RESUMEN
The molecular detection of Toxoplasma gondii DNA is a key tool for the diagnosis of disseminated and congenital toxoplasmosis. This multicentric study from the Molecular Biology Pole of the French National Reference Center for toxoplasmosis aimed to evaluate Toxoplasma gondii Real-TM PCR kit (Sacace). The study compared the analytical and clinical performances of this PCR assay with the reference PCRs used in proficient laboratories. PCR efficiencies varied from 90% to 112%; linearity zone extended over four log units (R2 > 0.99) and limit of detection varied from 0.01 to ≤1 Tg/mL depending on the center. Determined on 173 cryopreserved DNAs from a large range of clinical specimens, clinical sensitivity was 100% [106/106; 95 confidence interval (CI): 96.5%-100%] and specificity was 100% (67/67; 95 CI: 94.6%-100%). The study revealed two potential limitations of the Sacace PCR assay: the first was the inconsistency of the internal control (IC) when added to the PCR mixture. This point was not found under routine conditions when the IC was added during the extraction step. The second is a lack of practicality, as the mixture is distributed over several vials, requiring numerous pipetting operations. Overall, this study provides useful information for the molecular diagnosis of toxoplasmosis; the analytical and clinical performances of the Sacace PCR kit were satisfactory, the kit having sensitivity and specificity similar to those of expert center methods and being able to detect low parasite loads, at levels where multiplicative analysis gives inconsistently positive results. Finally, the study recommends multiplicative analysis in particular for amniotic fluids, aqueous humor, and other single specimens.
Asunto(s)
Toxoplasma , Toxoplasmosis Congénita , Toxoplasmosis , Humanos , Toxoplasma/genética , Toxoplasmosis/diagnóstico , Toxoplasmosis/parasitología , Toxoplasmosis Congénita/diagnóstico , Toxoplasmosis Congénita/parasitología , ADN , Juego de Reactivos para Diagnóstico , Sensibilidad y Especificidad , ADN Protozoario/genética , ADN Protozoario/análisisRESUMEN
Pneumocystis pneumonia (PcP) remains life-threatening in kidney transplant recipients (KTR). Our study investigated risk factors one-year before PcP. We conducted a monocentric, case-control study including all KTR at the Dijon University Hospital (France) with a diagnosis of PcP between 2005 and 2022 (cases), and matched control KTR with no history of PcP (3 controls/case). Among all 1,135 KTR, 57 cases (5%) and 169 matched-controls were included. PcP was associated with 18% mortality. Compared to controls, cases were older, with a higher immunological risk, and CMV infection was more frequent in the year preceding the occurrence of PcP (23% vs. 4%; p < 0.001). As early as 1 year before PcP, lymphocyte counts were lower and serum creatinine levels were higher in cases, but immunosuppressive regimens were not significantly different. Multivariable analysis identified lymphocyte count, serum creatinine level, being treated by immunosuppressive therapy other than anti-rejection drugs, and CMV infection in the year preceding the time PcP as independently associated with the occurrence of PcP. PcP was associated with an increased risk of subsequent chronic rejection (27% vs. 3%; p = 0.001) and return to dialysis (20% vs. 3%; p = 0.002). The occurrence of CMV infection and a low lymphocyte count could redefine the indications for continuation or reinitiation of anti-Pneumocystis prophylaxis.
