Juan Alberola

Enumeration of cytomegalovirus-specific interferonγ CD8+ and CD4+ T cells early after allogeneic stem cell transplantation may identify patients at risk of active cytomegalovirus infection

Recovery of functional cytomegalovirus (CMV)-specific T lymphocytes is critical for protection from active CMV infection and disease in allogeneic stem cell transplant recipients (Allo-SCT).[1][1]–[6][2] To date, assessment of CMV-specific T-cell immunity has not had a major impact on the clinical

Early neutralizing and glycoprotein B (gB)-specific antibody responses to human cytomegalovirus (HCMV) in immunocompetent individuals with distinct clinical presentations of primary HCMV infection

BACKGROUND Antibodies with functional anti-Human Cytomegalovirus (HCMV) activity are likely to be involved in preventing virus dissemination and thus may contribute to minimize the clinical manifestations of infection. OBJECTIVES To investigate the role of humoral immunity in modulating the clinical expression of primary Human Cytomegalovirus (HCMV) infection in immunocompetent persons. STUDY DESIGN Neutralizing (NA) and glycoprotein B (gB)-specific antibodies were quantitated in acute-phase and late-convalescence phase sera from 19 individuals who developed either HCMV mononucleosis (12) or oligosymptomatic hepatitis (seven). RESULTS The levels of NA in sera drawn early after infection were significantly lower in the former patients than in the latter (P=0. 032). This difference was not related to either the total serum IgG levels and anti-HCMV IgGs avidity or to the presence of higher viral loads in blood, as assessed by detecting serum HCMV DNA by PCR, in patients experiencing mononucleosis. Increased NA titers were seen in all available late-convalescence sera. In these sera, median NA levels were not significantly different among the study groups. Antibodies to HCMV gB of both IgG and IgM classes were detected in all acute-phase sera analyzed. Median anti-gB IgG and IgM titers did not differ significantly between study groups. Likewise, the IgG subclass reactivity pattern against gB was found to be similar for both groups. CONCLUSIONS The data revealed that an intense and early antibody response to gB developed in patients undergoing primary HCMV infection irrespective of the clinical manifestation of the disease. In contrast, a deficient NA response was observed in patients with HCMV mononucleosis versus that observed in patients displaying a milder form of disease-suggesting that the strength of NA response to HCMV generated early after infection might determine the severity of primary HCMV infection.

Influence of inadequate antimicrobial therapy on prognosis in elderly patients with severe urinary tract infections

BACKGROUND Inadequate empirical antimicrobial therapy (IEAT) in intensive care unit (ICU) is associated with adverse outcomes. However, the influence of IEAT on prognosis for elderly patients with urinary tract infection (UTI) in non-ICU settings is unknown. METHODS A retrospective cross-sectional study of elderly patients admitted to a non-ICU ward in a university hospital with a primary diagnosis of UTI over a 3-year period was done. Data relating to age, sex, background comorbidities, severity of infection, bacteremia, microorganisms isolated in urine, treatment given, length of stay and prognosis were obtained using chart review. Cases were segregated according to the adequacy of empirical antimicrobial therapy. In-hospital mortality rate was the main outcome variable evaluated. RESULTS A total of 270 patients with a mean age of 83.7years were studied. Sixty-eight percent were health-care associated infections. Seventy-nine (29.3%) cases received IEAT. IEAT was associated with previous hospitalization, urinary catheter and previous antibiotic. A Gram stain of urine with a gram-positive cocci was predictive of IEAT by multivariate analysis (OR, 6.29; 95% CI, 1.05-37.49). In-hospital mortality rate was 8.9%. IEAT (OR, 3.47; 95% CI, 1.42-8.48) was an independent risk factor for mortality along with APACHE II ≥15 (OR, 3.14; 95% CI, 1.24-7.90), dementia (OR, 3.10; 95% CI, 1.19-8.07) and neoplasia (OR, 3.49; 95% CI, 1.13-10.77). IEAT was not associated with length of stay in hospital. CONCLUSION IEAT is associated with mortality in elderly patients with UTI admitted to a non-ICU ward, suggesting that improving empirical antimicrobial therapy could have a favorable impact on prognosis.

