Rapid identification,fluconazole and micafungin susceptibility testing of Candida species from blood culture by a short incubation method.

This study aimed to develop and validate a rapid method for identification by MALDI-TOF system and determination of the susceptibility to Fluconazole and Micafungin by broth microdilution among Candidaspecies causing bloodstream infections. Subcultures from blood culture bottles were incubated for 5 hours (+/- 1h) and used to perform the tests, so that the turnaround time of rapid identification and susceptibility profile was about 5 and 24 hours, respectively. The rapid identification showed agreement of 92.05 %. Regarding the rapid broth microdilution for Fluconazole and Micafungin, the agreement was 97.06 % (p<0.001) and 100 % (p<0.001), and the Kappa coefficient was 0.91 (p<0.001) and 1.0 (p<0.001), respectively. To conclude, both rapid methods showed to be reproducible, inexpensive, easy to perform and time-saving. Thus, these methodologies could be useful to guide and adjust empirical antifungal therapy.

Rapid bacterial identification by MALDI-TOF MS directly from blood cultures and rapid susceptibility testing: A simple approach to reduce the turnaround time of blood cultures.

Antimicrobial treatment of patients with bloodstream infections (BSI) is time-sensitive. In an era of increasing antimicrobial resistance, rapid detection and identification of bacteria with antimicrobial susceptibility are critical for targeted therapy early in the disease course. This study describes the performance of a rapid method for identifying and testing antimicrobial susceptibility of Gram-negative bacteria performed directly from blood culture bottles in a routine microbiology laboratory. A total of 284, 120, and 24 samples were analyzed by rapid identification (Rid), rapid susceptibility testing (RAST), and rapid broth microdilution for polymyxin B (rMIC), respectively, and compared with standard methods. Our protocol was able to identify 93% of isolates at the species level using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). We obtained 100% agreement for RAST compared to the standard method and 96% agreement for rMIC. Our protocol has proven to be an excellent tool for rapid identification of Gram-negative bacilli causing BSIs. It can also be used in microbiology laboratory routine along with RAST and faster polymyxin microdilution, especially for carbapenemase-producing bacteria, allowing for rapid, simple, accurate, and cost-effective diagnosis.

Rapid bacterial identification by MALDI-TOF MS directly from blood cultures and rapid susceptibility testing: A simple approach to reduce the turnaround time of blood cultures

Abstract Antimicrobial treatment of patients with bloodstream infections (BSI) is time-sensitive. In an era of increasing antimicrobial resistance, rapid detection and identification of bacteria with antimicrobial susceptibility are critical for targeted therapy early in the disease course. This study describes the performance of a rapid method for identifying and testing antimicrobial susceptibility of Gram-negative bacteria performed directly from blood culture bottles in a routine microbiology laboratory. A total of 284, 120, and 24 samples were analyzed by rapid identification (Rid), rapid susceptibility testing (RAST), and rapid broth microdilution for polymyxin B (rMIC), respectively, and compared with standard methods. Our protocol was able to identify 93% of isolates at the species level using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). We obtained 100% agreement for RAST compared to the standard method and 96% agreement for rMIC. Our protocol has proven to be an excellent tool for rapid identification of Gram-negative bacilli causing BSIs. It can also be used in microbiology laboratory routine along with RAST and faster polymyxin microdilution, especially for carbapenemase-producing bacteria, allowing for rapid, simple, accurate, and cost-effective diagnosis.

Diversity of Fusarium species causing invasive and disseminated infections.

