemic antifungal therapy were also examined. Prior to the use of antifungal drugs the routine microbiological tests and GC detection of the fungus metabolites were performed. The fungus was isolated with the cultural method from the blood in 2 patients (6.3 per cent), from the mucosa in 25 (71.4 per cent) out of 32 patients with fungal complications at the background of cytostatic therapy and neutropenia, from the liquor in 3 (21.4 per cent) out of 14 patients with meningitis and from the urine in 8 (100 per cent) out of 8 patients with urinary infection. The GC examination revealed increased levels of the Candida metabolites in 96 per cent of the children. A favourable time course of the infection at the background of amphotericin B or fluconasol use was recorded by the clinical indices which correlated with a reliable decrease of the contents of D-arabinitol and mannose to the normal. The use of GC is recommended in express diagnosis of candidiasis especially when the results of the cultural tests are negative as well as in monitoring of the fungal therapy efficacy.

Diagnosis and therapeutic monitoring of invasive candidiasis by rapid enzymatic detection of serum D-arabinitol.

BACKGROUND: Using a rapid automated enzymatic assay, we prospectively investigated serum D-arabinitol (DA), a biochemical marker of invasive candidiasis, in a large population of high-risk patients to determine its potential diagnostic, therapeutic, and prognostic significance in invasive candidiasis. PATIENTS AND METHODS: A total of 3,223 serum samples were collected from 274 patients with cancer. Serum DA concentrations were determined in coded serum samples analyzed by rapid enzymatic assay. Creatinine also was analyzed in the same system to determine a serum DA and creatinine ratio (DA/Cr). The sensitivity, specificity, correlation with therapeutic response, and prognostic significance were analyzed for all patient study groups. RESULTS: A DA/Cr of > or = 4.0 mumol/L per mg/dL was detected in 31 (74%) of all 42 cases of fungemia and 25 (83%) of the 30 cases of the subset of persistent fungemia. Elevated DA/Cr was detected in 4 (40%) of 10 patients with tissue-proven, deeply invasive candidiasis and negative blood cultures (eg, hepatosplenic candidiasis or localized abscess) and 7 (44%) of 16 cases of deep mucosal candidiasis (eg, esophageal candidiasis). Elevated serial DA/Cr levels also were detected in persistently febrile and granulocytopenic patients requiring empirical amphotericin B. Among 26 assessable cases of fungemia, abnormally elevated DA/Cr values were detected in 14 (54%) before, 10 (38%) after, and 2 (8%) simultaneously with the first microbiologic report of fungemia. The trends of serial DA/Cr values correlated with therapeutic response in 29 (85%) of 34 patients with assessable cases of fungemia, decreasing in 8 (89%) of 9 patients with clearance of fungemia and increasing in 21 (84%) of 25 patients with persistence of fungemia. Among the 34 assessable patients with fungemia, mortality was directly related to the trend of serial DA/Cr determinations over time: 71% among fungemic patients who had persistently elevated or increasing DA/Cr, and 18% among the fungemic patients who had resolving DA/Cr or never had elevated DA/Cr (P < 0.01). CONCLUSIONS: Rapid enzymatic detection of DA in serially collected serum samples from high-risk cancer patients permitted detection of invasive candidiasis, early recognition of fungemia, and therapeutic monitoring in DA-positive cases. Serially collected serum DA determinations complement blood cultures for improving detection and monitoring therapeutic response in patients at risk for invasive candidiasis.

Capillary gas chromatography combined with ion trap detection for quantitative profiling of polyols in cerebrospinal fluid and plasma.

Polyol species in cerebrospinal fluid and plasma--ribitol, arabitol, xylitol, 1,5-anhydrosorbitol, myo-inositol, mannitol, sorbitol, and galactitol--simultaneously were quantitated by a capillary gas chromatography/ion trap (mass spectrometric) detection method. The details of the methodology are discussed and the results of analysis of polyols in healthy human subjects are reported. Microliter volumes of cerebrospinal fluid or plasma were mixed with internal standard (deuterium labeled myo-inositol), deproteinized, and evaporated to dryness. Polyols were acetylated in the presence of pyridine catalyst and washed with sodium bicarbonate solution and the acetate derivatives were recovered. Standard curve solutions were similarly treated. The polyol components were resolved on a capillary column bonded with 50% phenyl-50% methyl polysiloxane. Chemical ionization mass spectra for the acetate derivatives of polyols were generated in an ion trap using acetonitrile as reagent gas. Each polyol yielded a fragment ion in 100% abundance arising probably from the loss of one acetate moiety from the protonated molecule. These ions were monitored. The relative standard deviation (within-day) for quantitation of polyols was not greater than 8% for cerebrospinal fluid and 15% for plasma matrix. A polyol profile in cerebrospinal fluid and plasma was determined in healthy human subjects and a cerebrospinal fluid/plasma concentration ratio larger than 1.0 was found for all polyol species except 1,5-anhydrosorbitol and xylitol. This assay technique will be used to study the role of polyols in central nervous system diseases.

