Persistence of Zika virus in semen 93 days after the onset of symptoms

INTRODUCTION: Zika virus is mainly transmitted through the bites of infected Aedes mosquitoes, although mother-to-child and sexual transmission have also been described. The presence of Zika virus in semen after infection seems to be not uncommon, but the duration of viral persistence has not been well-determined. METHODS: Molecular, serological and cell culture methods were used for the diagnosis and follow up of a case of Zika virus infection imported from Venezuela. Serial samples of serum, urine and semen were analyzed to investigate the persistence of the Zika virus. RESULTS: Zika virus was detected in semen samples up to 93 days after the onset of symptoms. CONCLUSIONS: Our results confirm the persistence of Zika virus in semen samples for long periods after infection

Evaluation of a novel microfluidic immuno-magnetic agglutination assay method for detection of dengue virus NS1 antigen

BACKGROUND: Dengue virus (DENV) is the most important arbovirus worldwide, causing infections in endemic countries and returning travellers from these areas. Rapid diagnostic tests are needed to improve patient management and monitor local transmission. The detection of DENV non-structural protein 1 (NS1) is a useful tool for the diagnosis, but the currently available methods can be time consuming or lack sensitivity. The objective of our study was to evaluate a new rapid and semi-quantitative microfluidic DENV NS1 immuno-magnetic agglutination assay based on aggregation of magnetic nanoparticles detected by an electronic reader (Virotrack Dengue Acute and Blubox, Blusense diagnostics, Copenhagen, Denmark). METHODOLOGY/PRINCIPAL FINDINGS: A panel of 135 serum samples from travelers returning from dengue endemic countries was analyzed (74 DENV positive samples including the four DENV serotypes, 26 Zika virus positive samples, 25 chikungunya virus positive samples, 5 malaria positive samples and 5 negative samples). Samples were tested by three different antigen detection methods: SD Dengue NS1 Ag ELISA, SD BIOLINE Dengue Duo and ViroTrack Dengue Acute. The sensitivity observed for SD Dengue NS1 Ag ELISA, ViroTrack Dengue Acute and SD BIOLINE Dengue Duo was 97.2%, 91.1% and 68.1%, respectively. All methods showed high specificity (98.4% for ViroTrack Dengue Acute and 100% for both SD Dengue NS1 Ag ELISA and SD BIOLINE Dengue Duo). SD Dengue NS1 Ag ELISA and ViroTrack Dengue Acute only failed to detect samples positive for DENV-2. CONCLUSIONS/SIGNIFICANCE: ViroTrack Dengue Acute is a sensitive and specific assay for DENV NS1 detection. It provides faster results than the ELISA method and a better performance than the rapid immunochromatographic tests. ViroTrack Dengue Acute could represent a valuable tool for rapid diagnosis of DENV infections in returning travellers from endemic countries

In vitro activity of 12 antimicrobial peptides against Mycobacterium tuberculosis and Mycobacterium avium clinical isolates

Tuberculosis (TB) remains a major threat to human health worldwide. The increasing incidence of non-tuberculous mycobacterial infections and particularly those produced by Mycobacterium avium has emphasized the need to develop new drugs. Additionally, high levels of natural drug resistance in non-tuberculous mycobacteria (NTM) and the emergence of multidrug-resistant (MDR) TB is of great concern. Antimicrobial peptides (AMPs) are antibiotics with broad-spectrum antimicrobial activity. The objective was to assess the activity of AMPs against Mycobacterium tuberculosis and M. avium clinical isolates. MICs were determined using microtitre plates and the resazurin assay. Mastoparan and melittin showed the greatest activity against M. tuberculosis, while indolicidin had the lowest MIC against M. avium. In conclusion, AMPs could be alternatives for the treatment of mycobacterial infections. Further investigation of AMPs' activity in combination and associated with conventional antibiotics and their loading into drug-delivery systems could lead to their use in clinical practice

Cutaneous infection by Phaeoacremonium parasiticum

Background: Phaeoacremonium parasiticum is considered a rare infectious agent that is part of a heterogeneous group of fungi causing phaeohyphomycosis. This organism is capable of producing subcutaneous infections, eumycetomas, osteomyelitis, arthritis, myositis and also disseminated diseases, such as fungemia and endocarditis. Case report: We describe a case of cutaneous infection by P. parasiticum in a kidney transplant patient. The identification of this microorganism was performed by microbiological and histopathological studies and confirmed with the sequence of the gene encoding β-tubulin and a real time panfungal PCR targeting 18S ribosomal RNA gene. The microorganism was correctly identified by phenotypic and molecular methods. The patient was treated with oral antifungal therapy and a debulking surgery and evolved without any complication. Conclusions: The diagnosis of this infection is difficult and usually affects kidney transplant patients, but the reasons of this association are still unknown