INTRODUCTION: Invasive fusariosis (IF) is considered an emerging fungal disease and an important problem worldwide that increasingly affects immunocompromised individuals. There is currently concern about establishing the genetic diversity and phylogenetic relationship of the species Fusarium causing invasive fusariosis. MATERIALS AND METHODS: The aim of this study was to characterize the molecular profile and morphological characteristics of Fusarium species isolated from 21 patients with invasive fusariosis. Multilocus sequence typing was performed for molecular identification of the following genes: the second largest subunit of the RNA polymerase gene (RPB2) and elongation factor 1 alpha (EF-1α). The morphological features of different species were carefully described and revised by experienced mycologists. RESULTS: Morphological and molecular analyses revealed that the F. solani species complex (FSSC) and F. oxysporum species complex (FOSC) were the most common species isolated from patients with invasive fusariosis; FSSC-2 h (5), FSSC-1 (2) and FOSC-183 (2) were the most frequent haplotypes. The macroscopic characterization revealed great variation in the tonalities of the FSSC colonies and particularities amongst the species in relation to the macroconidia structures, while the FOSC was more homogeneous and presented shades from white to lilac. CONCLUSIONS: Our study characterized the diversity, haplotypes, and morphological aspects of Fusarium species and the haplotypes prevalent in patients with invasive fusariosis. FSSC and FSSC-2 h were the predominant species and haplotype, respectively. Although we have described interesting morphological aspects in Fusarium species, particularly haplotypes, their identification cannot rely on phenotypical aspects. Molecular biology techniques are necessary and should be introduced for routine use in mycology laboratories.

Diagnostic Accuracy of CD64 for Sepsis in Emergency Department.

INTRODUCTION: Sepsis is a systemic inflammatory response to suspected or confirmed infection. Clinical evaluations are essential for its early detection and treatment. Blood cultures may take as long as 2 days to yield a result and are not always reliable. However, recent studies have suggested that neutrophil CD64 expression may be a sensitive and specific alternative for the diagnosis of systemic infection. OBJECTIVE: The objective of the study was to analyze the difference in CD64 values between subjects with systemic inflammatory response syndrome (SIRS), suspected or confirmed sepsis, who meet diagnostic criteria for SIRS upon arriving at an emergency department. MATERIALS AND METHODS: This was a prospective observational cohort study, an accuracy study of CD64 prospectively evaluated. The sample consisted of 109 patients aged 18 years with criteria for SIRS on arrival to emergency department. CD64 expression was measured within 6 h of hospital admission and once again after 48 h. RESULTS: ROC curve analysis suggested that a cutoff of 1.45 for CD64 expression could diagnose sepsis with a sensitivity of 0.85, a specificity of 0.75, an accuracy of 82.08%, a positive predictive value of 0.96, a negative predictive value of 0.38 and a positive likelihood ratio of 3.33. The area under the curve was 0.83. CONCLUSION: CD64 seems to be a useful, sensitive, and specific biomarker in discriminating between SIRS and sepsis.

Antifungal Susceptibility, Morphological and Molecular Characterization of Lasiodiplodia theobromae Isolated from a Patient with Keratitis.

Lasiodiplodia theobromae is a rare ocular pathogen. We report a patient with fungal keratitis caused by L. theobromae. The patient was a 75-year-old male, a farmer with diabetes type II, and no previous history of ocular trauma. Histopathology analysis revealed the presence fungi invading Descemet's membrane of the cornea. The fungus was characterized by septate, highly bulged fungal filaments involving full corneal thickness in the corresponding histopathology specimens. A dematiaceous mold was isolated and initally identified as L. theobromae by microscopic and macroscopic morphology, and further confirmed by PCR-based determination of internal transcribed spacer (ITS) regions of ribosomal DNA. Antifungal susceptibility tests showed sensitivity to amphotericin B (AMB) and voriconazole ( VRC), and resistance to other azoles, including itraconazole (ITC) and fluconazole (FLC). Corneal transplant was performed. Despite in vitro itraconazole resistance, the patient was successfully treated with oral itraconazole, topical voriconazole and natamycin, combined with ocular injections of amphotericin B and voriconazole.

Relationship between intracranial pressure and antifungal agents levels in the CSF of patients with cryptococcal meningitis.

