Evaluation of a new rapid diagnostic test for the detection of influenza and RSV / Evaluación de un nuevo test de diagnóstico rápido para la detección de influenza y VRS

INTRODUCTION: Influenza viruses and respiratory syncytial virus (RSV) can cause an acute respiratory disease that occurs seasonally in epidemic waves. This retrospective study was conducted to evaluate the Sofia®Influenza A + B and the Sofia® RSV fluorescence immunoassays (FIAs), two novel rapid detection tests (RDTs) for influenza A and B and RSV. METHODS: Two hundred and nine breath samples were selected from patients with respiratory symptoms determined to be positive/negative for influenza A, influenza B or RSV using one of the reference diagnostic techniques, cell culture and/or RT-PCR (Simplexa™Flu A/B & RSV). The Sofia Influenza A + B FIA was tested on 123 samples (63 from children and 60 from adults) and the Sofia RSV FIA was tested on 86 pediatric samples. Sensitivity and specificity values of both assays were calculated assuming the reference techniques as the gold standard. RESULTS: Sensitivity and specificity values for the Sofia Influenza A + B FIA were 73.1% and 97.8%, respectively. Sensitivity and specificity values for the Sofia RSV FIA were 87.5% and 86.7%, respectively. CONCLUSION: The sensitivity results obtained for the two assays were considerably higher than those reported for other RDTs. In conclusion, the Sofia Influenza A + B and the Sofia RSV FIAs are appropriate tools for the rapid diagnosis of these viruses

INTRODUCCIÓN: Los virus influenza A y B y el virus respiratorio sincitial (VRS) causan infecciones respiratorias agudas de forma estacional en olas epidémicas. Este estudio restrospectivo se llevó a cabo para evaluar dos nuevos tests de diagnóstico rápido para detectar los virus influenza A y B y VRS: Sofia® Influenza A + B y Sofia® RSV Fluorescence Immunoassays (FIAs). MÉTODOS: Se seleccionaron 209 muestras respiratorias procedentes de pacientes con síntomas respiratorios en las que se había establecido el diagnóstico de presencia o ausencia de influenza A, influenza B o VRS mediante las técnicas de referencia: cultivo celular o RT-PCR PCR (Simplexa™Flu A/B & RSV). Sofia Influenza A + B se realizó en 123 muestras (63 de niños y 60 de adultos) y Sofia RSV en 86 muestras pediátricas. Se calcularon los valores de sensibilidad y especificidad de ambos test tomando las técnicas de referencia como gold standard. RESULTADOS: Los valores de sensibilidad y especificidad de Sofia Influenza A + B fueron 73,1 y 97,8%. Los valores de sensibilidad y especificidad de Sofia RSV fueron 87,5 y 86,7%. CONCLUSIONES: Los valores de sensibilidad obtenidos para ambos tests son mayores que los descritos para otras técnicas de diagnóstico rápido. Por lo tanto, Sofia Influenza A + B FIA y Sofia RSV FIA son test adecuados para el diagnóstico rápido de influenza y VRS

Multiclonal spread of VIM-1-producing Enterobacter cloacae isolates associated with In624 and In488 integrons located in an IncHI2 plasmid.

Over a 6-year period (2007-2012), the emergence of Enterobacter cloacae isolates resistant to ß-lactams and with reduced susceptibility to carbapenems was observed in Hospital Universitario 12 de Octubre (Madrid, Spain). To determine the possible role of metallo-ß-lactamases (MBLs) in the resistance profile of these isolates, a molecular and clinical epidemiological study was performed, including determination of patients' clinical characteristics, genetic diversity of strains, resistance mechanisms to carbapenems, and the genetic environment of VIM-1. A total of 73 E. cloacae isolates showed resistance to extended-spectrum cephalosporins and reduced susceptibility to at least one carbapenem during 2007-2012. PCR amplification revealed the presence of bla(VIM-1) gene in 37 isolates, bla(VIM-2) in 1 isolate and bla(KPC) in 5 isolates. Molecular typing showed high clonal diversity of E. cloacae isolates carrying bla(VIM-1). The genetic environment of bla(VIM-1) was investigated and two integron structures were found: intI-bla(VIM-1)-aacA4-dfrB1-aadA1-catB2-qacEΔ1/sul1 (In624); and intI-bla(VIM-1)-aacA4-aadA1-qacEΔ1/sul1 (In488). Isolates belonging to three clones (A, F and G) harboured different types of integron (In624 or In488) despite belonging to the same clone. Conjugal experiments showed an association with a conjugative plasmid of ca. 300 kb belonging to IncHI2 group, which is common in Spanish hospitals, suggesting that the widespread dissemination of bla(VIM-1) may be due to horizontal transfer of mobile genetic determinants rather than the result of spreading of a few clones. These results have implications for infection control programmes in the hospital.