Evaluation of a multiplex panel for the diagnosis of acute infectious diarrhea in immunocompromised hematologic patients

Introduction: diarrhea is a frequent complication in hematologic patients, being an infectious cause frequently suspected. Rapid and accurate detection of gastrointestinal pathogens is vital in immunocompromised hosts. The aim of this study was to compare routine diagnostic methods versus a multiplex polymerase chain reaction (PCR) assay for the diagnosis of infectious diarrhea in immunocompromised hematologic patients. Material and methods: we conducted a prospective observational study from March 2015 to January 2016 to compare conventional methods for the diagnosis of infectious diarrhea with FIlmArray GI Panel (BioFire-bioMérieux, France). Samples from adult immunocompromised hematologic patients with acute diarrhea were collected. In cases with discordant results, a second multiplex assay was performed (Allplex, Seegene, Korea). The result was considered positive or negative when the same result was obtained by at least two of the methods. Results: a total of 95 samples were obtained from 95 patients (median age of 52 years (46-64)). Sixty-one (64%) episodes were hospital-acquired and 34 (36%) were community-acquired diarrhea. Twenty-five (26%) patients had a positive microbiological result, being Clostridium difficile the most frequent pathogen, followed by Campylobacter spp and norovirus. The concordance between FilmArray methods was good (k = 0.79). The FilmArray GI panel showed a sensitivity of 95%, a specificity of 100% for positive results. The time required to obtain results was markedly reduced with the use of multiplex PCR methods. Conclusions: multiplex molecular panels provide a rapid and sensitive tool for the diagnosis of infectious diarrhea, thereby allowing more timely clinical decisions in immunocompromised hematologic patients

Screening for Zika virus infection in 1057 potentially exposed pregnant women, Catalonia (northeastern Spain)

Dear editor: A recent editorial in Travel Medicine and Infectious Diseases highlighted the lack of studies about Zika virus (ZIKV) in pregnancy and its implications in many countries [1]. Zika virus infection can induce congenital defects in the newborn such as microcephaly and miscarriage when mothers are infected during pregnancy [2]. However, relevant questions remain to be completely understood, such as the risk of infection for pregnant women and of subsequent congenital defects, and the ratio between symptomatic and asymptomatic ZIKV infections in the general population and in pregnant women. Here, we describe the results of a ZIKV screening of pregnant women in Catalonia, northeastern Spain. Testing for ZIKV was recommended for all pregnant women with history of travel to ZIKV endemic areas during pregnancy or in the 8 weeks before conception [3]. Symptomatic patients were screened by serological methods from day four after the onset of symptoms and by molecular methods within the first week (serum) and two weeks (urine) after illness. Asymptomatic patients were tested by serological methods. Seroneutralization assay for ZIKV was perfomed in samples positive for antibodies. Commercial diagnostic assays were used (RT-PCR, Altona Diagnostics and IIFT, Euroimmun). Neutralization titers ≥1/32 were considered indicative of the presence of ZIKV neutralizing antibodies. Follow up at two designated reference obstetrical departments for early detection of microcephaly or other malformations was offered to pregnant women with laboratory evidence of ZIKV infection. When available, amniotic fluid and placental tissue samples were tested for ZIKV by RT-PCR in cases of microcephaly or miscarriage, respectively

Time-to-positivity, type of culture media and oxidase test performed on positive blood culture vials to predict Pseudomonas aeruginosa in patients with Gram-negative bacilli bacteraemia

OBJECTIVE: The aim of this study was to determine the usefulness of oxidase test and time-to-positivity (TTP) in aerobic and anaerobic blood culture vials to detect the presence of Pseudomonas aeruginosa in patients with Gram-negative bacilli (GNB) bacteraemia. METHODS: TTP was recorded for each aerobic and anaerobic blood culture vial of monomicrobial bacteraemia due to GNB. Oxidase test was performed in a pellet of the centrifuged content of the positive blood culture. An algorithm was developed in order to perform the oxidase test efficiently taking into account TTP and type of vial. RESULTS: A total of 341 episodes of GNB bacteraemia were analysed. Sensitivity, specificity, positive predictive value and negative predictive value of the oxidase test performed on positive vials with GNB to predict P. aeruginosa were 95%, 99%, 91%, and 99%, respectively. When growth was first or exclusively detected in anaerobic vials, P. aeruginosa was never identified hence the performance of the oxidase test could be avoided. When growth was only or first detected in aerobic vials, a TTP≥8h predicted P. aeruginosa in 37% or cases (63 of 169), therefore oxidase test is highly recommended. CONCLUSIONS: Oxidase test performed onto positive blood culture vials previously selected by TTP and type of vials is an easy and inexpensive way to predict P. aeruginosa. In most cases, this can lead to optimization of treatment in less than 24 hours