The purpose of this study was to evaluate the influence of intracranial hypertension in the cerebrospinal fluid (CSF) levels of amphotericin B and fluconazole levels of patients with cryptococcal meningitis. CSF samples and intracranial pressure were obtained by means of routine punctures performed at days 1, 7, and 14 of therapy, respectively. Amphotericin B and fluconazole CSF levels were measured by HPLC method as previously described. The minimum inhibitory concentration for amphotericin B, fluconazole, 5΄flucytosine, and voriconazole of each Cryptococcus isolate was performed according to CLSI. The predominant Cryptococcus species found was C. neoformans, and the major underlying condition was AIDS. Only one CSF sample had a detectable level for amphotericin B during the 14 days of therapy. Fluconazole CSF levels progressively increased from day 1 to day 14 of therapy for most cases. Fluconazole levels in the CSF were above the minimum inhibitory concentrations (MICs) for Cryptococcus during the initial 14 days of antifungal therapy. Variations of intracranial pressure did not affect amphotericin B and fluconazole levels in the CSF. The generalized estimating correlation (GEE) and Spearman correlation test (SCT) showed no significant correlation between the amphotericin B or fluconazole concentrations in the CSF and intracranial pressure (P = .953 and P = .093, respectively for GEE test and P = .477 and P = .847, respectively, for SCT). Combination therapy of amphotericin B with fluconazole was effective in 60% of the patients considering CSF cultures were negative in 9 of 15 patients after 14 days of therapy. Further studies are necessary to evaluate the role of intracranial hypertension on the therapeutic efficacy of different antifungal agents in patients with cryptococcal meningitis.

Human immunodeficiency virus in a cohort of systemic lupus erythematosus patients.

BACKGROUND: Systemic lupus erythematosus (SLE) and acquired immunodeficiency syndrome (AIDS) share many clinical manifestations and laboratory findings, therefore, concomitant diagnosis of SLE and human immunodeficiency virus (HIV) can be challenging. METHODS: Prospective cohort with 602 patients with SLE who attended the Rheumatology Clinic of the Hospital de Clínicas de Porto Alegre since 2000. All patients were followed until 01 May 2015 or until death, if earlier. Demographic, clinical and laboratory data were prospectively collected. RESULTS: Out of the 602 patients, 11 presented with the diagnosis of AIDS (1.59%). The following variables were significantly more prevalent in patients with concomitant HIV and SLE: neuropsychiatric lupus (10.9% vs. 36.4%; p = 0.028) and smoking (37.6% vs. 80%; p = 0.0009) while malar rash was significantly less prevalent in this population (56% vs. 18.2%; p = 0.015). Nephritis (40.5% vs. 63.6%; p = 0.134) and hemolytic anemia (28.6% vs. 54.5%; p = 0.089) were more prevalent in SLE patients with HIV, but with no statistical significance compared with SLE patients without HIV. The SLICC damage index median in the last medical consultation was significantly higher in SLE patients with HIV (1 vs. 2; p = 0,047). CONCLUSIONS: Our patients with concomitant HIV and SLE have clinically more neuropsychiatric manifestations. For the first time, according to our knowledge, higher cumulative damage was described in lupus patients with concomitant HIV infection. Further studies are needed to elucidate this complex association, its outcomes, prognosis and which therapeutic approach it's best for each case.

Outcomes and Risk Factors Associated with Clostridium difficile Diarrhea in Hospitalized Adult Patients.

Background. The epidemiology of Clostridium difficile infection has changed over time. Therefore, it is essential to monitor the characteristics of patients at risk of infection and factors associated with poor prognosis. Objective. To evaluate factors associated with C. difficile infection and with poor prognosis in those with documented C. difficile colitis. Methods. A retrospective case-control study of 75 patients with documented C. difficile colitis and 75 controls with hospital-acquired diarrhea of other causes. Stepwise multiple logistic regression was used to identify factors associated with C. difficile infection among patients with hospital-acquired diarrhea. Results. Previous antibiotic treatment (odds ratio (OR), 13.3; 95% confidence interval (CI), 1.40-126.90), abdominal distension (OR, 3.85; 95% CI, 1.35-10.98), and fecal leukocytes (OR, 8.79; 95% CI, 1.41-54.61) are considered as predictors of C. difficile colitis; anorexia was negatively associated with C. difficile infection (OR, 0.15; 95% CI, 0.03-0.66). Enteral tube feeding was independently associated with a composite outcome that included in-hospital mortality, intensive care unit admission, and treatment failure (OR, 3.75; 95%CI, 1.24-11.29). Conclusions. Previous antibiotic use and presence of fecal leukocytes in patients with hospital-acquired diarrhea are associated with C. difficile colitis and enteral tube support with complications associated with C. difficile colitis.