Could polymerase chain reaction tests on conjunctival swabs be useful to diagnose herpetic keratitis?

INTRODUCTION: Diagnosis of HSV-1 keratitis (HK) is frequently based on clinical findings. Invasive specimens (corneal scrapings, biopsies) are required for microbiological diagnosis. Methods Corneal scrapings and conjunctival swabs were collected on patients with/without clinical suspicion of HK from 2007 to 2012.ResultsThe sensitivity, specificity, positive and negative predictive values for conjunctival swabs by PCR was 77.8, 92.1, 84.4 and 88.3, respectively. Discussion Conjunctival swabs by PCR may help in the diagnosis of HK, despite the limited sensitivity

INTRODUCCIÓN: El diagnóstico de queratitis herpética (QH) está basado normalmente en hallazgos clínicos. Para el diagnóstico microbiológico se requieren muestras invasivas (raspado corneal, biopsias).MÉTODOS: Raspados corneales y exudados conjuntivales fueron obtenidos de pacientes con/sin sospecha clínica de QH del ano˜ 2007 al 2012.RESULTADOS: La sensibilidad, la especificidad y los valores predictivos positivos y negativos para la PCR enexudados conjuntivales fueron 77,8, 92,1, 84,4 y 88,3, respectivamente. DISCUSIÓN: La PCR en exudados conjuntivales puede ayudar en el diagnóstico, a pesar de su limitada sensibilidad

Relationship between agr dysfunction and reduced vancomycin susceptibility in methicillin-susceptible Staphylococcus aureus causing bacteraemia.

OBJECTIVES: Limited data exist regarding the role of agr dysfunction in reducing susceptibility to vancomycin in methicillin-susceptible Staphylococcus aureus (MSSA). This study investigated the clinical and molecular epidemiology of MSSA causing bacteraemia, with emphasis on the reduced susceptibility to vancomycin (RSV) phenotype (MIC ≥ 1.5 mg/L) and its relationship with agr dysfunction. METHODS: All MSSA bloodstream isolates obtained at our hospital during 2010 were analysed. Antimicrobial susceptibility was determined and time-kill experiments were performed for oxacillin. Multilocus sequence type and agr genotype were determined and DNA microarray analysis of virulence factors was performed. agr dysfunction was assessed phenotypically and by RT-PCR quantification of RNAIII. RESULTS: Of 84 MSSA, 55 (65.5%) exhibited the RSV phenotype, comprising 13 clonal complexes. agr II polymorphism was more prevalent in RSV than non-RSV isolates (41.8% versus 17.2%, P = 0.023) and average levels of RNAIII gene expression were higher in RSV than non-RSV isolates (ΔCt 4.05 ± 3.29 versus 1.5 ± 2.11, P = 0.005), implying greater agr dysfunction in RSV MSSA. CONCLUSIONS: We demonstrated a correlation between RSV phenotype in MSSA and reduced agr expression, particularly in association with the agr II genotype. These results may help to understand the role of agr dysfunction in the increased mortality in MSSA infections.

Could polymerase chain reaction tests on conjunctival swabs be useful to diagnose herpetic keratitis?

INTRODUCTION: Diagnosis of HSV-1 keratitis (HK) is frequently based on clinical findings. Invasive specimens (corneal scrapings, biopsies) are required for microbiological diagnosis. METHODS: Corneal scrapings and conjunctival swabs were collected on patients with/without clinical suspicion of HK from 2007 to 2012. RESULTS: The sensitivity, specificity, positive and negative predictive values for conjunctival swabs by PCR was 77.8, 92.1, 84.4 and 88.3, respectively. DISCUSSION: Conjunctival swabs by PCR may help in the diagnosis of HK, despite the limited sensitivity.