Genomics And Susceptibility Profiles Of Extensively Drug-resistant Pseudomonas Aeruginosa Isolates From Spain

This study assessed the molecular epidemiology, resistance mechanisms, and susceptibility profiles of a collection of 150 extensively drug-resistant (XDR) Pseudomonas aeruginosa clinical isolates obtained from a 2015 Spanish multicenter study, with a particular focus on resistome analysis in relation to ceftolozane-tazobactam susceptibility. Broth microdilution MICs revealed that nearly all (> 95%) of the isolates were nonsusceptible to piperacillin-tazobactam, ceftazidime, cefepime, aztreonam, imipenem, meropenem, and ciprofloxacin. Most of them were also resistant to tobramycin (77%), whereas nonsusceptibility rates were lower for ceftolozane-tazobactam (31%), amikacin (7%), and colistin (2%). Pulsed-field gel electrophoresis-multilocus sequence typing (PFGE-MLST) analysis revealed that nearly all of the isolates belonged to previously described high-risk clones. Sequence type 175 (ST175) was detected in all 9 participating hospitals and accounted for 68% (n = 101) of the XDR isolates, distantly followed by ST244 (n = 16), ST253 (n = 12), ST235 (n = 8), and ST111 (n = 2), which were detected only in 1 to 2 hospitals. Through phenotypic and molecular methods, the presence of horizontally acquired carbapenemases was detected in 21% of the isolates, mostly VIM (17%) and GES enzymes (4%). At least two representative isolates from each clone and hospital (n = 44) were fully sequenced on an illumina MiSeq. Classical mutational mechanisms, such as those leading to the overexpression of the beta-lactamase AmpC or efflux pumps, OprD inactivation, and/or quinolone resistance-determining regions (QRDR) mutations, were confirmed in most isolates and correlated well with the resistance phenotypes in the absence of horizontally acquired determinants. Ceftolozane-tazobactam resistance was not detected in carbapenemase-negative isolates, in agreement with sequencing data showing the absence of ampC mutations. The unique set of mutations responsible for the XDR phenotype of ST175 clone documented 7 years earlier were found to be conserved, denoting the long-term persistence of this specific XDR lineage in Spanish hospitals. Finally, other potentially relevant mutations were evidenced, including those in penicillin-binding protein 3 (PBP3), which is involved in beta-lactam (including ceftolozane-tazobactam) resistance, and FusA1, which is linked to aminoglycoside resistance

Molecular characterization of imported and autochthonous dengue in northeastern spain

Dengue is the most significant arbovirus worldwide and a public health threat to nonendemic areas in which Aedes vectors are present. Autochthonous dengue transmission has been reported in several European countries in the last decade. Infected travelers from endemic regions arriving to areas colonized by Aedes albopictus in Europe need to be monitored in surveillance and control programs. We aimed to perform molecular characterization of RT-PCR-positive dengue cases detected in Catalonia, northeastern Spain, from 2013 to 2018. The basic demographic information and the geographical regions of importation were also analyzed. One-hundred four dengue cases were studied (103 imported infections and the first autochthonous case in our region). The dengue virus strains detected were serotyped and genotyped using molecular methods, and phylogenetic analyses were conducted. All four dengue serotypes were detected in travelers, including up to 10 different genotypes, reflecting the global circulation of dengue in endemic areas. The primary travel-related case of the 2018 autochthonous transmission was not identified, but the molecular analysis revealed dengue serotype 1, genotype I of Asian origin. Our results highlight the diversity of imported dengue virus strains and the role of molecular epidemiology in supporting arbovirus surveillance programs