Asunto(s)
Infecciones por Citomegalovirus , Trasplante de Riñón , Linfopenia , Pneumocystis carinii , Neumonía por Pneumocystis , Trombocitopenia , Humanos , Neumonía por Pneumocystis/diagnóstico , Neumonía por Pneumocystis/complicaciones , Neumonía por Pneumocystis/tratamiento farmacológico , Estudios de Casos y Controles , Trasplante de Riñón/efectos adversos , Creatinina , Infecciones por Citomegalovirus/complicaciones , Infecciones por Citomegalovirus/diagnóstico , Infecciones por Citomegalovirus/epidemiología , Factores de Riesgo , Linfopenia/complicaciones , Trombocitopenia/complicaciones , Receptores de Trasplantes , Estudios RetrospectivosRESUMEN
BACKGROUND: Secretory IgA interacts with commensal bacteria, but its impact on human mycobiota ecology has not been widely explored. In particular, whether human IgA-deficiency is associated with gut fungal dysbiosis remains unknown. OBJECTIVES: Our goal was to study the impact of IgA on gut mycobiota ecology. METHODS: The Fungi-Flow method was used to characterize fecal, systemic, and maternal IgA, IgM, and IgG responses against 14 representative fungal strains (yeast/spores or hyphae forms) in healthy donors (HDs) (n = 34, 31, and 20, respectively) and to also compare gut mycobiota opsonization by secretory antibodies in HDs (n = 28) and patients with selective IgA deficiency (SIgAd) (n = 12). Stool mycobiota composition was determined by internal transcribed spacer gene sequencing in HDs (n = 23) and patients with SIgAd (n = 17). Circulating CD4+ T-cell cytokine secretion profiles were determined by intracellular staining. The impact of secretory IgA, purified from breast milk (n = 9), on Candidaalbicans growth and intestinal Caco-2 cell invasion was tested in vitro. RESULTS: Homeostatic IgA binds commensal fungi with a body fluid-selective pattern of recognition. In patients with SIgAd, fungal gut ecology is preserved by compensatory IgM binding to commensal fungi. Gut Calbicans overgrowth nevertheless occurs in this condition but only in clinically symptomatic patients with decreased TH17/TH22 T-cell responses. Indeed, secretory IgA can reduce in vitro budding and invasion of intestinal cells by Calbicans and therefore exert control on this pathobiont. CONCLUSION: IgA has a selective impact on Calbicans ecology to preserve fungal-host mutualism.
Asunto(s)
Candida albicans , Deficiencia de IgA , Femenino , Humanos , Células CACO-2 , Inmunoglobulina A , Inmunoglobulina A Secretora , Inmunoglobulina MRESUMEN
BACKGROUND: Early diagnosis and prompt initiation of specific antifungal treatment are essential for improving the prognosis of mucormycosis. We aimed to assess the performance of serum Mucorales quantitative polymerase chain reaction (qPCR) for the early diagnosis and follow-up of mucormycosis. METHODS: We prospectively enrolled 232 patients with suspicion of invasive mold disease, evaluated using standard imaging and mycological procedures. Thirteen additional patients with proven or probable mucormycosis were included to analyze DNA load kinetics. Serum samples were collected twice-a-week for Mucorales qPCR tests targeting the Mucorales genera Lichtheimia, Rhizomucor, and Mucor/Rhizopus. RESULTS: The sensitivity was 85.2%, specificity 89.8%, and positive and negative likelihood ratios 8.3 and 0.17, respectively in this prospective study. The first Mucorales qPCR-positive serum was observed a median of 4 days (interquartile range [IQR], 0-9) before sampling of the first mycological or histological positive specimen and a median of one day (IQR, -2 to 6) before the first imaging was performed. Negativity of Mucorales qPCR within seven days after liposomal-amphotericin B initiation was associated with an 85% lower 30-day mortality rate (adjusted hazard ratioâ =â 0·15, 95% confidence interval [.03-.73], Pâ =â .02). CONCLUSIONS: Our study argues for the inclusion of qPCR for the detection of circulating Mucorales DNA for mucormycosis diagnosis and follow-up after treatment initiation. Positive results should be added to the criteria for the consensual definitions from the European Organization for the Research and Treatment of Cancer/Mycoses Study Group Education and Research Consortium (EORTC/MSGERC), as already done for Aspergillus PCR.
Asunto(s)
Mucorales , Mucormicosis , Anfotericina B , Antifúngicos/uso terapéutico , Detección Precoz del Cáncer , Humanos , Mucorales/genética , Mucormicosis/diagnóstico , Reacción en Cadena de la Polimerasa/métodos , Estudios ProspectivosRESUMEN
In the non-AIDS group, several underlying conditions and immune defects could lead to different PCP presentations. This study compared PCP presentation and outcome according to the underlying disease. A secondary analysis of a previously published prospective observational study including 544 PCP patients was done. Only non-AIDS patients were included. Underlying disease was defined as chronic lymphocytic leukemia (CLL), organ transplantation, solid cancer, allogeneic hematopoietic stem cell transplant (AHSCT), other hematological diseases, and immunosuppressive treatment. Clinical characteristics and outcomes were compared between groups. Multiple correspondent analyses compared clinical characteristics at diagnosis. Day 30 mortality was analyzed. Three hundred and twenty-one patients were included in the study. The underlying diseases were hematological malignancy (n = 75), AHSCT (n = 14), CLL (n = 19), solid organ transplant (n = 94), solid tumor (n = 39), and immunosuppressive treatment (n = 57). Compared with other underlying diseases, PCP related to CLL was closer to PCP related to AIDS presentation (long duration of symptoms before diagnosis, high level of dyspnea, and low oxygen saturation at diagnosis). Day 30 mortality was associated with underlying disease, oxygen flow, and shock at ICU admission. PCP presentations may vary according to the underlying reason for immunosuppression. Response to treatment and adjuvant steroid therapy should be analyzed regarding this result.
