Montserrat Giménez

Epidemiology and Predictors of Mortality in Cases of Candida Bloodstream Infection: Results from Population-Based Surveillance, Barcelona, Spain, from 2002 to 2003

ABSTRACT We conducted population-based surveillance for Candida bloodstream infections in Spain to determine its incidence, the extent of antifungal resistance, and risk factors for mortality. A case was defined as the first positive blood culture for any Candida spp. in a resident of Barcelona, from 1 January 2002 to 31 December 2003. We defined early mortality as occurring between days 3 to 7 after candidemia and late mortality as occurring between days 8 to 30. We detected 345 cases of candidemia, for an average annual incidence of 4.3 cases/100,000 population, 0.53 cases/1,000 hospital discharges, and 0.73 cases/10,000 patient-days. Outpatients comprised 11% of the cases, and 89% had a central venous catheter (CVC) at diagnosis. Overall mortality was 44%. Candida albicans was the most frequent species (51% of cases), followed by Candida parapsilosis (23%), Candida tropicalis (10%), Candida glabrata (8%), Candida krusei (4%), and other species (3%). Twenty-four isolates (7%) had decreased susceptibility to fluconazole (MIC ≥ 16 μg/ml). On multivariable analysis, early death was independently associated with hematological malignancy (odds ratio [OR], 3.5; 95% confidence interval [CI], 1.1 to 10.4). Treatment with antifungals (OR, 0.05; 95% CI, 0.01 to 0.2) and removal of CVCs (OR, 0.3; 95% CI, 0.1 to 0.9) were protective factors for early death. Receiving adequate treatment, defined as having CVCs removed and administration of an antifungal medication (OR, 0.2; 95% CI, 0.08 to 0.8), was associated with lower odds of late mortality; intubation (OR, 7.5; 95% CI, 2.6 to 21.1) was associated with higher odds. The incidence of candidemia and prevalence of fluconazole resistance are similar to other European countries, indicating that routine antifungal susceptibility testing is not warranted. Antifungal medication and catheter removal are critical in preventing mortality.

Procalcitonin, C-reactive protein and leukocyte count in children with lower respiratory tract infection

Background. Lower respiratory tract infection is the most common infection leading to unnecessary antibiotic treatment in children. Etiologic diagnosis is not immediately achieved, and the pathogen remains unidentified in a large number of cases. Neither clinical nor laboratory factors allow for a rapid distinction between bacterial and viral etiology. The aim of our study was to evaluate the reliability of procalcitonin (PCT), C-reactive protein (CRP) and leukocyte count in distinguishing pneumococcal, atypical and viral lower respiratory tract infection. Methods. PCT, CRP and leukocyte count were measured in children with microbiologically documented diagnoses of lower respiratory tract infection. The results were compared of children with pneumococcal, atypical and viral etiologies. Results. PCT and CRP showed significant correlation with a bacterial etiology of lower respiratory tract infection. No significance was found for leukocyte count. Using a cutoff point of 2 ng/ml for PCT and 65 mg/l for CRP, the sensitivities and specificities for distinguishing bacterial from viral lower respiratory tract infections were 68.6 and 79.4% for PCT and 79.1 and 67.1% for CRP. The sensitivities and specificities for distinguishing pneumococcal from other etiologies were 90.3 and 74.1% for PCT and 90.3 and 60% for CRP, respectively. Conclusions. High PCT and CRP values show a significant correlation with the bacterial etiology of lower respiratory tract infection. PCT and CRP show good sensitivity for distinguishing pneumococcal from other etiologies. PCT shows higher specificity than CRP. PCT and CRP can help make decisions about antibiotic therapy in children with lower respiratory tract infections.

Rapid diagnosis of bloodstream infections with PCR followed by mass spectrometry.

Achieving a rapid microbiological diagnosis is crucial for decreasing morbidity and mortality of patients with a bloodstream infection, as it leads to the administration of an appropriate empiric antimicrobial therapy. Molecular methods may offer a rapid alternative to conventional microbiological diagnosis involving blood culture. In this study, the performance of a new technology that uses broad-spectrum PCR coupled with mass spectrometry (PCR/ESI-MS) was evaluated for the detection of microorganisms directly from whole blood. A total of 247 whole blood samples and paired blood cultures were prospectively obtained from 175 patients with a suspicion of sepsis. Both sample types were analyzed using the PCR/ESI-MS technology, and the results were compared with those obtained by conventional identification methods. The overall agreement between conventional methods and PCR/ESI-MS performed in blood culture aliquots was 94.2% with 96.8% sensitivity and 98.5% specificity for the molecular method. When comparing conventional methods with PCR/ESI-MS performed in whole blood specimens, the overall agreement was 77.1% with 50% sensitivity and 93.8% specificity for the molecular method. Interestingly, the PCR/ESI-MS technology led to the additional identification of 13 pathogens that were not found by conventional methods. Using the PCR/ESI-MS technology the microbiological diagnosis of bloodstream infections could be anticipated in about half of the patients in our setting, including a small but significant proportion of patients newly diagnosed. Thus, this promising technology could be very useful for the rapid diagnosis of sepsis in combination with traditional methods.