[A comparative clinical study on flucytosine alone and in combination with fluconazole in hematological malignancies: a multicenter study using the envelope method].

Two different flucytosine (5-FC) treatment regimens, one by itself and the other in combination with fluconazole (FLCZ) were compared in chemotherapy against mycotic infections in 60 patients with hematological diseases. The patients in a randomized fashion were assigned to the two treatments. In the combination regimen, the two drugs were used in half doses. beta-D-Glucan and D-arabinitol in the sera of patients were measured to document mycotic infections, and bacterial examinations were also performed. The efficacy of the combination therapy was 60.0% (18/30) and that of 5-FC alone was 65.5% (19/29). The stratified evaluation indicated that no factor was found to contribute to the efficacy in the two treatments with statistical significance. The side effects occurred in few cases and none of those was serious; including, one case of subjective symptom in each groups and two episodes of liver dysfunction in combination treatment. Changes in beta-D-glucan concentrations in the sera reflected well the pathophysiology of mycotic infections and clinical improvement. These results suggested that a combination of 5-FC and FLCZ at half doses provided a clinical benefit comparable to 5-FC alone at the ordinary dose, and the safety was considered satisfactory.

Trichosporon cutaneum fungemia in patients with acute myeloblastic leukemia and measurement of serum D-arabinitol, Candida antigen (CAND-TEC), and beta-D-glucan.

Two patients with acute myeloblastic leukemia are described who developed fungemia due to Trichosporon cutaneum. Fungemia occurred at the leukocyte nadir following the administration of anti-cancer chemotherapy. One patient was cured but the other died. Both patients received prednisolone continuously and had central venous catheters in place for parenteral hyperalimentation. T. cutaneum isolates were resistant to 5-fluorocytosine and moderately susceptible to fluconazole. One case was complicated by endophthalmitis due to T. cutaneum; this is the second report of such a complication in the world. We investigated the serum levels of beta-D-glucan, D-arabinitol, and Candida antigen (CAND-TEC); beta-D-glucan was elevated in both cases, which suggests that simultaneous measurements of these laboratory values are useful for the diagnosis and possibly for the evaluation of therapy for this fungal infection.

Diagnosis and monitoring of disseminated candidiasis based on serum/urine D/L-arabinitol ratios.

Disseminated candidiasis, a devastating disease with high morbidity and mortality in immunosuppressed patients, is difficult to diagnose because of the protean nature of symptoms and the lack of rapid and reliable laboratory diagnostic procedures. The subject of this review is the status of gas chromatographic-mass spectrometric techniques for the determination of D-arabinitol, a unique metabolite of pathogenic Candida species, in serum and urine. The enantiomers are separated by chiral chromatography followed by specific and sensitive detection using chemical ionization and selected ion monitoring. Using D/L-arabinitol ratios, instead of individual concentrations, eliminates the need for knowing the volume of samples and for calibration curves. A new filter paper technique requires only an unmeasured drop of whole blood (venous or finger/heel puncture) or urine; paper spots are mailable. Parallel determinations of D/L-arabinitol ratios in serum and urine in normal subjects and cancer patients with both normal and increased D/L-arabinitol ratios revealed constant (1.2-1.3 range) ratios of serum D/L-arabinitol/urine D/L-arabinitol for all populations studied. Analyzing two body fluids taken at the same time increases reliability by reducing false positives.

Measurement of serum arabinitol by gas-liquid chromatography: limitations for detection of systemic candida infections.

The measurement of serum arabinitol in the diagnosis of systemic candidosis was evaluated using a gas-liquid chromatography technique in a cohort of at risk patients. The prevalence of seropositivity was low and did not correlate with evidence of infection. This technique is unlikely to achieve acceptance because it does not discriminate between patients with and without infection; it requires specialised equipment and it is expensive.