Asunto(s)
Neumonía por Pneumocystis/complicaciones , Insuficiencia Respiratoria/etiología , Insuficiencia Respiratoria/mortalidad , Enfermedad Aguda , Anciano , Femenino , Enfermedades Hematológicas/complicaciones , Humanos , Leucemia Linfoide/complicaciones , Masculino , Persona de Mediana Edad , Neumonía por Pneumocystis/diagnóstico , Estudios ProspectivosRESUMEN
Tight junctions play a major role in maintaining the integrity and impermeability of the intestinal barrier. As such, they act as an ideal target for pathogens to promote their translocation through the intestinal mucosa and invade their host. Different strategies are used by pathogens, aimed at directly destabilizing the junctional network or modulating the different signaling pathways involved in the modulation of these junctions. After a brief presentation of the organization and modulation of tight junctions, we provide the state of the art of the molecular mechanisms leading to permeability breakdown of the gut barrier as a consequence of tight junctions' attack by pathogens, including bacteria, viruses, fungi, and parasites.
Asunto(s)
Bacterias/patogenicidad , Células Epiteliales/fisiología , Infecciones/fisiopatología , Enfermedades Intestinales/fisiopatología , Mucosa Intestinal/fisiología , Uniones Estrechas/fisiología , Animales , Permeabilidad de la Membrana Celular , Humanos , Infecciones/microbiología , Enfermedades Intestinales/microbiología , Transducción de SeñalRESUMEN
The burden of nosocomial Pneumocystis infections in transplantation units in France was evaluated through a retrospective survey. Over 12 years, 16 outbreaks occurred, including 13 among renal transplant recipients (RTRs). We performed Pneumocystis jirovecii genotyping in 5 outbreaks, which suggested that specific strains may have been selected by RTRs.
Asunto(s)
Trasplante de Órganos , Pneumocystis carinii , Neumonía por Pneumocystis , Brotes de Enfermedades , Francia/epidemiología , Genotipo , Humanos , Trasplante de Órganos/efectos adversos , Pneumocystis carinii/genética , Neumonía por Pneumocystis/epidemiología , Estudios Retrospectivos , Encuestas y CuestionariosRESUMEN
Candida albicans is part of the human gastrointestinal (GI) microbiota. To better understand how C. albicans efficiently establishes GI colonisation, we competitively challenged growth of 572 signature-tagged strains (~10% genome coverage), each conditionally overexpressing a single gene, in the murine gut. We identified CRZ2, a transcription factor whose overexpression and deletion respectively increased and decreased early GI colonisation. Using clues from genome-wide expression and gene-set enrichment analyses, we found that the optimal activity of Crz2p occurs under hypoxia at 37°C, as evidenced by both phenotypic and transcriptomic analyses following CRZ2 genetic perturbation. Consistent with early colonisation of the GI tract, we show that CRZ2 overexpression confers resistance to acidic pH and bile salts, suggesting an adaptation to the upper sections of the gut. Genome-wide location analyses revealed that Crz2p directly modulates the expression of many mannosyltransferase- and cell-wall protein-encoding genes, suggesting a link with cell-wall function. We show that CRZ2 overexpression alters cell-wall phosphomannan abundance and increases sensitivity to tunicamycin, suggesting a role in protein glycosylation. Our study reflects the powerful use of gene overexpression as a complementary approach to gene deletion to identify relevant biological pathways involved in C. albicans interaction with the host environment.