Use of Quantitative and Semiquantitative Procalcitonin Measurements to Identify Children with Sepsis and Meningitis

During infancy and childhood, clinical signs of infection and conventional laboratory markers are not specific in the early phase of disease [1]. The availability of a parameter that more rapidly identifies children suspected to have bacterial sepsis before microbiological results are available would minimize unnecessary treatments and hospitalization. Since its original description, the importance of procalcitonin (PCT) as an indicator of systemic bacterial infection has been demonstrated in many reports [2, 3, 4, 5]. The aim of our study was to evaluate the reliability of PCT measurement by quantitative luminometric immunoassay (LIA) in distinguishing between systemic bacterial infection (sepsis and/or meningitis), localized bacterial infection and aseptic meningitis in children, compared to leukocyte count and C-reactive protein (CRP) levels. We also evaluated the correlation of a rapid immunochromatographic test (ICT) for semiquantitative PCT measurement in comparison with quantitative test results. The study was carried out on selected children aged between 1 month and 12 years who were admitted to the pediatric emergency department of our hospital after presenting with fever of less than 12-h duration. At the time of admission, blood samples were collected for leukocyte count, CRP and PCT measurement. Clinical specimens were collected for microbiological testing in order to correctly establish the etiology. Serial serum samples for PCT and CRP assays were collected daily when possible. Patients were grouped retrospectively according to the type of illness. Group 1 included 25 children diagnosed with bacterial sepsis and/or meningitis by culture of blood and/or cerebrospinal fluid (CSF) samples: Neisseria meningitidis was isolated in 18 cases, Streptococcus pneumoniae in 6 and Haemophilus influenzae in 1. Bacteria were isolated from CSF culture only in 8 patients, blood culture only in 10 and both culture types in 7. Group 2 included 18 children diagnosed with aseptic meningitis by compatible clinical presentation, negative Gram stain, cultures and antigen tests for bacterial infection using blood and CSF samples, compatible CSF analysis (pleocytosis with a predominance of mononuclear cells) and successful recovery without antibiotic therapy. Viral cultures were not performed. Group 3 included 22 children presenting with localized bacterial infection such as purulent conjunctivitis, acute otitis media, streptococcal pharyngitis, cellulitis or sinusitis, confirmed by culture of the site of infection and negative blood cultures. The bacteria isolated were Staphylococcus aureus, Streptococcus pneumoniae and Streptococcus pyogenes. In order to establish normal PCT levels for comparison, serum samples were also collected from a control group of 25 similarly aged healthy children admitted to the hospital for elective surgery. PCT was measured using LIA (Lumitest PCT; Brahms Diagnostica, Germany) for quantitative detection and ICT (PCT-Q; Brahms Diagnostica) for semiquantitative detection following the manufacturer’s instructions. LIA requires 20 l of serum and 90–120 min to be performed. ICT requires 200 l of serum and results can be obtained in 30 min; samples can be measured individually. CRP was measured using a turbidimetric assay (C-Reactive Protein FlexT reagent cartridge, Dimension; Dade Behring, USA) and leukocyte count was measured with Coulter Counter Maxm (Coulter, USA). Comparison between groups for quantitative parameters was performed using the non-parametric MannWhitney U test for CRP, leukocyte count and PCT. Diagnostic accuracy and optimum cut-off points were determined using a receiver operating characteristic curve. Comparisons between the two methods were made C. Prat ()) · J. Dom nguez · M. Gim nez · S. Blanco · V. Ausina Servei de Microbiologia, Hospital Universitari Germans Trias i Pujol, C/Canyet s/n, 08916 Badalona, Spain e-mail: crisprat@ns.hugtip.scs.es Tel.: +34-93-4978894 Fax: +34-93-4978895

Noninvasive Method for Diagnosis of Visceral Leishmaniasis by a Latex Agglutination Test for Detection of Antigens in Urine Samples