[Differentiation of intestinal candidial colonization from invasive candidiasis by measuring serum level of D-arabinitol in combination with oral administration of low dose amphotericin B].

The measurement of D-arabinitol in serum has been reported to be useful for the diagnosis of invasive candidiasis. However, excessive proliferation of Candida species in intestinal tract often leads false positive result of serum D-arabinitol. Based on the evidence that amphotericin B (AMPH) is scarcely absorbed from intestinal tract and inhibits the proliferation of Candida species only in intestinal tract, we have developed a simple differentiation method of intestinal candida colonization from invasive candidiasis by measuring serum level of D-arabinitol in combination with oral administration of low dose AMPH. AMPH, 600 mg/day for 2 days was orally administered to five patients with hematological malignancies who showed more than 1.7 mumol/mg of D-arabinitol/creatinine ratio (D/C ratio) in serum without any evidence of invasive candidiasis. D/C ratios were markedly decreased and normalized after the oral administration of low dose AMPH. While, in a patient with invasive candidiasis in whom Candida species was detected by blood cultures, D/C ratio remained unchanged in spite of oral administration of AMPH. These observations suggest that this method is a simple and reliable diagnostic method to distinguish intestinal candida colonization from true invasive candidiasis.

[A case of candida endocarditis with consecutive measurement of serum mannan and D-arabinitol concentrations].

We encountered a case of an 83-year-old patient with candida endocarditis. He had been diagnosed as having aortic regurgitation with moderately calcified aortic valve 5 years previously, and had received medication. He was admitted to our hospital because of pancreatic cancer. He had high fever from the time of his admission and antibiotics produced no effect. Candida albicans was detected in arterial blood culture. We also detected antibody against Candida albicans, and investigated serum mannan and the D-arabinitol creatinine ratio several times. We performed echo-cardiographic examination and recorded a vegetation at the aortic valve. Rising antibody titers against Candida albicans, mannan antigenemia and an elevated creatinine ratio were also observed. So we concluded that these examinations were also effective in the diagnosis of candida infection. Finally, this patient died of cerebral hemorrhage and was autopsied. Macroscopic findings showed mass-like vegetation involving the aortic valve, and microscopic findings showed candida organisms scattered within the vegetation.

Separation and quantification by gas chromatography-mass spectrometry of arabinitol enantiomers to aid the differential diagnosis of disseminated candidiasis.

To differentiate increased arabinitol due to fungal (only D-arabinitol) and non-fungal origin, O-trifluoroacetyl derivatives of the enantiomers were separated using alpha-perpentylated cyclodextrin columns and measured by selected ion monitoring. Mean +/- S.D. D/L in normal serum: 1.40 +/- 0.42. D/L ratios greater than 2.24, defined as normal mean + 2S.D., were considered outside normal range. D/L was greater than 2.2 in 10 of 12 confirmed candidiasis cases with one false negative and one borderline. Renal dysfunction without candidiasis yielded normal D/L despite high arabinitol concentrations. D/L in normal urine was nearly identical to that in serum despite 60 times larger concentration. D/L ratios, determined by peak heights or areas, could be used without the need to determine concentrations.

[The determination of the D-arabinitol content of the blood serum--a method for the diagnosis and monitoring of an invasive candidal infection in cytopenia patients (preliminary data)]. / Opredelenie soderzhaniia D-arabinitola v syvorotke krovi--metod diagnostiki i monitoringa invazivnoi kandidoznoi infektsii u bol'nykh s tsitopeniiami (predvaritel'nye dannye).

Gas chromatography was used to identify D-arabinitol over time (5-6 times on the average) in the blood serum of 24 patients with fever of unclear etiology. The patients were in a state of cytostatic and/or cytostatic and radiation cytopenia. In 20 donors, D-arabinitol was identified once. In the majority of the donors, that indicator did not exceed 1 microgram/ml, amounting on the average to 0.7 +/- 0.3. In patients with invasive candidiasis, the concentration of D-arabinitol surpassed 1 microgram/ml, reaching 4.0-5.5 micrograms/ml in some patients. In patients colonized with Candida, that indicator also exceeded 1 microgram/ml, not reaching, however, such high values as those seen in invasive candidiasis. Identification of the D-arabinitol level reflects the total activity of the fungi of the Candida genus contaminating the patients' mucous membranes at the moment of investigation irrespective of the process site, whereas monitoring allows one to follow tendencies of that activity and to correct therapy.