Asunto(s)
Candida albicans/fisiología , Proteínas Fúngicas/genética , Tracto Gastrointestinal/microbiología , Animales , Candida albicans/efectos de los fármacos , Candida albicans/genética , Pared Celular/metabolismo , Femenino , Proteínas Fúngicas/metabolismo , Microbioma Gastrointestinal , Regulación Fúngica de la Expresión Génica , Redes Reguladoras de Genes , Concentración de Iones de Hidrógeno , Mananos/metabolismo , Manosiltransferasas/genética , Ratones Endogámicos BALB C , Microorganismos Modificados Genéticamente , Regiones Promotoras Genéticas , Tunicamicina/farmacologíaRESUMEN
BACKGROUND: External ophtalmomyiasis (EOM) is a zoonosis related to the presence of Oestrus ovis larvae at the ocular level in small ruminants (i.e. ovine, caprine). In humans, EOM is a rare cosmopolitan disorder, mostly described in warm and dry rural areas in patients living close to livestock areas. In metropolitan France (excluding Corsica), EOM is an exceptional disease with less than 25 cases recorded since 1917. CASE PRESENTATION: We report a case of EOM in a 19-years old man in the last week of September 2016 in Burgundy. CONCLUSION: The diagnosis of an EOM in Burgundy, a French region described as cold and humid, is surprising and could be due to a more marked climatic warming during the vegetative season in Burgundy resulting in the implantation of Diptera of the genus Oestrus sp. in this region.
Asunto(s)
Dípteros , Infecciones Parasitarias del Ojo/diagnóstico , Ojo/parasitología , Miasis/diagnóstico , Animales , Francia , Humanos , Masculino , Adulto JovenRESUMEN
Cryptosporidiosis is a common disease in children and immunodeficient individuals. In 2006, a national network was set up on the surveillance of human cryptosporidiosis in France. Since January 2015, the 41 tertiary care hospitals and the 3 private laboratories of the French National Network on the surveillance of human cryptosporidiosis have been able to declare confirmed cases of cryptosporidiosis online. Between 2015 and 2017, 210 cases of cryptosporidiosis were declared in immunodeficient patients in France; Cryptosporidium parvum and Cryptosporidium hominis represented 66% and 22% of cases, respectively. A peak was observed in autumn. Cryptosporidiosis occurred mainly in a context of solid organ transplantation (SOT) (49%) and of HIV infection (30%). In SOT recipients, cryptosporidiosis appeared more frequently in the first 6 months post transplantation. Regarding cases declared in SOT recipients, mycophenolate mofetil was used in 68%. A mortality rate of 6% was observed. Present results underline the importance of screening for cryptosporidiosis in immunocompromised patients suffering from diarrhea, especially in the course of major cell mediated immunodeficiency or even systematic screening before SOT. Exclusive Cryptosporidium free water feeding could be suggested during major cell mediated immunodeficiency.
Asunto(s)
Criptosporidiosis/epidemiología , Huésped Inmunocomprometido , Inmunoterapia/efectos adversos , Adolescente , Adulto , Anciano , Niño , Preescolar , Criptosporidiosis/inmunología , Ciclosporina/efectos adversos , Ciclosporina/uso terapéutico , Femenino , Francia/epidemiología , Humanos , Terapia de Inmunosupresión/efectos adversos , Terapia de Inmunosupresión/métodos , Inmunosupresores/efectos adversos , Inmunosupresores/uso terapéutico , Inmunoterapia/métodos , Lactante , Recién Nacido , Masculino , Persona de Mediana Edad , Ácido Micofenólico/efectos adversos , Ácido Micofenólico/uso terapéutico , Factores de Riesgo , Tacrolimus/efectos adversos , Tacrolimus/uso terapéutico , Adulto JovenRESUMEN
Fungal respiratory colonization of cystic fibrosis (CF) patients emerges as a new concern; however, the heterogeneity of mycological protocols limits investigations. We first aimed at setting up an efficient standardized protocol for mycological analysis of CF sputa that was assessed during a prospective, multicenter study: "MucoFong" program (PHRC-06/1902). Sputa from 243 CF patients from seven centers in France were collected over a 15-month period and submitted to a standardized protocol based on 6 semi-selective media. After mucolytic pretreatment, sputa were plated in parallel on cycloheximide-enriched (ACT37), erythritol-enriched (ERY37), benomyl dichloran-rose bengal (BENO37) and chromogenic (CAN37) media incubated at 37 °C and on Sabouraud-chloramphenicol (SAB27) and erythritol-enriched (ERY27) media incubated at 20-27 °C. Each plate was checked twice a week during 3 weeks. Fungi were conventionally identified; time for detection of fungal growth was noted for each species. Fungal prevalences and media performances were assessed; an optimal combination of media was determined using the Chi-squared automatic interaction detector method. At least one fungal species was isolated from 81% of sputa. Candida albicans was the most prevalent species (58.8%), followed by Aspergillus fumigatus (35.4%). Cultivation on CAN37, SAB27, ACT37 and ERY27 during 16 days provided an optimal combination, detecting C. albicans, A. fumigatus, Scedosporium apiospermum complex and Exophiala spp. with sensitivities of 96.5, 98.8, 100 and 100%. Combination of these four culture media is recommended to ensure the growth of key fungal pathogens in CF respiratory specimens. The use of such consensual protocol is of major interest for merging results from future epidemiological studies.