Diagnóstico y vigilancia de las infecciones por arbovirus

[spa] INTRODUCCIÓN: Los arbovirus son un grupo heterogéneo de virus transmitidos por vectores artrópodos. Entre ellos destacan los virus dengue (VDEN), chikungunya (VCHIK) y Zika (VZIK), que presentan similitudes en cuanto a su distribución geográfica, vectores transmisores y cuadro clínico que ocasionan. Existe un alto riesgo de emergencia y re-emergencia de los mismos debido, entre otros, al gran número de viajeros entre zonas endémicas y no endémicas y a la expansión de diferentes vectores. Unas técnicas diagnósticas fiables y rápidas son de vital importancia para hacer frente a este riesgo. HIPÓTESIS Y OBJETIVOS: Una evaluación rigurosa de diferentes métodos diagnósticos y de la cinética de los marcadores de infección permitirá la creación de algoritmos diagnósticos que mejoren el diagnóstico de las arbovirosis. La revisión de los resultados obtenidos por los protocolos vigentes permitirá estimar el riesgo de infección y afectación fetal por VZIK en embarazadas. Los objetivos de este trabajo son: estudiar la cinética de la infección por VZIK, evaluar el cribado serológico de VZIK llevado a cabo en embarazadas en Cataluña, analizar la utilidad de los parámetros analíticos para orientar el diagnóstico del síndrome febril en viajeros, evaluar el rendimiento de un nuevo ensayo de inmuno-aglutinación magnética (IMA) para detectar antígeno NS1 de VDEN e investigar la utilidad de otras técnicas serológicas para el diagnóstico de las infecciones por VDEN, VCHIK y VZIK. MATERIAL Y MÉTODOS : Se estudió la presencia de VZIK en muestras de suero y orina de 24 pacientes con infección por VZIK mediante RT-PCR en tiempo real. Se investigó la presencia de VZIK en semen en un paciente mediante métodos moleculares y cultivo celular utilizando muestras seriadas. Se evaluaron los resultados del cribado de VZIK y el impacto sobre el embarazo en 1.057 gestantes cribadas para VZIK en Cataluña en 2016. Se estudiaron los valores de plaquetas y leucocitos en 1.218 viajeros provenientes del trópico para determinar la utilidad de la plaquetopenia y leucopenia en el diagnóstico de malaria y arbovirosis. Se estudió el rendimiento diagnóstico de un método de IMA para detectar antígeno NS1 de VDEN en un panel de 135 muestras que incluía los cuatro serotipos de VDEN, y se comparó con las técnicas de ELISA e inmunocromatografía (ICT). También se exploró la utilidad diagnóstica de un test rápido de dengue, de un ELISA basado en la proteína NS1 de VZIK para detectar anticuerpos y de un ensayo de inmunoblot para diferenciar infecciones por VDEN, VZIK y VCHIK, comparando los resultados con los de las técnicas diagnósticas de rutina. RESULTADOS PRINCIPALES : Se detectó la presencia del VZIK por RT-PCR en tiempo real en muestras de suero y orina fuera de los períodos considerados óptimos en aquel momento por las recomendaciones de organismos internacionales. Se detectó la presencia de ARN de VZIK en semen 93 días desde el inicio de los síntomas, sin poder aislar el virus en cultivo celular. Los datos del cribado serológico de VZIK de 1.057 embarazadas en Cataluña mostraron un 13,4% de gestantes con alguna evidencia serológica de infección por VZIK, siendo la prevalencia de efectos fetales adversos (aborto o microcefalia) del 3,1%. La leucopenia (<4x109 leucocitos/L) y la trombocitopenia (<150x109 plaquetas/L) mostraron un valor predictivo negativo del 98,1% para el diagnóstico de arbovirosis y malaria respectivamente, en viajeros con síndrome febril a la vuelta de un viaje al trópico. El ensayo de IMA para detectar NS1 de VDEN mostró una elevada sensibilidad (91,9%) y especificidad (98,4%), comparables a la técnica de ELISA pero obteniendo los resultados de forma tan rápida como una ICT. El test rápido para VDEN basado en ICT mostró un la detección de IgM frente al virus. La técnica de ELISA para la detección de IgG utilizando como antígeno NS1 de VZIK mostró una sensibilidad del 92,9% y una especificidad del 87,0% comparándola con el ensayo de microneutralización. El ensayo basado en inmunoblot mostró una baja sensibilidad (<40%) para la detección de IgM frente a VDEN, VZIK y VCHIK. CONCLUSIONES: El testado simultáneo de suero y orina mediante métodos moleculares aumenta el rendimiento diagnóstico para la detección de infecciones por VZIK. El VZIK puede persistir en semen durante periodos prolongados de tiempo. La frecuencia de desenlaces adversos por VZIK en embarazadas atendidas en Cataluña fue similar a la descrita en la literatura en algunas zonas endémicas. Los valores de leucocitos y plaquetas pueden ayudar en el despistaje clínico inicial de pacientes con fiebre a la vuelta de un viaje al trópico. El ensayo de IMA es una herramienta útil para el diagnóstico rápido de dengue agudo. La evaluación de nuevas técnicas diagnósticas para arbovirosis es necesaria para determinar su utilidad a fin de intentar solucionar las limitaciones del diagnóstico serológico