Consensus of the Brazilian Society of Infectious Diseases and Brazilian Society of Clinical Oncology on the management and treatment of Kaposi's sarcoma

Kaposi's sarcoma is a multifocal vascular lesion of low-grade potential that is most often present in mucocutaneous sites and usually also affects lymph nodes and visceral organs. The condition may manifest through purplish lesions, flat or raised with an irregular shape, gastrointestinal bleeding due to lesions located in the digestive system, and dyspnea and hemoptysis associated with pulmonary lesions. In the early 1980s, the appearance of several cases of Kaposi's sarcoma in homosexual men was the first alarm about a newly identified epidemic, acquired immunodeficiency syndrome. In 1994, it was finally demonstrated that the presence of a herpes virus associated with Kaposi's sarcoma called HHV-8 or Kaposi's sarcoma herpes virus and its genetic sequence was rapidly deciphered. The prevalence of this virus is very high (about 50%) in some African populations, but stands between 2% and 8% for the entire world population. Kaposi's sarcoma only develops when the immune system is depressed, as in acquired immunodeficiency syndrome, which appears to be associated with a specific variant of the Kaposi's sarcoma herpes virus. There are no treatment guidelines for Kaposi's sarcoma established in Brazil, and thus the Brazilian Society of Clinical Oncology and the Brazilian Society of Infectious Diseases developed the treatment consensus presented here.

Aetiology of bacteraemia as a risk factor for septic shock at the onset of febrile neutropaenia in adult cancer patients.

Septic shock (SS) at the onset of febrile neutropaenia (FN) is an emergency situation that is associated with high morbidity and mortality. The impact of the specific aetiology of bloodstream infections (BSIs) in the development of SS at the time of FN is not well established. The aim of this study was to evaluate the association between the aetiology of BSIs and SS at the time of FN in hospitalised adult cancer patients. This prospective cohort study was performed at a single tertiary hospital from October 2009 to August 2011. All adult cancer patients admitted consecutively to the haematology ward with FN were evaluated. A stepwise logistic regression was conducted to verify the association between the microbiological characteristics of BSIs and SS at the onset of FN. In total, 307 cases of FN in adult cancer patients were evaluated. There were 115 cases with documented BSI. A multivariate analysis showed that polymicrobial bacteraemia (P = 0.01) was associated with SS. The specific blood isolates independently associated with SS were viridans streptococci (P = 0.02) and Escherichia coli (P = 0.01). Neutropaenic cancer patients with polymicrobial bacteraemia or BSI by viridans streptococci or Escherichia coli are at increased risk for SS at the time of FN.

Consensus of the Brazilian Society of Infectious Diseases and Brazilian Society of Clinical Oncology on the management and treatment of Kaposi's sarcoma.

Kaposi's sarcoma is a multifocal vascular lesion of low-grade potential that is most often present in mucocutaneous sites and usually also affects lymph nodes and visceral organs. The condition may manifest through purplish lesions, flat or raised with an irregular shape, gastrointestinal bleeding due to lesions located in the digestive system, and dyspnea and hemoptysis associated with pulmonary lesions. In the early 1980s, the appearance of several cases of Kaposi's sarcoma in homosexual men was the first alarm about a newly identified epidemic, acquired immunodeficiency syndrome. In 1994, it was finally demonstrated that the presence of a herpes virus associated with Kaposi's sarcoma called HHV-8 or Kaposi's sarcoma herpes virus and its genetic sequence was rapidly deciphered. The prevalence of this virus is very high (about 50%) in some African populations, but stands between 2% and 8% for the entire world population. Kaposi's sarcoma only develops when the immune system is depressed, as in acquired immunodeficiency syndrome, which appears to be associated with a specific variant of the Kaposi's sarcoma herpes virus. There are no treatment guidelines for Kaposi's sarcoma established in Brazil, and thus the Brazilian Society of Clinical Oncology and the Brazilian Society of Infectious Diseases developed the treatment consensus presented here.

The Higher Frequency of Blood Group B in a Brazilian Population with HIV Infection.