Asunto(s)
Fibrosis Quística/complicaciones , Hongos/clasificación , Hongos/aislamiento & purificación , Enfermedades Pulmonares Fúngicas/diagnóstico , Técnicas Microbiológicas/métodos , Técnicas Microbiológicas/normas , Esputo/microbiología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Niño , Femenino , Francia , Humanos , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Adulto JovenRESUMEN
Candida albicans is the most frequent yeast responsible for systemic infections in humans. These infections mainly originate from the gastrointestinal tract where C. albicans can invade the gut epithelial barrier to gain access to the bloodstream. Along the gut, pathogens can use Microfold (M) cells as a portal of entry to cross the epithelial barrier. M cells are specialized cells mainly located in the follicule-associated epithelium of Peyer patches. In this study, we used scanning electron and fluorescence microscopy, adhesion and invasion assays and fungal mutants to investigate the interactions of C. albicans with M cells obtained in an established in vitro model whereby enterocyte-like Caco-2 cells co-cultured with the Raji B cell line undergo a phenotypic switch to morphologically and functionally resembling M cells. Our data demonstrate that C. albicans co-localizes with and invades preferentially M cells, providing evidence that the fungus can use M cells as a portal of entry into the intestinal barrier. In addition to active penetration, F-actin dependent endocytosis contributes to internalization of the fungus into M cells through a mechanism involving hypha-associated invasins including Ssa1 and Als3.
Asunto(s)
Candida albicans/fisiología , Candidemia/microbiología , Tracto Gastrointestinal/microbiología , Interacciones Huésped-Patógeno , Ganglios Linfáticos Agregados/microbiología , Linfocitos B/fisiología , Adhesión Celular , Línea Celular , Técnicas de Cocultivo , Endocitosis , Células Epiteliales/microbiología , Células Epiteliales/fisiología , Humanos , Microscopía Electrónica de Rastreo , Microscopía FluorescenteRESUMEN
Fusarium oxysporum is typically a soilborne fungus but can also be found in aquatic environments. In hospitals, water distribution systems may be reservoirs for the fungi responsible for nosocomial infections. F. oxysporum was previously detected in the water distribution systems of five French hospitals. Sixty-eight isolates from water representative of all hospital units that were previously sampled and characterized by translation elongation factor 1α sequence typing were subjected to microsatellite analysis and full-length ribosomal intergenic spacer (IGS) sequence typing. All but three isolates shared common microsatellite loci and a common two-locus sequence type (ST). This ST has an international geographical distribution in both the water networks of hospitals and among clinical isolates. The ST dominant in water was not detected among 300 isolates of F. oxysporum that originated from surrounding soils. Further characterization of 15 isolates by vegetative compatibility testing allowed us to conclude that a clonal lineage of F. oxysporum circulates in the tap water of the different hospitals. IMPORTANCE: We demonstrated that a clonal lineage of Fusarium oxysporum inhabits the water distribution systems of several French hospitals. This clonal lineage, which appears to be particularly adapted to water networks, represents a potential risk for human infection and raises questions about its worldwide distribution.
Asunto(s)
Agua Potable/microbiología , Fusarium/genética , Fusarium/aislamiento & purificación , Hospitales , ADN de Hongos/aislamiento & purificación , ADN Intergénico , Francia/epidemiología , Fusariosis/epidemiología , Fusariosis/etiología , Fusariosis/microbiología , Fusarium/clasificación , Humanos , Repeticiones de Microsatélite , Factor 1 de Elongación Peptídica/genética , Filogenia , Análisis de Secuencia de ADNRESUMEN
Unlike bacterial infections, the value of procalcitonin (PCT) in detecting fungal infections in leukaemia patients is not clear. To determine whether the monitoring of PCT coupled with C-reactive protein (CRP) and fibrinogen (Fib) could be helpful in the management of pulmonary aspergillosis (IPA) or mucormycosis (PM), we retrospectively analysed the evolution of PCT, CRP and Fib levels in 94 leukaemia patients with proven/probable IPA (n = 77) or PM (n = 17) from D-12 to D12 relative to IFI onset defined as D0. Overall, 2140 assays were performed. From D-12 to D0, 12%, 5% and 1.4% of patients had PCT >0.5, 1 and 1.5 µg l(-1) , respectively, while CRP was >50, 75 and 100 mg l(-1) in 84%, 70% and 57% and Fib was >4, 5 and 6 g l(-1) in 96%, 80% and 61% of cases respectively (P < 10(-7) ). The same trends were observed from D1 to D12. Overall, between D-12 and D12, only 6.4% of patients had PCT >1.5 µg l(-1) , while CRP >100 mg l(-1) and Fib >6 g l(-1) were observed in 80% and 75% of cases respectively (P < 10(-7) ). In leukaemia patients, IPA or PM was accompanied by a significant increase in CRP and Fib while PCT remained low.