Antibody response to human cytomegalovirus (HCMV) glycoprotein B (gB) in AIDS patients with HCMV end‐organ disease

Human cytomegalovirus (HCMV)‐specific antibody responses in HIV‐1 infected individuals either with or without HCMV end‐organ disease were examined to determine the whether development of HCMV disease was associated with a particular deficit in the antibody response. Anti‐whole HCMV, anti‐glycoprotein B (gB), and neutralizing antibody levels were higher in HIV‐1 infected individuals than in healthy immunocompetent subjects, particularly in patients with AIDS either with or without HCMV‐associated disease. Irrespective of location and spread of HCMV disease, patients who had received anti‐HCMV therapy prior to sampling exhibited significantly higher anti‐gB and neutralizing antibody titers than those who remained untreated. Likewise, patients with HCMV disease who were antigenemic or viremic had significantly lower anti‐gB and neutralizing antibody titers than those who tested negative in either assay. Patients with untreated HCMV disease had significantly lower antibody titers than AIDS patients without disease. Analysis of the IgG subclass antibody responses to gB revealed no significant differences among HIV‐1 infected individuals. These results suggest that levels of detectable anti‐gB and HCMV neutralizing antibodies are inversely related to systemic viral load. Thus, antibodies with such specificities may be relevant in preventing the establishment of HCMV‐associated disease or in modulating its progression. J. Med. Virol. 55:272–280, 1998.

Effect of Bacteremia in Elderly Patients With Urinary Tract Infection

Introduction The clinical effect of bacteremia on outcomes in urinary tract infection (UTI) is still debated. This study aims to examine the clinical effect of bacteremia in elderly patients with UTI requiring hospital admission. Methods This retrospective observational study recorded the clinical features, microbiology and outcomes in a Spanish cohort of patients aged ≥65 years hospitalized for UTI in whom blood cultures were performed in the emergency department. The primary outcome of the study was in‐hospital mortality. Results Of 333 patients, with a mean age of 81.6 years, 137 (41.1%) had positive blood cultures. Escherichia coli, with 223 (66.9%) cases, was the most common microorganism isolated. Independent risk factors of bacteremia were temperature >38°C, heart rate >90 bpm and inversely both Enterococcus faecalis and Pseudomonas aeruginosa. Bacteremia was not associated with the length of stay in hospital (6.96 ± 3.50 days versus 7.33 ± 5.54 days, P = 0.456). Mortality rate was 9.3% with no significant difference between bacteremic and nonbacteremic cases (8.8% and 9.7%, respectively, P = 0.773). In‐hospital mortality analyzed by logistic regression was associated with McCabe index >2 (20.5% survival versus 66.7% death, adjusted odds ratio = 6.31, 95% CI: 2.71‐14.67; P < 0.001) but not with bacteremia (41.4% survival versus 38.7% death, adjusted odds ratio = 0.99, 95% CI: 0.43‐2.29; P = 0.992). Conclusions Our study suggests that the presence or absence of bacteremia in elderly people with UTI requiring hospitalization does not have an influence on outcomes such as in‐hospital mortality or length of stay.

Becton Dickinson Directigen EZ Flu A+B assay in the diagnosis of pandemic influenza A H1N1 2009 virus infection in adult patients.