Colonización nasal por Staphylococcus aureus en estudiantes de medicina: importancia en la transmisión hospitalaria / Staphylococcus aureus nasal colonization in medical students: Importance in nosocomial transmission

INTRODUCCIÓN: Staphylococcus aureus es el principal patógeno causante de infecciones nosocomiales. Los profesionales sanitarios, incluyendo los estudiantes de medicina, pueden ser una fuente de transmisión. Los objetivos del estudio fueron determinar la tasa de portadores nasales de S. aureus sensible y resistente a la meticilina (SARM) y evaluar el conocimiento y la adherencia que tenían los estudiantes sobre la higiene de manos. Métodos En el estudio participaron estudiantes de medicina adscritos al Hospital Universitario 12 de Octubre. Se realizó la toma de muestras de ambos vestíbulos nasales, y en todos los aislamientos se determinó la sensibilidad antibiótica. La obtención de datos se realizó mediante una encuesta autoadministrada que incluía factores de riesgo de colonización, hábitos higiénicos y conocimiento del protocolo de higiene de manos. Resultados De los 140 estudiantes incluidos, se detectaron 55 (39,3%) colonizados por S. aureus, con 3 (2,1%) SARM. La exposición a los antibióticos en los últimos 3 meses fue menor en los estudiantes colonizados (12,3% vs. 25,9%, p = 0,03). La autoevaluación mostró que el 56,4% de los estudiantes no se lavaban las manos casi nunca antes de atender al primer paciente, y solo el 38,6% se lavaban siempre después de explorar a los pacientes. El 35,7% desconocían el protocolo de higiene de manos, y el 38,6% no habían recibido formación específica. Conclusiones Los estudiantes de medicina deben ser tenidos en cuenta en los programas de control de la infección hospitalaria. La formación sobre higiene de manos debería impartirse antes de que los estudiantes comenzasen sus prácticas en el hospital

BACKGROUND: Staphylococcus aureus is the main pathogen causing nosocomial infections. Health profesionals, including medical students, could be a source of transmission. The aims of the study were to determine the rate of nasal carriage of S. aureus susceptible and resistant to methicillin (MRSA) and evaluate the knowledge and adherence that students had about hand hygiene. METHODS: The study included medical students attached to the Hospital Universitario 12 de Octubre(Madrid, Spain). We collected samples from both nasal vestibules, and the antimicrobial susceptibility was determined on all isolates. Data collection was performed using a self-administered questionnaire that included risk factors for colonization, hygiene habits and knowledge of hand hygiene protocols. RESULTS: Of the 140 students included, 55 (39.3%) were colonized by S. aureus, and 3 (2,1%) by MRSA. The exposure to antibiotics in the last 3 months was lower in colonized students (12.3% vs. 25.9%, P = .03). Self-assessment showed that 56.4% of students almost never washed their hands before to attending to the first patient, and only 38.6% always washed after examining patients. More than a third (35.7%) ignored the hand hygiene protocol, and 38.6% had not received specific formation. CONCLUSIONS: Medical students should be included in hospital infection control programs. Hand hygiene training should be given to students before they begin their practices in the hospital

Draft Whole-Genome Sequence of VIM-1-Producing Multidrug-Resistant Enterobacter cloacae EC_38VIM1.

The VIM-1-producing multidrug-resistant strain Enterobacter cloacae was isolated from blood culture. The strain showed multiple resistances to clinically used antibiotics, including all ß-lactams, fluoroquinolones, aminoglycosides, and sulfonamides. Sequence analysis showed the presence of 14 genes associated with resistance to antibiotics, including the metallo-ß-lactamase VIM-1 gene, which was located in a class 1 integron.

Draft Genome Sequence of Strain SA_ST125_MupR of Methicillin-Resistant Staphylococcus aureus ST125, a Major Clone in Spain.

Here, we report the draft genome sequence of a methicillin-resistant Staphylococcus aureus (MRSA) strain with high-level mupirocin resistance (SA_ST125_MupR), isolated from a patient with recurrent bacteremia. This strain belonged to sequence type ST125, which was responsible for more than 50% of the health care-associated infections caused by MRSA in Spain.