Asunto(s)
Proteína C-Reactiva/análisis , Calcitonina/sangre , Fibrinógeno/análisis , Aspergilosis Pulmonar Invasiva/diagnóstico , Leucemia/complicaciones , Mucormicosis/diagnóstico , Neutropenia/complicaciones , Adulto , Anciano , Anciano de 80 o más Años , Aspergillus/aislamiento & purificación , Biomarcadores/sangre , Femenino , Humanos , Inflamación/sangre , Inflamación/microbiología , Aspergilosis Pulmonar Invasiva/sangre , Aspergilosis Pulmonar Invasiva/complicaciones , Aspergilosis Pulmonar Invasiva/microbiología , Leucemia/sangre , Leucemia/tratamiento farmacológico , Masculino , Persona de Mediana Edad , Mucormicosis/sangre , Mucormicosis/microbiología , Neutropenia/sangre , Neutropenia/tratamiento farmacológico , Estudios Retrospectivos , Adulto JovenRESUMEN
Toxoplasmosis is a life-threatening infection in immunocompromised patients (ICPs). The definitive diagnosis relies on parasite DNA detection, but little is known about the incidence and burden of disease in HIV-negative patients. A 3-year retrospective study was conducted in 15 reference laboratories from the network of the French National Reference Center for Toxoplasmosis, in order to record the frequency of Toxoplasma gondii DNA detection in ICPs and to review the molecular methods used for diagnosis and the prevention measures implemented in transplant patients. During the study period, of 31,640 PCRs performed on samples from ICPs, 610 were positive (323 patients). Blood (n = 337 samples), cerebrospinal fluid (n = 101 samples), and aqueous humor (n = 100 samples) were more frequently positive. Chemoprophylaxis schemes in transplant patients differed between centers. PCR follow-up of allogeneic hematopoietic stem cell transplant (allo-HSCT) patients was implemented in 8/15 centers. Data from 180 patients (13 centers) were further analyzed regarding clinical setting and outcome. Only 68/180 (38%) patients were HIV(+); the remaining 62% consisted of 72 HSCT, 14 solid organ transplant, and 26 miscellaneous immunodeficiency patients. Cerebral toxoplasmosis and disseminated toxoplasmosis were most frequently observed in HIV and transplant patients, respectively. Of 72 allo-HSCT patients with a positive PCR result, 23 were asymptomatic; all were diagnosed in centers performing systematic blood PCR follow-up, and they received specific treatment. Overall survival of allo-HSCT patients at 2 months was better in centers with PCR follow-up than in other centers (P < 0.01). This study provides updated data on the frequency of toxoplasmosis in HIV-negative ICPs and suggests that regular PCR follow-up of allo-HSCT patients could guide preemptive treatment and improve outcome.
Asunto(s)
Huésped Inmunocomprometido , Técnicas Microbiológicas , Técnicas de Diagnóstico Molecular , Reacción en Cadena de la Polimerasa , Toxoplasma/aislamiento & purificación , Toxoplasmosis/epidemiología , Francia/epidemiología , Humanos , Prevalencia , Estudios Retrospectivos , Análisis de Supervivencia , Toxoplasma/genética , Toxoplasmosis/diagnóstico , Toxoplasmosis/parasitología , Toxoplasmosis/patologíaRESUMEN
The detection of Toxoplasma gondii in amniotic fluid is an essential tool for the prenatal diagnosis of congenital toxoplasmosis and is currently essentially based on the use of PCR. Although some consensus is emerging, this molecular diagnosis suffers from a lack of standardization and an extreme diversity of laboratory-developed methods. Commercial kits for the detection of T. gondii by PCR were recently developed and offer certain advantages; however, they must be assessed in comparison with optimized reference PCR assays. The present multicentric study aimed to compare the performances of the Bio-Evolution T. gondii detection kit and laboratory-developed PCR assays set up in eight proficient centers in France. The study compared 157 amniotic fluid samples and found concordances of 99% and 100% using 76 T. gondii-infected samples and 81 uninfected samples, respectively. Moreover, taking into account the classification of the European Research Network on Congenital Toxoplasmosis, the overall diagnostic sensitivity of all assays was identical and calculated to be 86% (54/63); specificity was 100% for all assays. Finally, the relative quantification results were in good agreement between the kit and the laboratory-developed assays. The good performances of this commercial kit are probably in part linked to the use of a number of good practices: detection in multiplicate, amplification of the repetitive DNA target rep529, and the use of an internal control for the detection of PCR inhibitors. The only drawbacks noted at the time of the study were the absence of uracil-N-glycosylase and small defects in the reliability of the production of different reagents.