To the editor: The recent emergence and spread of the pandemic influenza A H1N1 2009 virus demands the evaluation of rapid antigen assays for their ability to detect this novel subtype of influenza A virus. Data on the ability of BD Directigen EZ Flu A+B immunochromatographic (IC) assay (Beckton Dickinson and Company, Sparks, MD, USA) to detect the pandemic influenza A virus strain in fresh clinical samples have been recently published. 1 , 2 , 3 , 4 , 5 In these studies, the majority of specimens were collected from pediatric patients, and the sensitivities reported ranged from 46·8% to 76·6%. As viral shedding in the upper respiratory tract during influenza virus infection is of greater magnitude in children than in adults, the clinical utility of IC tests may indeed depend on the patient age. 6 We wish to report on our experience regarding the diagnostic and analytical performance characteristics of the Directigen EZ Flu A+B in a cohort of adults (≥18 years old) presenting with an influenza‐like syndrome at a tertiary Spanish hospital (Peset Aleixandre, Valencia Spain). A total 274 nasopharyngeal swabs from unique patients (median age of 50 years, range 18–97 years; 145 women and 129 men) and collected between July and September 2009 were included in the study. The specimens were obtained within 72 hours after the onset of symptoms by means of flexible nasopharyngeal nylon flocked swabs, placed in 3 ml of transport medium (Universal transport medium; Beckton Dickinson) and delivered to the Microbiology laboratory within 1 hours of collection. The specimens were vortexed and tested by the IC assay following the instructions of the manufacturer. 7 Samples were assayed by RT‐PCR within 24 hours after reception. Total RNA was extracted by the MagNApure extraction kit in the MagNA Pure robot (Roche Diagnostics, Basel, Switzerland), and RT‐PCR was performed by use of the Realtime Ready Influenza A/H1N1 Detection Set on the LightCycler® 2.0 instrument (Roche Diagnostics). 8 , 9 The overall positive rate for novel influenza A virus RNA as determined by real‐time PCR was 15·3%. Forty‐two specimens tested positive by RT‐PCR, of which 18 gave a positive IC result. The remaining 232 specimens tested negative by RT‐PCR. All these specimens gave a negative result in the IC assay. The overall agreement between the two assays was 91·2% (250/274), and the sensitivity, specificity, positive predictive value, and negative predictive value (adjusted to the prevalence in our cohort) were of 42·8%, 100%, 100%, and 79·8%, respectively. Cycle threshold (Ct) values for samples testing positive by the IC assay (median, 24·1, range, 20·5–33·6) were significantly lower (P = 0·001, by the Mann–Whitney test) than those for specimens yielding a negative result (median, 31·5, range, 30·2–34·5). To determine the analytical sensitivity of the IC assay, a local influenza strain (A/Valencia/1/2009H1N1v) isolated in Mardin Darby Canine Kidney cells was used. The viral stock (50% tissue culture infectious dose‐TCID50‐/ml of log10 7·0) was serially diluted in viral transport medium and tested in duplicate by IC. The limit of detection of the BD Directigen assay was approximately log10 4·5 TCID50/ml, which is in keeping with previous estimations. 3 , 10 In summary, the sensitivity of the Directigen EZ Flu A+B assay for the diagnosis of pandemic influenza A virus infection is clearly suboptimal and appears to be lower than that reported in studies conducted in either pediatric or mixed children and adult cohorts. Thus, molecular testing should be mandatory when a negative IC result is obtained, particularly in adult patients with a high pretest probability of infection. Nevertheless, given the specificity of the assay, a positive IC result may be safely used in making decisions regarding the instauration of antiviral treatments or implementation of infection control measures.

How is post-mortem microbiology appraised by pathologists? Results from a practice survey conducted by ESGFOR

Post-mortem microbiology (PMM) is an important tool in forensic pathology, assisting to determine the cause and manner of death. However, there is a lack of standardisation of PMM sampling. In order to get a better insight into the methods used, the available technical options and developmental needs, ESCMID Study Group for Forensic and Postmortem Microbiology (ESGFOR) members designed a survey aimed at pathologists regarding common practices of PMM in clinical and forensic autopsies. Multiple choice questions were developed based on Cumulative Techniques and Procedures in Clinical Microbiology (Cumitech). The questionnaire was sent to pathologists mainly across Europe and Turkey using SurveyMonkey. The survey had 147 respondents. Although all pathologists were aware of the existence of PMM, 39% (19/49) of the participants were not using it. The three main indications for PMM were: (i) clinical suspicion of an infection not confirmed antemortem (83%), (ii) infectious signs at autopsy (83%) and (iii) as part of a standard protocol for foetal/perinatal or paediatric death (67%). Almost 80% of the participants using PMM stated taking 1–10 samples per case. Of the requested examinations, a general bacteriological culture (96%) and a specific polymerase chain reaction (PCR) assay for a particular infectious agent (34%) were most popular. The most frequent samples were: heart blood (66%), peripheral femoral blood (49%), spleen (64%) and lung (56%). Eighty-nine percent of the participants considered PMM a useful resource when investigating the cause of death. Although there are some common uses, this survey indicates that there is a need for improvement towards standardising sampling procedures in PMM.