Draft Genome Sequence of VIM-2-Producing Multidrug-Resistant Pseudomonas aeruginosa ST175, an Epidemic High-Risk Clone.

The VIM-2-producing multidrug-resistant high-risk clone Pseudomonas aeruginosa sequence type (ST) 175 was isolated in the setting of a large outbreak in Hospital Universitario 12 de Octubre (Spain) from 2007 to 2010. This strain was resistant to all ß-lactams, fluoroquinolones, and aminoglycosides, with the exception of amikacin, and has become an endemic clone in our institution.

Molecular characterization of Achromobacter isolates from cystic fibrosis and non-cystic fibrosis patients in Madrid, Spain.

Multilocus sequence typing and nrdA sequence analysis identified 6 different species or genogroups and 13 sequence types (STs) among 15 Achromobacter isolates from cystic fibrosis (CF) patients and 7 species or genogroups and 11 STs among 11 isolates from non-CF patients. Achromobacter xylosoxidans was the most frequently isolated species among CF patients.

[Staphylococcus aureus nasal colonization in medical students: importance in nosocomial transmission]. / Colonización nasal por Staphylococcus aureus en estudiantes de medicina: importancia en la transmisión hospitalaria.

BACKGROUND: Staphylococcus aureus is the main pathogen causing nosocomial infections. Health professionals, including medical students, could be a source of transmission. The aims of the study were to determine the rate of nasal carriage of S.aureus susceptible and resistant to methicillin (MRSA) and evaluate the knowledge and adherence that students had about hand hygiene. METHODS: The study included medical students attached to the Hospital Universitario 12 de Octubre (Madrid, Spain). We collected samples from both nasal vestibules, and the antimicrobial susceptibility was determined on all isolates. Data collection was performed using a self-administered questionnaire that included risk factors for colonization, hygiene habits and knowledge of hand hygiene protocols. RESULTS: Of the 140 students included, 55 (39.3%) were colonized by S.aureus, and 3 (2,1%) by MRSA. The exposure to antibiotics in the last 3 months was lower in colonized students (12.3% vs. 25.9%, P=.03). Self-assessment showed that 56.4% of students almost never washed their hands before to attending to the first patient, and only 38.6% always washed after examining patients. More than a third (35.7%) ignored the hand hygiene protocol, and 38.6% had not received specific formation. CONCLUSIONS: Medical students should be included in hospital infection control programs. Hand hygiene training should be given to students before they begin their practices in the hospital.

VIM-2-producing multidrug-resistant Pseudomonas aeruginosa ST175 clone, Spain.

A total of 183 patients were colonized or infected with multidrug-resistant Pseudomonas aeruginosa isolates at a hospital in Spain during 2007-2010; prevalence increased over this period from 2.8% to 15.3%. To characterize these isolates, we performed molecular epidemiologic and drug resistance analysis. Genotyping showed that 104 (56.8%) isolates belonged to a single major clone (clone B), which was identified by multilocus sequence typing as sequence type (ST) 175. This clone was initially isolated from 5 patients in 2008, and then isolated from 23 patients in 2009 and 76 patients in 2010. PCR analysis of clone B isolates identified the bla(VIM-2) gene in all but 1 isolate, which harbored bla(IMP-22). ST175 isolates were susceptible to only amikacin (75%) and colistin (100%). Emergence of the ST175 clone represents a major health problem because it compromises therapy for treatment of P. aeruginosa nosocomial infections.

Long-term evolution of multiple outbreaks of Serratia marcescens bacteremia in a neonatal intensive care unit.

The annual incidence of Serratia marcescens bacteremia in a neonatal intensive care unit increased significantly between 2002 and 2010. Molecular epidemiology studies revealed that 8 clones were responsible for 85.2% of cases. Given that these infections are potentially preventable, even the appearance of 1 case of bacteremia should be an indicator for outbreak management.

Lack of the detection of XMRV or polytropic MLV-related sequences in blood cells from HIV-1-infected patients in Spain.

