Clinical utility of a panfungal polymerase chain reaction assay for invasive fungal diseases in patients with haematologic disorders.

OBJECTIVES: Invasive fungal diseases (IFDs) are life-threatening events in patients with haematologic disorders, and the spectrum of the aetiological pathogens continues to expand. This study aimed to evaluate the clinical utility of a panfungal polymerase chain reaction (PCR) assay for the management of IFDs in such patients. METHODS: We prospectively analysed 273 consecutive blood samples from 64 risk episodes in 51 patients with haematologic disorders at high risk for IFD who were treated at our hospital between April 2007 and October 2010. RESULTS: PCR-positive results were obtained in 18 of 64 risk episodes (35.3%). IFD was documented in 14 episodes (21.9%, 9 probable IFDs and 5 possible IFDs) according to the revised criteria of the European Organization for Research and Treatment of Cancer/Mycoses Study Group. PCR was positive in all of these 14 episodes, and in 4 of the 50 episodes with no IFD category. Sensitivity, specificity, positive predictive value, and negative predictive value of our assay were 100%, 92%, 78% and 100% respectively. A considerable number of fungi (44.4%) that are less common than Aspergillus and Candida species were positive by PCR. Molecular diagnoses of Cunninghamella species, Aspergillus ustus, Fusarium species, Scedosporium apiospermum, Rhodotorula species and Rhizopus species were beneficial in selecting suitable treatments. CONCLUSIONS: Our panfungal PCR approach allows for the highly sensitive and specific detection and identification of a wide spectrum of fungal pathogens, which provides indispensable information for managing IFDs, especially refractory or breakthrough IFDs during antifungal therapy in high-risk patients with haematologic disorders.

Diurnal differences in urine flow in healthy young men in a light-controlled environment: a randomized crossover design.

BACKGROUND: Older men often experience nocturnal urination difficulties, reflected by diurnal differences in maximum urine flow (Qmax). Since lower urinary tract symptoms and pathological comorbidities are frequent in older men, it remains unclear whether this diurnal variation is a physiological or pathological phenomenon. Our aim was to quantify the diurnal variability of Qmax in healthy young participants under varying daylight conditions in a stable environment to discern potential underlying causes of nocturnal urination difficulties. METHODS: Twenty-one healthy young men were recruited in a 4-day study utilizing daytime (08:00-18:00) exposure with two light conditions in randomized order: dim (< 50 lx) or bright (~2500 lx). Day 1 was for acclimation, and urine flow was assessed from day 2. The participants urinated ad libitum during day 2 and then at fixed 3-4-h intervals thereafter (days 3-4). Regular urination Qmax at late night (04:00) on day 4 was compared with the nearest voided volume during daytime of day 3 (mDay). RESULTS: Morning Qmax scores (after bed-11:00) on day 2 were significantly lower than evening (17:00-before pre-sleep) in bright conditions and those of daytime (11:00-17:00), evening (17:00-before pre-sleep), and pre-sleep in dim conditions. Pre-sleep Qmax during the ad libitum period was significantly higher in dim than bright conditions. Late-night Qmax values (04:00) on day 4 were significantly lower than Qmax scores of mDay on day 3 in both light conditions. CONCLUSIONS: Healthy young men had a clear diurnal Qmax difference that decreased during late night and morning. In addition, the pre-sleep Qmax values in dim daylight were significantly higher than in bright daylight. Taken together, we conclude that late-night and morning decreases in Qmax are an instinctive physiological phenomenon in humans, and the diurnal difference of Qmax can be influenced by daylight conditions.

Diurnal rhythms of urine volume and electrolyte excretion in healthy young men under differing intensities of daytime light exposure.