Asunto(s)
Reacción en Cadena de la Polimerasa , Juego de Reactivos para Diagnóstico , Toxoplasma/genética , Toxoplasmosis Congénita/diagnóstico , Toxoplasmosis Congénita/parasitología , Líquido Amniótico/parasitología , Estudios de Cohortes , Femenino , Humanos , Ensayos de Aptitud de Laboratorios , Reacción en Cadena de la Polimerasa/métodos , Reacción en Cadena de la Polimerasa/normas , Embarazo , Juego de Reactivos para Diagnóstico/normas , Reproducibilidad de los Resultados , Estudios Retrospectivos , Sensibilidad y EspecificidadRESUMEN
Pneumocystis jirovecii pneumonia (PCP) in patients without AIDS is increasingly common. We conducted a prospective cohort study of consecutive patients with proven PCP; of 544 patients, 223 (41%) had AIDS (AIDS patients) and 321 (59%) had other immunosuppressive disorders (non-AIDS patients). Fewer AIDS than non-AIDS patients required intensive care or ventilation, and the rate of hospital deaths--17.4% overall--was significantly lower for AIDS versus non-AIDS patients (4% vs. 27%; p<0.0001). Multivariable analysis showed the odds of hospital death increased with older age, receipt of allogeneic bone marrow transplant, immediate use of oxygen, need for mechanical ventilation, and longer time to treatment; HIV-positive status or receipt of a solid organ transplant decreased odds for death. PCP is more often fatal in non-AIDS patients, but time to diagnosis affects survival and is longer for non-AIDS patients. Clinicians must maintain a high index of suspicion for PCP in immunocompromised patients who do not have AIDS.
Asunto(s)
Pneumocystis carinii , Neumonía por Pneumocystis/epidemiología , Infecciones Oportunistas Relacionadas con el SIDA/diagnóstico , Infecciones Oportunistas Relacionadas con el SIDA/epidemiología , Síndrome de Inmunodeficiencia Adquirida/diagnóstico , Síndrome de Inmunodeficiencia Adquirida/epidemiología , Adulto , Femenino , Francia/epidemiología , Humanos , Masculino , Persona de Mediana Edad , Mortalidad , Oportunidad Relativa , Neumonía por Pneumocystis/diagnóstico , Neumonía por Pneumocystis/tratamiento farmacológico , Estudios Prospectivos , Factores de RiesgoRESUMEN
The molecular diagnosis of toxoplasmosis essentially relies upon laboratory-developed methods and suffers from lack of standardization, hence the large diversity of performances between laboratories. Moreover, quantifications of parasitic loads differ among centers, a fact which prevents the possible prediction of the severity of this disease as a function of parasitic loads. The objectives of this multicentric study performed in eight proficient laboratories of the Molecular Biology Pole of the French National Reference Center for Toxoplasmosis (NRC-T) were (i) to assess the suitability of a lyophilized preparation of Toxoplasma gondii as a common standard for use in this PCR-based molecular diagnosis and (ii) to make this standard available to the community. High-quality written procedures were used for the production and qualification of this standard. Three independent batches of this standard, containing concentrations ranging from 10(4) to 0.01 T. gondii genome equivalents per PCR, were first assessed: the linear dynamic range was ≥ 6 log, the intra-assay coefficients of variation (CV) from a sample containing 10 T. gondii organisms per PCR were 0.3% to 0.42%, and the interassay CV over a 2-week period was 0.76% to 1.47%. A further assessment in eight diagnostic centers showed that the standard is stable, robust, and reliable. These lyophilized standards can easily be produced at a larger scale when needed and can be made widely available at the national level. To our knowledge, this is the first quality control assessment of a common standard which is usable both for self-evaluation in laboratories and for accurate quantification of parasitic loads in T. gondii prenatal infections.