BACKGROUND: Xenotropic murine leukemia virus-related virus (XMRV) and polytropic murine leukemia virus (MLV)-related virus are recently described human gammaretroviruses that have been associated with prostate cancer and chronic fatigue syndrome. These studies have been controversial because a number of laboratories have been unable to find evidence of XMRV in similar groups of patients or controls. Because the existence of XMRV raises many questions, we decided to study its presence in a group of patients infected with HIV-1 with a high proportion of intravenous drug use and coinfection by hepatitis C virus. METHODS: Forty HIV-1-infected patients under follow-up in our institution were screened for XMRV/MLV by nested polymerase chain reaction using primers targeting the gag and env region. Specific primers for mouse mitochondrial DNA were used to rule out contamination. RESULTS: No evidence of XMRV or polytropic MLV-related sequences was found in any sample from patients or controls. Four samples yielded polymerase chain reaction bands whose sequence corresponded to murine endogenous retroviral sequences, however, contamination with mouse cell DNA was subsequently confirmed. CONCLUSIONS: XMRV/MLV viruses do not seem to be associated with HIV-1 infection or intravenous drug use. Contamination of samples or reagents by genomic murine DNA or XMRV vectors could account for the sporadic detection of positive samples for XMRV and related agents.

Multidrug-resistant Acinetobacter baumannii Harboring OXA-24 carbapenemase, Spain.

In February 2006, a patient colonized with a multidrug-resistant sequence type 56 Acinetobacter baumannii strain was admitted to a hospital in Madrid, Spain. This strain spread rapidly and caused a large outbreak in the hospital. Clinicians should be alert for this strain because its spread would have serious health consequences.

Endoluminal colonization as a risk factor for coagulase-negative staphylococcal catheter-related bloodstream infections in haemodialysis patients.

BACKGROUND: Approximately 25% of haemodialysis (HD) patients use catheters as vascular access. Catheter-related bloodstream infections (CRBSI) are a major risk in this population. The objective of our study was to determine whether endoluminal catheter colonization (ECC) predicts CRBSI. METHODS: We followed up a cohort of HD patients in our institution who underwent HD with tunnelled cuffed central venous catheters (TCC) between December 2006 and June 2008. Colonization of the inner catheter lumen was assessed every 15 days immediately before HD by culture of blood-heparin mixture and the time to positivity (TTP) was recorded by the BacT/Alert automated system. CRBSI was confirmed by differential TTP (> 2 h) between TCC and peripheral blood cultures. RESULTS: We studied 51 patients who required 64 TCC. The incidence of CRBSI was 1.65 episodes per 1000 catheter-days, with Staphylococcus epidermidis being the most common cause of infection (76.2%). ECC was more frequent in the CRSBI group than in the non-CRBSI group (100 vs 5.4%, P < 0.001). For S. epidermidis CRBSIs, the median time from ECC to CRBSI was 31.5 days (interquartile range, 27.0-79.0). The sensitivity, specificity and negative and positive predictive values of arterial lumen cultures for S. epidermidis CRBSIs were 100, 96.3, 92.3 and 100%, respectively, while for venous culture, these values were 92.3, 96.3, 92.3 and 96.3%, respectively. For predicting S. epidermidis CRBSI, endoluminal cultures with a TTP of ≤ 14 h had sensitivity and specificity of 52.1 and 97.7%, respectively. CONCLUSIONS: This study shows that ECC may predict the risk of developing CRSBI. Surveillance cultures could, therefore, be used to triage individual HD patients who might benefit from specific intervention measures.

Acute graft pyelonephritis in renal transplant recipients: incidence, risk factors and long-term outcome.

BACKGROUND: The influence of acute graft pyelonephritis (AGPN) on graft outcome in renal transplant recipients still remains controversial. METHODS: We retrospectively analysed 189 patients (113 males; mean age: 49.7 ± 13.1 years) undergoing renal transplantation at the University Hospital 12 de Octubre (Madrid, Spain) from January 2002 to December 2004, with a minimum follow-up of 36 months. Factors associated with AGPN were assessed by logistic regression analysis. Long-term graft function was compared according to the occurrence of this complication during follow-up. 'Decline in renal graft function' was defined as the increase in serum creatinine (SC) levels > 0.33 mg/dL between Month 3 and Year 1 after transplantation. RESULTS: Nineteen patients (10.0%) were diagnosed with 25 episodes of AGPN (incidence rate: 4.4 episodes per 100 patient-years). The presence of glomerulonephritis as the underlying disease [odds ratio (OR) 4.2; 95% confidence interval (95%CI): 1.3-14.1] and the previous occurrence of two to five (OR 9.4; 95%CI: 1.5-56.8) or more than five episodes of asymptomatic bacteriuria after transplantation (OR 19.8; 95%CI: 2.4-160.2) emerged as independent predictors for AGPN. A near-significant association was found for cytomegalovirus infection (OR 4.2; 95%CI: 0.9-18.4), whereas receiving a single-kidney transplant (vs. double-kidney) showed a protective effect (OR 0.2; 95%CI: 0.0-0.8). During the 36-month follow-up, levels of SC, creatinine clearance and 24-h proteinuria did not differ significantly between patients with or without AGPN, and this complication did not exert any effect on the risk for decline in renal graft function. CONCLUSIONS: AGPN does not impair long-term graft function in renal transplant recipients.