In humans, most renal functions, including urine volume and electrolyte excretions, have a circadian rhythm. Light is a strong circadian entrainment factor and daytime-light exposure is known to affect the circadian rhythm of rectal temperature (RT). The effects of daytime-light exposure on the diurnal rhythm of urinary excretion have yet to be clarified. The aim of this study was to clarify whether and how daytime exposure to bright-light affects urinary excretions. Twenty-one healthy men (21-27 years old) participated in a 4-day study involving daytime (08:00-18:00 h) exposure to two light conditions, Dim (< 50 lx) and Bright (~ 2500 lx), in a random order. During the experiment, RT was measured continuously. Urine samples were collected every 3 ~ 4 h. Compared to the Dim condition, under the Bright condition, the RT nadir time was 45 min earlier (p = 0.017) and sodium (Na), chloride (Cl), and uric acid (UA) excretion and urine volumes were greater (all p < 0.001), from 11:00 h to 13:00 h without a difference in total daily urine volume. The present results suggest that daytime bright light exposure can induce a phase shift advance in urine volume and urinary Na, Cl, and UA excretion rhythms.

Diagnostic value of PCR analysis of bacteria and fungi from blood in empiric-therapy-resistant febrile neutropenia.

This study aimed to assess the clinical utility of PCR for the analysis of bacteria and fungi from blood for the management of febrile neutropenic patients with hematologic malignancies. Using a PCR system able to detect a broad range of bacteria and fungi, we conducted a prospective pilot study of periodic analyses of blood from patients following intensive chemotherapy. When fever occurred, it was treated with empirical antibiotic therapy, basically without knowledge of the PCR results. In 23 febrile episodes during the neutropenic period, bacteria were detected by PCR in 11 cases, while the same species were identified by blood culture in 3 cases. In 10 out of 11 PCR-positive cases, fever could be managed by empirical therapy. In the empirical-therapy-resistant case, the identification of Stenotrophomonas maltophilia by PCR led to improvement of fever. No fungi were detected by PCR in febrile cases, while Aspergillus fumigatus was detected in one afebrile patient, several days before a clinical diagnosis was made. In subsequent sporadic PCR analyses in 15 cases of febrile neutropenia, bacteria were detected by both PCR and blood culture in 7 cases and by PCR alone in 6. Fungi were not detected. While fever was improved by empirical therapy in 12 out of the 13 PCR-positive cases, the identification of Pseudomonas aeruginosa by PCR in one therapy-resistant case contributed to the successful treatment of persistent fever. Our results indicate that PCR analysis of bacteria from blood provides essential information for managing empirical-therapy-resistant febrile neutropenia.

Metallo-beta-lactamase IMP-1 in Providencia rettgeri from two different hospitals in Japan.

In 2002, 495 indole-positive proteae strains were isolated from patients at 60 hospitals in Japan. Nine indole-positive proteae strains had reduced susceptibility to imipenem (MIC > or = 8 microg ml(-1)) and were identified as Providencia rettgeri by BD Phoenix. Eight of the nine Prov. rettgeri isolates were confirmed as metallo-beta-lactamase producers by the double-disc synergy test. All the metallo-beta-lactamases were classified as IMP-1 by PCR and DNA sequence analysis. These bla(IMP-1) genes were encoded in the integron structure on conjugative plasmids. These plasmids could transfer from Prov. rettgeri clinical isolates to Escherichia coli ML4903 at a frequency between 1.5 x 10(-5) and 5.5 x 10(-7). The eight bla(IMP)-positive strains were isolated from two hospitals, and showed two different PFGE patterns, two different integron structures and two different incompatibility groups, which corresponded to the two hospitals. These results strongly suggest the possibility of nosocomial infections by bla(IMP-1)-producing Prov. rettgeri isolates.

[Early and differential diagnosis for fungal infection by PCR in case of living donor liver transplantation].

Although opportunistic infection including fungal infection is often associated with living donor liver transplantation followed by immunosuppressive therapy, antifungal agents are empirically given to most patients without a definitive diagnosis of fungal infection. Indeed, there is no diagnostic test available, that shows both high sensitivity and specificity for fungal infection. In this study, we developed a polymerase chain reaction(PCR)-based system for rapid diagnosis of fungal infection. This system consisted of two PCR steps: the first PCR for most species of fungi and 4 kinds of nested PCR for Aspergillus or Penicillium (ASP/PEN), Candida glabrata (C. glab), other Candida species including Candida albicans, Candida parapsilosis, Candida tropicalis, and Candida guiliermondii (CAN), and a broad spectrum of fungi (Broad). The newly developed PCR-based system was applied to 28 recipients of with living donor liver transplantation to determine its clinical usefulness in early and differential diagnosis of fungal infection. A total of 514 blood samples from 28 patients ware analyzed. Nested PCR assays were positive in 118 samples from 19 patients: 4 patients (30 samples) with ASP/PEN, 5 patients (29 samples) with C. glab, 12 patients (61 samples) with CAN. All of these samples were positive with nested PCR for Broad as well. Even in samples that were negative on blood culture or ELISA for Aspergillus antigen, nested PCR assays were able to amplify the fungal DNA. Furthermore, all samples positive for Aspergillus antigen tested positive on nested PCR. In conclusion, the PCR-based system we developed was thought to be useful for early diagnosis of fungal infection.