Asunto(s)
Técnicas de Diagnóstico Molecular/métodos , Técnicas de Diagnóstico Molecular/normas , Técnicas de Amplificación de Ácido Nucleico/métodos , Técnicas de Amplificación de Ácido Nucleico/normas , Estándares de Referencia , Toxoplasma/aislamiento & purificación , Toxoplasmosis/diagnóstico , Francia , Humanos , Carga de Parásitos/métodos , Carga de Parásitos/normas , Toxoplasma/genéticaRESUMEN
BACKGROUND: Invasive candidiasis (IC) is a life-threatening ICU-acquired infection. A strong correlation between time to antifungal therapy (AFT) administration and outcome has been established. Empirical therapy benefit should be balanced with the risk of echinocandin overuse. We assessed therefore a decision rule that aimed at guiding empirical therapy. METHODS: A 45-month prospective cohort study in a teaching medical ICU. All of the patients with suspected IC (uncontrolled sepsis despite broad spectrum antibiotics without any bacterial proven infection in patients with Candida score ≥ 3 points including multifocal Candida sp. colonization) were eligible. The primary endpoint was proven IC diagnosis (i.e., candidemia) following treatment onset. Timing of AFT administration was also investigated in those latter patients. Antifungal therapy step-down and discontinuation was done according to international guidelines in patients with candidemia. Otherwise, echinocandin discontinuation was encouraged in patients without proven IC, excepting when a clinical improvement was achieved without any other explanation that antifungals initiation (i.e., probable IC). In addition, a survival multivariate analysis using a Cox model was conducted. RESULTS: Fifty-one patients were given an echinocandin with respect to our decision rule. Among them, candidemia was diagnosed thereafter in 9 patients. Over the same period, antifungal therapy was triggered by candidemia announcement (i.e., definite therapy) in 12 patients who did not fulfill criteria for empirical therapy before. Time elapsed from candidemia onset to echinocandin therapy initiation was shortened (0.4 [0.5] vs. 2.4 [2.8] hours; p = 0.04) when it was given empirically. In addition, 18 patients clinically improved under empirical antifungal therapy without any obvious other explanation, despite IC remained unproven. Moreover, echinocandin exposure duration was independently related to survival in those patients. Over the same period, our predefined criteria for empirical therapy were overruled in 55 cases. None of them develop IC thereafter. Finally, Our decision rule allowed IC early recognition of proven/probable IC with sensitivity, specificity, positive and negative predictive value of 69.2%, 82.1%, 69.2% and 82.1%, respectively. CONCLUSION: Implementation of pragmatic guidelines for empirical AFT based on CS and fungal colonization assessment could be useful in selecting patients who really benefit from an echinocandin.
Asunto(s)
Antifúngicos/administración & dosificación , Candidiasis/tratamiento farmacológico , Equinocandinas/administración & dosificación , Índice de Severidad de la Enfermedad , Anciano , Antifúngicos/uso terapéutico , Candidiasis/epidemiología , Candidiasis/patología , Estudios de Cohortes , Enfermedad Crítica , Femenino , Francia/epidemiología , Humanos , Unidades de Cuidados Intensivos , Masculino , Evaluación de Programas y Proyectos de Salud , Estudios Prospectivos , Sensibilidad y EspecificidadRESUMEN
This study aimed to validate the effectiveness of a standardised procedure for the MALDI-TOF mass spectrometry (MS)-based identification on a large sample of filamentous fungi routinely identified in university hospitals' laboratories. Non-dermatophyte filamentous fungi prospectively isolated in the routine activity of five teaching hospitals in France were first identified by conventional methods in each laboratory and then by MS in one centre. DNA sequence-based identification resolved discrepancies between both methods. In this study, of the 625 analysed filamentous fungi of 58 species, 501 (80%) and 556 (89%) were correctly identified by conventional methods and MS respectively. Compared with the conventional method, MS dramatically enhanced the performance of the identification of the non-Aspergillus filamentous fungi with a 31-61% increase in correct identification rate. In conclusion, this study on a large sample of clinical filamentous fungi taxa demonstrates that species identification is significantly improved by MS compared with the conventional method. The main limitation is that MS identification is possible only if the species is included in the reference spectra library. Nevertheless, for the routine clinical laboratory, MS provides the means to attain markedly accurate results in filamentous fungi identification, which was previously restricted to only a few reference laboratories.