Systematic screening and treatment of asymptomatic bacteriuria in renal transplant recipients.

We sought to examine the impact of asymptomatic bacteriuria on renal transplant outcome by retrospectively analyzing 189 renal transplant recipients for whom systematic screening uncovered 298 episodes of asymptomatic bacteriuria in 96 recipients. These patients were treated and all were followed for 36 months. Significant risk factors included female gender, glomerulonephritis as the disease that led to transplantation, and double renal transplant. There were no differences in serum creatinine, creatinine clearance, or proteinuria between patients with and without bacteriuria. The incidence of pyelonephritis in these patients was 7.6 episodes per 100 patient-years compared with 1.07 in those without asymptomatic bacteriuria. Between two to five and more than five bacteriuria episodes were significant independent factors associated with pyelonephritis whereas more than five episodes was a significant independent factor associated with rejection. Thus, we found no differences in renal function prognosis between patients who do not develop asymptomatic bacteriuria and those uncovered by systematic screening and who received treatment following kidney transplantation. Despite this treatment, the incidence of pyelonephritis was much higher in the group of patients with detected asymptomatic bacteriuria.

Clinical and molecular characteristics of infections with CO2-dependent small-colony variants of Staphylococcus aureus.

Most Staphylococcus aureus small-colony variants (SCVs) are auxotrophs for menadione, hemin, or thymidine but rarely for CO(2). We conducted a prospective investigation of all clinical cases of CO(2)-dependent S. aureus during a 3-year period. We found 14 CO(2)-dependent isolates of S. aureus from 14 patients that fulfilled all requirements to be considered SCVs, 9 of which were methicillin resistant. The clinical presentations included four cases of catheter-related bacteremia, one complicated by endocarditis; two deep infections (mediastinitis and spondylodiscitis); four wound infections; two respiratory infections; and two cases of nasal colonization. Pulsed-field gel electrophoresis typing showed that the 14 isolates were distributed into 4 types corresponding to sequence types ST125-agr group II (agrII), ST30-agrIII, ST34-agrIII, and ST45-agrI. An array hybridization technique performed on the 14 CO(2)-dependent isolates and 20 S. aureus isolates with normal phenotype and representing the same sequence types showed that all possessed the enterotoxin gene cluster egc, as well as the genes for alpha-hemolysin and delta-hemolysin; biofilm genes icaA, icaC, and icaD; several microbial surface components recognizing adhesive matrix molecules (MSCRAMM) genes (clfA, clfB, ebh, eno, fib, ebpS, sdrC, and vw); and the isaB gene. Our study confirms the importance of CO(2)-dependent SCVs of S. aureus as significant pathogens. Clinical microbiologists should be aware of this kind of auxotrophy because recovery and identification are challenging and not routine. Further studies are necessary to determine the incidence of CO(2) auxotrophs of S. aureus, the factors that select these strains in the host, and the genetic basis of this type of auxotrophy.

Detection of HIV-1 at between 20 and 49 copies per milliliter by the Cobas TaqMan HIV-1 v2.0 assay is associated with higher pretherapy viral load and less time on antiretroviral therapy.

New commercial techniques for determination of the viral load (VL) in plasma are able to detect as few as 20 copies of HIV-1 RNA/ml. The relevance of this new technical threshold is uncertain. Upon multivariate analysis, factors associated with detection of VLs between 20 and 49 copies/ml by the Cobas TaqMan HIV-1 v2.0 assay in an HIV clinic were the basal VL and time on antiretroviral therapy.