Prospective cohort study of risk factors for extended-spectrum ß-lactamase-producing Escherichia coli urinary tract infections in elderly patients admitted to hospital

Extended‐spectrum beta‐lactamase (ESBL)‐producing Escherichia coli are currently common in community‐onset infections, limiting therapeutic options. In this work we aim to identify the prevalence of and risk factors for ESBL‐producing E. coli in elderly patients with urinary tract infections (UTI) admitted to hospital.

Post-mortem microbiology in sudden death: sampling protocols proposed in different clinical settings

BACKGROUND Autopsies, including minimally invasive autopsies, are a powerful tool for determination of the cause of death. When a patient dies from an infection, microbiology is crucial to identify the causative organism. Post-mortem microbiology (PMM) aims to detect unexpected infections causing sudden deaths; confirm clinically suspected but unproven infection; evaluate the efficacy of antimicrobial therapy; identify emergent pathogens; and recognize medical errors. Additionally, the analysis of the thanatomicrobiome may help to estimate the post-mortem interval. AIMS The aim was to provide advice in the collection of PMM samples and to propose sampling guidelines for microbiologists advising autopsy pathologists facing different sudden death scenarios. SOURCES A multidisciplinary team with experts in various fields of microbiology and autopsies on behalf of the ESGFOR (ESCMID - European Society of Clinical Microbiology and Infectious Diseases - study group of forensic and post-mortem microbiology and in collaboration with the European Society of Pathology) developed this narrative review based on a literature search using MedLine and Scopus electronic databases supplemented with their own expertise. CONTENT These guidelines address measures to prevent sample contamination in autopsy microbiology; general PMM sampling technique; protocols for PMM sampling in different scenarios and using minimally invasive autopsy; and potential use of the evolving post-mortem microbiome to estimate the post-mortem interval. IMPLICATIONS Adequate sampling is paramount to identify the causative organism. Meaningful interpretation of PMM results requires careful evaluation in the context of clinical history, macroscopic and histological findings. Networking and closer collaboration among microbiologists and autopsy pathologists is vital to maximize the yield of PMM.

Assessing the risk of cytomegalovirus DNAaemia in allogeneic stem cell transplant recipients by monitoring oxidative-stress markers in plasma

The level of antioxidants, such as thiol-containing tripeptide glutathione (GSH), in cytomegalovirus (CMV)-infected cells is notably increased. We previously showed that GSH levels in plasma, as measured by untargeted 1H nuclear magnetic resonance, are higher in allogeneic stem cell transplant (allo-SCT) recipients who subsequently develop CMV viraemia. We hypothesized that the net level of oxidative-stress markers present in plasma may be reduced in patients who develop CMV DNAaemia compared to those who do not. We serially monitored the levels of malondialdehyde (MDA) and carbonylated proteins (CPs) early after allo-SCT and assessed whether they could predict the occurrence of CMV DNAaemia. MDA levels were measured in 43 patients (28 had CMV DNAaemia) and CPs were quantified in 53 patients (38 patients developed CMV DNAaemia). The area under the curve (AUC) value for MDA, but not for CPs, was significantly lower in patients who subsequently developed CMV DNAaemia compared to those who remained DNAaemia-free (P=0.043). A trend toward lower MDA AUC values was observed in episodes of CMV DNAaemia with faster CMV replicative kinetics and in those who reached higher peak CMV DNA levels. Moreover, receiver operating characteristic curve analyses indicated that the MDA biomarker had the predictive ability to discriminate between patients with or without subsequent CMV DNAaemia (AUC=0.69, 95 % confidence interval 0.51-0.85, P=0.05). In summary, serial quantitation of MDA may be useful for individualizing antiviral prophylaxis therapies (targeted prophylaxis) in the upcoming era of new antiviral drugs with improved safety profiles.