OBJECTIVE: To analyze the frequency of and odds for and against HIV infection based on ABO blood type in a large sample of blood donors. BACKGROUND: Coevolution between pathogens and hosts may explain the ABO system of polymorphisms. HIV-infected cells add ABO(H) blood group antigens to the viral envelope. Naturally occurring antibodies against ABO(H) antigens that are present in normal human sera are able to neutralize ABO-expressing HIV in vitro. Blood donors are ideal for studying blood groups and HIV infection in vivo because all donors are routinely typed and tested. METHODS: All blood donors who donated blood between 1994 and 2010 were tested for HIV (ELISA antibody tests and Western blot test or immunofluorescence testing) and were ABO typed (direct and reverse grouping tests). HIV infection based on the ABO blood group was analyzed using the chi-square test and game theory. RESULTS: The total number of examined blood donors during this period was 271,410, of whom 389 were infected with HIV. B-group donors were more infected than non-B donors (p= 0.006). CONCLUSIONS: A more restricted antigen recognition capacity of anti-Galα1-3Gal in blood groups AB and B and a weaker antigen-binding capacity of anti-A antibodies may contribute to a higher frequency of HIV infection in blood group B.

Microbiologic isolates and risk factors associated with antimicrobial resistance in patients admitted to the intensive care unit in a tertiary care hospital.

This study reviewed the clinical and microbiologic data for patients admitted to the intensive care unit with hospital-acquired infections. In the multivariate analysis, AIDS and previous antibiotic use were associated with the emergence of multiresistant bacteria. Hematologic diseases, length of stay, number of days on central venous catheter, antimicrobial use, and presence of multiresistant bacteria were associated with death. The previous use of antibiotics and the length of the hospital stay contribute to the development of infections caused by multiresistant gram-negative bacteria.

Autologous transplant: microbial contamination of hematopoietic stem cell products.

Hematopoietic progenitor cells from peripheral blood (HPCPB) are commonly used for autologous and allogenic transplants in patients with most various onco-hematological diseases, and despite the utilization of sterile techniques during collection and processing of these products, bacterial contamination can occur. This study aimed to investigate the microbial contamination of HPCPB products. Microbial cultures of 837 HPCPB products between the year 2000 and 2009 were retrospectively analyzed to determine the incidence of culture positivity and identify the main organisms that cause contamination. The microbiological studies were performed with an automated system (BacT/Alert(®) bioMérieux Corporate). Thirty-six (4.3%) of 837 microbial cultures were contaminated. Coagulase-negative Staphylococcus was the most frequent bacteria isolated from HPCPB products (20 [56%] of the 36 positive microbial cultures). Considering the 36 contaminated samples, 22 HPCPB products were infused and 14 discarded. Pre- and post-infusion antibiotic therapy of the patients transfused with contaminated products was established based on the isolated microorganism and its antibiogram. Microbial contamination rate of HPCPB products was low. Clinically significant outcomes after infusion of contaminated HPCPB products were not observed.

Autologous transplant: microbial contamination of hematopoietic stem cell products

Hematopoietic progenitor cells from peripheral blood (HPCPB) are commonly used for autologous and allogenic transplants in patients with most various onco-hematological diseases, and despite the utilization of sterile techniques during collection and processing of these products, bacterial contamination can occur. This study aimed to investigate the microbial contamination of HPCPB products. Microbial cultures of 837 HPCPB products between the year 2000 and 2009 were retrospectively analyzed to determine the incidence of culture positivity and identify the main organisms that cause contamination. The microbiological studies were performed with an automated system (BacT/Alert® bioMérieux Corporate). Thirty-six (4.3%) of 837 microbial cultures were contaminated. Coagulase-negative Staphylococcus was the most frequent bacteria isolated from HPCPB products (20 [56%] of the 36 positive microbial cultures). Considering the 36 contaminated samples, 22 HPCPB products were infused and 14 discarded. Pre-and post-infusion antibiotic therapy of the patients transfused with contaminated products was established based on the isolated microorganism and its antibiogram. Microbial contamination rate of HPCPB products was low. Clinically significant outcomes after infusion of contaminated HPCPB products were not observed.