Enantioselective measurement of the Candida metabolite D-arabinitol in human serum using multi-dimensional gas chromatography and a new chiral phase.

A multi-dimensional gas chromatographic method was developed to measure the Candida metabolite D-arabinitol enantioselectively in human serum. The heptafluorobutyrate derivatives of D-arabinitol, L-arabinitol and ribitol (internal standard) were separated from other serum constituents with a 60 m X 0.32 mm fused-silica SPB-5 precolumn, and (after intermediate cold trapping) they were separated from each other with a 25 m x 0.25 mm fused-silica column coated with a new bonded chiral phase. Replicate analyses of spiked human sera showed that D-arabinitol could be quantified accurately and precisely. The D- and L-arabinitol concentrations in 24 normal adult sera were 0.20 +/- 0.053 and 0.11 +/- 0.040 mu/ml, respectively, and the D- and L-arabinitol/creatinine ratios were 0.023 +/- 0.011 and 0.012 +/- 0.0051, respectively (mean +/- S.D.). In a patient with Candida albicans fungemia, the D-arabinitol/creatinine ratios rose early during infection and fell with successful treatment, whereas L-arabinitol/creatinine ratios did not change significantly. This enantioselective analytical method is more practical than earlier ones; it should facilitate further investigation of D-arabinitol as a diagnostic marker for candidiasis.

Mannan and D-arabinitol concentrations in serum from a patient with Candida albicans endocarditis.

In an attempt to clarify the comparative values of serological and microbiological examinations for the early diagnosis of systemic candidiasis, antibodies against Candida albicans, serum mannan, and the D-arabinitol creatinine ratio were investigated in a patient with aortic valve endocarditis associated with carcinoma of the bile duct. Candida precipitins and the antibody titer against Candida cell wall mannan were examined by an immunodiffusion technique and hemagglutination test, respectively. Serum mannan was tested by enzyme-linked immunosorbent assay (ELISA) using the biotin-streptavidin procedure. The upper limit of negativity of the assay was determined by adding 0.06 to the absorbance of pooled serum from healthy laboratory workers. This value was about 0.8 ng/ml with ELISA. The D-arabinitol concentration in serum was examined by an enzymatic fluorometric method. Rising antibody titers against C. albicans, mannan antigenemia, and an elevated D-arabinitol creatinine ratio were first observed between the 11th and 12th hospital days. Blood cultures obtained on 8th, 9th, and 11th hospital days grew C. albicans after 3 to 4 days of incubation. Of 11 serum samples, 5 were positive for mannan, whereas D-arabinitol creatinine ratio was positive in 7 of 9 samples. Blood cultures was the earliest evidence of Candida infections in our cases. However, because of saprophytic nature of Candida species, tests for antibodies, antigenemia, and the D-arabinitol creatinine ratio in combination with blood cultures are necessary to confirm systemic candidiasis at an early stage of infection.

[Treatment of Candida urinary tract infections with reference to serum levels of anti-Candida antibody and D-arabinitol].

A clinical study was carried out on 11 patients with Candida urinary tract infection; 4 with chronic cystitis and 7 with acute pyelonephritis including 2 patients associated with candidemia. The patients received 1.5 to 6.0 g of 5-fluorocytosine (5-FC) orally daily for 6 to 22 days. The response was excellent in 6, moderate in 4 and poor in 1 case, overall efficacy rate being 90.9%. No patients showed any adverse reaction. The antibody against the Candida species was measured by an indirect hemagglutination test and the serum level of D-arabinitol, a major metabolite of Candida species, was determined by gas-liquid chromatography. A 4-fold or greater rise in anti-Candida antibody was detected in 4 of the 6 patients with pyelonephritis, including one patient associated with candidemia, 14 to 31 days following the treatment. Serum D-arabinitol levels in all of the 4 patients with pyelonephritis exceeded the detection limit of 0.2 microgram/ml before treatment. The levels decreased promptly during treatment and normalized 10 to 20 days after the start of treatment. These results suggest that 5-FC is an effective drug for Candida urinary tract infections. The determinations of anti-Candida antibody titer and serum D-arabinitol level may be clinically useful for the diagnosis of Candida pyelonephritis. In particular, the serum levels of D-arabinitol appear to be a useful parameter to follow-up the effect of anti-fungal treatment on Candida pyelonephritis.