Comparison of diagnostic methods for Clostridium difficile-associated diarrhea

Para comparar diferentes métodos de diagnóstico de diarreas asociadas a Clostridium difficile desarrollados en el marco de un estudio colaborativo, se analizaron filtrados de materia fecal de pacientes con sintomatología compatible con esta patología. Se evaluó la actividad biológica sobre células Vero (ensayo biológico), la reactividad frente a anticuerpos anti-TcdA y anti-TcdB (dot blot) y la presencia de secuencias del gen tcdB por PCR. De 177 muestras analizadas por el ensayo biológico, 44 tuvieron títulos mayores o iguales que 64. Diecinueve muestras fueron a la vez positivas en el ensayo biológico y en el análisis por PCR. Se analizaron 149 muestras por dot blot utilizando anticuerpos anti-TcdA y anti-TcdB; 46 muestras resultaron positivas para ambas toxinas, 12 muestras fueron positivas sólo para TcdB y 5 muestras sólo para TcdA. Las divergencias entre los diferentes métodos podrían estar relacionadas con la presencia de genes truncados, con un bajo número de microorganismos en las muestras analizadas o con la degradación de las toxinas. Los resultados presentados demuestran la necesidad de implementar alternativas diagnósticas que se adapten a la compleja realidad epidemiológica de este importante patógeno intestinal.In order to compare different methods for the diagnosis of Clostridium difficile-associated diarrhea, fecal filtrates from patients presenting symptoms compatible with this condition, were analyzed. Biological activity on Vero cells (biological assay), dot blot with antibodies anti-TcdA and anti-TcdB, and a PCR assay for the tcdB gene, were evaluated. The analysis by dot blot using antiTcdA and anti-TcdB antibodies showed that 46 samples out of 149 were positive for both toxins whereas 12 samples were only positive for TcdB, and 5 samples only positive for TcdA. Discrepancies in the different methods could be related to truncated genes, low number of microorganisms in the samples and toxin degradation. The results herein presented show the need for developing diagnostic approaches compatible with the complex epidemiological situation of this clinically relevant intestinal pathogenFil: Trejo, Fernando Miguel. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas. Centro de Investigación y Desarrollo en Criotecnología de Alimentos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Criotecnología de Alimentos. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Criotecnología de Alimentos; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Microbiología General; ArgentinaFil: Rusconi, M.E.. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas. Centro de Investigación y Desarrollo en Criotecnología de Alimentos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Criotecnología de Alimentos. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Criotecnología de Alimentos; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Microbiología General; ArgentinaFil: Guzzeti, L.. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Microbiología General; Argentina. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas. Centro de Investigación y Desarrollo en Criotecnología de Alimentos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Criotecnología de Alimentos. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Criotecnología de Alimentos; ArgentinaFil: Zamboni, M.I.. Provincia de Santa Fe. Municipalidad de Rosario. Secretaría de Salud. Centro de Especialidades Médicas Ambulatorias de Rosario; ArgentinaFil: Guardati, M.C.. Hospital de Emergencias Dr. Clemente Álvarez; ArgentinaFil: Lejona, Sergio. Provincia de Santa Fe. Municipalidad de Rosario. Secretaría de Salud. Centro de Especialidades Médicas Ambulatorias de Rosario; ArgentinaFil: Perez, Pablo Fernando. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas. Centro de Investigación y Desarrollo en Criotecnología de Alimentos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Criotecnología de Alimentos. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Criotecnología de Alimentos; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Microbiología General; Argentin

Molecular Characterization of the Mg(2+)-Responsive PhoP-PhoQ Regulon in Salmonella enterica

The PhoP/PhoQ two-component system controls the extracellular magnesium deprivation response in Salmonella enterica. In addition, several virulence-associated genes that are mainly required for intramacrophage survival during the infection process are under the control of its transcriptional regulation. Despite shared Mg(2+) modulation of the expression of the PhoP-activated genes, no consensus sequence common to all of them could be detected in their promoter regions. We have investigated the transcriptional regulation and the interaction of the response regulator PhoP with the promoter regions of the PhoP-activated loci phoPQ, mgtA, slyB, pmrD, pcgL, phoN, pagC, and mgtCB. A direct repeat of the heptanucleotide sequence (G/T)GTTTA(A/T) was identified as the conserved motif recognized by PhoP to directly control the gene expression of the first five loci, among which the first four are ancestral to enterobacteria. On the other hand, no direct interaction of the response regulator with the promoter of phoN, pagC, or mgtCB was apparent by either in vitro or in vivo assays. These loci are Salmonella specific and were probably acquired by horizontal DNA transfer. Besides, sequence analysis of pag promoters revealed the presence of a conserved PhoP box in 6 out of the 12 genes analyzed. Our results strongly suggest that the expression of a set of Mg(2+)-controlled genes is driven by PhoP via unknown intermediate regulatory mechanisms that could also involve ancillary factors