[The prevalence of beta-lactamase negative ampicillin resistant Haemophilus influenzae in Mie Prefecture].

Equivalent MIC breakpoints to detect beta-lactamase negative ampicillin resistant Haemophilus influenzae (BLNAR) were controversial. We studied the relationship of drug resistance with gene alterations in 74 clinical isolates of H. influenzae. Out of 74 isolates, 26 showed MIC of ampicillin (ABPC) > or = 1 microg/ml. All isolates, except one, with MIC of ABPC > or = 4 microg/ml were found to produce beta-lactamase, while all 19 isolates with MIC of ABPC at 1 or 2 microg/ml were non-producing. Twenty-six ABPC resistant isolates were subjected to the analysis of genes involved in the drug resistance such as pbp3-1 pbp3-2, and TEM by the Haemophilus influenzae gene detection kit (Wakunaga Pharmaceutical Co., Ltd.) according to the supplier's instructions. Three (21.4%) of 14 beta-lactamase non-producing isolates with ABPC-MIC of 1 microg/ml had mutations of pbp3-1 gene, while all 5 non-producing isolates with ABPC-MIC of 2 microg/ml showed mutations of both pbp3-1 and pbp3-2 genes. Accordingly, it seems appropriate to set ABPC-MIC > or = 2 microg/ml for detection of BLNAR. In this study, six (8.1%) of 74 isolates were found to be BLNAR, and all of these six isolates were derived from patients of 5 year-old or younger.

Basal metabolic rate and body composition of elite Japanese male athletes.

The estimated energy requirement is important for adequate nutritional management in athletes. The energy requirement can be estimated from the basal metabolic rate (BMR). However, there is little data regarding the BMR of Japanese athletes. This study measured the BMR and body composition of 81 elite Japanese male athletes in different sports categories: endurance (E), strength, power and sprint (S) and ball game (B). The factors influencing the BMR were also investigated. The BMR and body composition were measured by indirect calorimetry and an air-displacement plentysmograph device (the BOD POD), respectively. The BMR per lean body mass (LBM) differed significantly among the three groups. The BMR was significantly correlated with the body weight (BW) and LBM in all groups. A multiple-regression analysis showed that the LBM was the most powerful predictor in the E and S groups, whereas the BW was the most powerful predictor in the B group. The BW appears to become an important predictor as the BW of athletes increases. Additionally, height was the second explanatory variable in the S and B groups, thus suggesting that height needs to be considered for the BMR in these groups. Therefore, the BMR in elite athletes needs to be estimated according to their body composition.

Efficacy of procalcitonin in the early diagnosis of bacterial infections in a critical care unit.

Procalcitonin (PCT) is a marker of severe bacterial infections and organ failure due to sepsis. The purpose of the present study was to identify the appropriate cutoff level of PCT based on the findings of a blood culture and polymerase chain reaction (PCR). The PCT levels were measured in 116 patients in an intensive care unit who were suspected of having bacteremia, to examine its relationship with a blood culture or PCR. The PCT levels were significantly high in patients with bacteremia, but they were also moderately high in some patients who were positive for fungus DNA. The area under the curve was significantly higher for PCT than for C-reactive protein. The appropriate cutoff values of PCT for bacteremia were 0.38 microg/L for the high negative predictive value and 0.83 microg/L for the high positive predictive value. Procalcitonin was slightly related to mortality, and the combination of a blood culture and PCR was thus found to increase the sensitivity for mortality. These findings suggest that PCT is useful for the diagnosis of bacteremia and that the diagnostic value of PCT in combination a with blood culture and PCR for bacterial infection or mortality further increases.