Since the appearance of human immunodeficiency virus (HIV), visceral leishmaniasis (VL) has emerged as an opportunistic infection in developed countries (1, 3, 10). It appears in advanced stages of HIV infection and is supposed to accelerate the progression of AIDS (3, 5, 8). Therapeutic failures and relapses are common. Accurate diagnosis is usually difficult in HIV-coinfected patients because VL has atypical clinical expressions and because serological diagnosis becomes unreliable (1). Demonstration of the parasite in cultured samples or in stained preparations is considered the “gold standard” (GS) for diagnosis but requires invasive techniques. A noninvasive and accurate test is needed in order to improve diagnosis of VL, especially in individuals with an increased risk of suffering complications after invasive methods (2, 6). We have evaluated the effectiveness of a rapid latex agglutination test (LAT) (KATEX; Kalon Biological Ltd., Aldershot, Hants, United Kingdom) in the detection of a leishmanial antigen (9) in urine from patients with VL and its usefulness in treatment monitoring. A bone marrow aspirate and a fresh urine specimen were collected from 85 patients with suggestive clinical symptoms and signs of VL. The demonstration of the parasite by culture or Giemsa stain in bone marrow aspirate was considered the GS. In 16 out of 89 cases the VL clinical diagnosis was confirmed by the GS (group 1), while in 73 it was not (group 2). Urine samples from 73 patients without any suspicion of VL were collected as negative controls (group 3). The percentages of HIV-infected individuals were as follows: 100% in group 1, 91% in group 2, and 21% in group 3. The LAT was performed in previously boiled urine specimens according to the manufacturers instructions. A clear agglutination was recorded as positive. All specimens from group 1, three from group 2, and none from group 3 were positive by the LAT (Table ​(Table1).1). One of the three patients from group 2 who tested positive had been diagnosed with VL 1 year before. The sensitivity was 100%, and specificity was 96% (Table ​(Table11). TABLE 1. Leishmanial urinary antigen detection in different patient groups We followed up the cases of 13 of the 16 patients from group 1. A urine specimen was collected each time they came to the hospital, and a bone marrow aspirate was also collected if a VL relapse was suspected. The LAT was performed for all urine samples obtained. The leishmanial antigen could be detected in urine up to 1 year after VL diagnosis in HIV-coinfected patients, probably due to their inability to control the parasitosis (4, 7). Therefore, this technique does not seem very helpful in monitoring treatment and predicting relapses. However, our results show that the test is a sensitive and specific noninvasive tool for diagnosing VL. It is easy to perform and interpret, so it could be used as a screening test in developing areas and among susceptible populations. However, due to the low number of patients with VL included in our evaluation, further studies are needed to confirm these data.

Performance of VITEK-2 Compact and overnight MicroScan panels for direct identification and susceptibility testing of Gram-negative bacilli from positive FAN BacT/ALERT blood culture bottles

We describe the reliability of the VITEK-2 Compact and overnight MicroScan panels for direct identification and susceptibility testing from the BacT/ALERT blood culture system when using FAN (FA and FN) bottles. A simple procedure, in two centrifugation steps, was designed to remove the charcoal particles present in FA and FN bottles. A total of 113 positive blood cultures showing Gram-negative rods were investigated. Enterobacteriaceae were isolated in 104 cases, and Pseudomonas aeruginosa in nine. The MicroScan system correctly identified 106 (93.8%) of the 113 isolates. The seven identificaction errors included P. aeruginosa (three), Enterobacter cloacae (one), Escherichia coli (one), Klebsiella oxytoca (one), and Klebsiella pneumoniae (one). The VITEK-2 system correctly identified 109 (96.5%) of the 113 samples obtained directly from the blood culture bottles. The four unidentified isolates were Enterobacter cloacae (two), Escherichia coli (one), and P. aeruginosa (one). MicroScan yielded 4/779 (0.5%) very major errors and 28/2825 (0.9%) minor errors. VITEK-2 yielded 2/550 (0.36%) very major errors, 1/1718 (0.05%) major error, and 32/2373 (1.3%) minor errors. Both systems provided excellent identification (correlation of >90%) and susceptibility (correlation of >98%) results. The average times required to obtain identification and susceptibility results using the direct test applied to the VITEK-2 Compact system were 4.57 +/- 1.37 h and 6.52 +/- 1.64 h, respectively. The VITEK-2 compact system provided results on the same day that the blood culture was found to be positive.

Improving the Diagnosis of Bloodstream Infections: PCR Coupled with Mass Spectrometry

The reference method for the diagnosis of bloodstream infections is blood culture followed by biochemical identification and antibiotic susceptibility testing of the isolated pathogen. This process requires 48 to 72 hours. The rapid administration of the most appropriate antimicrobial treatment is crucial for the survival of septic patients; therefore, a rapid method that enables diagnosis directly from analysis of a blood sample without culture is needed. A recently developed platform that couples broad-range PCR amplification of pathogen DNA with electrospray ionization mass spectrometry (PCR/ESI-MS) has the ability to identify virtually any microorganism from direct clinical specimens. To date, two clinical evaluations of the PCR/ESI-MS technology for the diagnosis of bloodstream infections from whole blood have been published. Here we discuss them and describe recent improvements that result in an enhanced sensitivity. Other commercially available assays for the molecular diagnosis of bloodstream infections from whole blood are also reviewed. The use of highly sensitive molecular diagnostic methods in combination with conventional procedures could substantially improve the management of septic patients.