Hand hygiene, and not ertapenem use, contributed to reduction of carbapenem-resistant Pseudomonas aeruginosa rates.

OBJECTIVE: To evaluate the impact of ertapenem use in Pseudomonas aeruginosa carbapenem resistance, taking into account the volume of antimicrobial consumption, the consumption by the entire hospital of alcohol-based antiseptic hand rub, and the density rate of invasive practices. DESIGN: Before-and-after trial. SETTING: A tertiary care university hospital in southern Brazil. METHODS: Ertapenem was first added to the hospital formulary in June 2006, and it was excluded in February 2009. We evaluated Pseudomonas aeruginosa resistance rates through 3 study periods: period 1, before ertapenem use (17 months); period 2, during ertapenem use (33 months); and period 3, after exclusion of ertapenem (15 months). RESULTS: After introduction of ertapenem, there was a significant decrease in median consumption of imipenem or meropenem, from 2.6 to 2.2 defined daily doses (DDDs) per 100 patient-days (level change from 0.04 to -1.08; P < .01), and an increase in the use of these medications after ertapenem exclusion, from 2.2 to 3.3 DDDs per 100 patient-days (level change from -0.14 to 0.91; P < .01), by segmented regression analysis. There was no difference in the incidence density of carbapenem-resistant P. aeruginosa infection related to ertapenem use throughout the study periods. However, by multiple regression analysis, the reduction in the rate of carbapenem-resistant P. aeruginosa infection correlated significantly with the increase in the volume of alcohol used as hand sanitizer, which was from 660.7 mL per 100 patient-days in period 1 to 2,955.1 mL per 100 patient-days in period 3 (P = .04). Ertapenem use did not impact the rate of carbapenem-resistant P. aeruginosa infection. CONCLUSIONS: Use of alcohol-based hand gel, rather than ertapenem, was associated with a reduction in the rates of carbapenem-resistant P. aeruginosa infection. Measures to reduce resistance must include factors other than just antimicrobial stewardship programs alone.

Disseminated Histoplasma capsulatum and Cryptococcus neoformans co-infection in patients with AIDS.

Histoplasmosis and cryptococcosis are the most prevalent systemic mycoses in HIV-infected patients. The authors report a 20-year-old Brazilian HIV-positive woman with concomitant disseminated histoplasmosis and cryptococcosis. In addition, we review the reported cases described in the medical literature.

Biopsy of peripheral lymph nodes: a useful tool to diagnose opportunistic diseases in HIV-infected patients.

Peripheral lymphadenopathy is commonly present in HIV-infected patients and has a wide spectrum of differential diagnoses. We carried out a cross-sectional study of peripheral lymph node biopsies performed from January 2004 to December 2008 in HIV-infected patients who attended a tertiary-care hospital in southern Brazil. Only 60 of 210 peripheral lymph node biopsies performed (28%) were non-diagnostic. The most common diagnoses included: mycobacteriosis (105 cases; 50.2%); lymphoma (19 cases; 9.0%); systemic mycosis (12 cases; 5.7%) including histoplasmosis, cryptococcosis and histoplasmosis; and metastatic cancer (2.9%). Peripheral lymph node biopsy is a simple and useful tool to diagnose opportunistic diseases in HIV-infected patients.

In-hospital mortality of disseminated tuberculosis in patients infected with the human immunodeficiency virus.

BACKGROUND: Tuberculosis (TB) is a cause of significant morbidity and mortality in patients with AIDS. The goal of our study was to determine predictors of in-hospital mortality in patients with AIDS and disseminated tuberculosis in a middle-income country. MATERIAL AND METHODS: We conducted a retrospective cohort study in a tertiary care center, for patients with AIDS in southern Brazil. From 1996 to 2008, all patients with the diagnosis of disseminated TB were included. RESULTS: Eighty patients were included. In-hospital mortality was 35% (N = 28). On multivariate Cox regression analysis, low basal albumin (P < .01) was associated with death, and fever at admission was related to better survival (P < .01). CONCLUSION: Albumin levels or fever are independent predictors of survival in patients with HIV and disseminated TB. They can serve as indirect markers of immunodeficiency in patients with disseminated TB and AIDS.