Prospective study of drug-resistant tuberculosis in a spanish urban population including patients at risk for HIV infection

From January 1988 to October 1992, the primary resistance to first-line antituberculous drugs in 501 tuberculous patients was evaluated prospectively. Three-hundred and seventeen patients were HIV-negative and 184 were HIV-positive; these patients had several different clinical forms of tuberculosis. Moreover, the acquired resistance to antituberculous drugs was studied in 295 non-AIDS patients and in 42 AIDS patients with evidence of antecedent tuberculosis treatment. The data indicated that during these five years there was no consistent and clear-cut trend toward greater frequency of primary drug resistance to any of the first-line antituberculous drugs. Primary drug resistance in HIV-positive patients (7.1 %) did not differ significantly (p>0.05) from that found in HIV-negative patients (8.2 %). Among HIV-positive patients, the acquired drug resistance pattern was similar to that detected in HIV-negative patients although the frequency of resistance in the former (69 %) was significantly higher (p<0.01). During the study, resistance to isoniazid was almost constant in the acquired-resistance cases and was frequently associated with resistance to other drugs. Furthermore, the acquired resistance to isoniazid was often of a higher level (1 to 10 mg/l) than the primary resistance (0.2 mg/l), and those strains were usually catalase and peroxidase negative.

Comparison of a Nonradiometric System with Bactec 12B and Culture on Egg-Based Media for Recovery of Mycobacteria from Clinical Specimens

Abstract The MB/BacT (Organon-Teknika, USA) is a fully automated, rapid, nonradiometric system for the culture of mycobacteria from clinical samples. The rate of recovery of mycobacteria and the time to detection obtained with the MB/BacT were compared with those obtained with Löwenstein-Jensen and Coletsos solid media and Bactec 7H12 (12B) (Becton-Dickinson, USA) broth when 600 processed specimens were inoculated into all media in parallel. Specimens included 383 respiratory samples, 20 urine samples, 23 purulent exudates, 13 stool samples, 103 blood samples, 14 bone marrow aspirates, and 44 body fluid samples or aspirates. Overall, 106 mycobacterial isolates comprising six species were recovered, of which 100 (94.3%) were detected with MB/BacT, 98 (92.5%) with egg-based media, and 96 (90.2%) with Bactec 12B. The recovery rates of Mycobacteriumtuberculosis complex with MB/BacT, egg-based media, and Bactec 12B were 98.7%, 93.7, and 89.9%, respectively. The average number of days to detection of single mycobacterial isolates was 14.2 days for MB/BacT, 26.1 days for egg-based media, and 11.7 days for Bactec 12B. The contamination rates were higher in MB/BacT (5%) than in Bactec 12B (1.8%) or egg-based media (1.5%). MB/BacT is a reliable, nonradiometric, less labor-intensive alternative to Bactec 12B for recovery of mycobacteria, but, as with other liquid culture methods, MB/BacT should be used in combination with a solid medium, not on its own.

Pneumococcal bacteremia in children: an 8-year review in two hospitals in Barcelona

In this study, 90 episodes of pneumococcal bacteremia that occurred over an 8-year period in two hospitals in Barcelona were analyzed retrospectively to determine the clinical and bacteriological characteristics of pneumococcal bacteremia, the risk factors for antibiotic resistance, the outcome, and the vaccine coverage. The mean age of the patients was 3.1 years and the male/female ratio was 1.7. The overall rates of penicillin-non-susceptible, cefotaxime-non-susceptible, and erythromycin-resistant isolates were 48.8, 24.4, and 25.5%, respectively. Antibiotic resistance was associated with children under the age of 2 years and with previous antibiotic treatment. The percentage of antibiotic resistance was higher in the nine episodes that occurred in patients with an underlying illness. The most prevalent serotypes identified were 1, 14, 6B, 18C, 5, and 19A. Serotypes 6A/B, 14, and 19A/F were isolated primarily from children under 2, whereas serotypes 1 and 5 were recovered more frequently from older children. Apparent relationships between serotypes 6A/B, 14, and 19A/F and occult bacteremia and between serotypes 1 and 5 and bacteremic pneumonia were confounded by the age variable. The proportion of bacteremic episodes preventable by all (7-valent, 9-valent, and 11-valent) of the conjugate pneumococcal vaccines was 60% in children under 2. In older children, the serotype coverage rate for the three formulations was 48, 87, and 87%, respectively. In summary, these data expand upon previous Spanish studies in which serotypes 1 and 5 were reported to be among the leading causes of severe systemic pneumococcal infections in children over 2, findings that should be taken into consideration when planning vaccine programmes.