PhoP Can Activate Its Target Genes in a PhoQ-Independent Manner

The PhoP/PhoQ two-component system controls the extracellular magnesium depletion response in Salmonella enterica. Previous studies have shown that PhoP is unable to up-regulate its target genes in the absence of PhoQ function. In this work, we demonstrate that PhoP overexpression can substitute for PhoQ- and phosphorylation-dependent activation. Either a high concentration of PhoP or activation via phosphorylation stimulates PhoP self-association

Impact of influenza in the post-pandemic phase: Clinical features in hospitalized patients with influenza A (H1N1) pdm09 and H3N2 viruses, during 2013 in Santa Fe, Argentina

It is important to characterize the clinical and epidemiological pattern of the influenza A (H1N1) pdm09 virus and compare it with influenza A (H3N2) virus, as surveyed in just a few studies, in order to contribute to the implementation and strengthening of influenza control and prevention strategies. The aims in this study were to describe influenza clinical and epidemiological characteristics in hospitalized patients, caused by influenza A (H1N1)pdm09 and influenza A (H3N2) viruses during 2013, in Santa Fe, Argentina. A retrospective study was conducted over 2013 among hospitalized patients with laboratory-confirmed influenza diagnosis. In contrast to patients with influenza A (H3N2) (20.5%), a higher proportion of hospitalizations associated with influenza H1N1pdm were reported among adults aged 35-65 years (42.8%). Of all patients, 73.6% had an underlying medical condition. Hospitalized patients with H1N1pdm were subject to 2.6 (95%CI, 1.0-6.8) times higher risk of severity, than those hospitalized with influenza A (H3N2). This results demonstrate the impact in the post-pandemic era of H1N1pdm virus, with increased risk of severe disease, in relation to H3N2 virus, both viruses co-circulating during 2013

Impact of influenza in the post-pandemic phase: Clinical features in hospitalized patients with influenza A (H1N1) pdm09 and H3N2 viruses, during 2013 in Santa Fe, Argentina

It is important to characterize the clinical and epidemiological pattern of the influenza A (H1N1) pdm09 virus and compare it with influenza A (H3N2) virus, as surveyed in just a few studies, in order to contribute to the implementation and strengthening of influenza control and prevention strategies. The aims in this study were to describe influenza clinical and epidemiological characteristics in hospitalized patients, caused by influenza A (H1N1)pdm09 and influenza A (H3N2) viruses during 2013, in Santa Fe, Argentina. A retrospective study was conducted over 2013 among hospitalized patients with laboratory-confirmed influenza diagnosis. In contrast to patients with influenza A (H3N2) (20.5%), a higher proportion of hospitalizations associated with influenza H1N1pdm were reported among adults aged 35-65 years (42.8%). Of all patients, 73.6% had an underlying medical condition. Hospitalized patients with H1N1pdm were subject to 2.6 (95%CI, 1.0-6.8) times higher risk of severity, than those hospitalized with influenza A (H3N2). This results demonstrate the impact in the post-pandemic era of H1N1pdm virus, with increased risk of severe disease, in relation to H3N2 virus, both viruses co-circulating during 2013

Trypanosoma cruzi discrete typing units in Chagas disease patients from endemic and non-endemic regions of Argentina

Fil: Cura, C I. Laboratorio de Biología Molecular de la Enfermedad de Chagas, Instituto de Ingeniería Genética y Biología Molecular (INGEBI-CONICET), Vuelta de Obligado 2490, 2do piso, 1428, Ciudad de Buenos Aires; Argentina.Fil: Lucero, Raúl Horacio. Instituto de Medicina Regional, Universidad Nacional del Nordeste, Av. Las Heras 727, 3500, Resistencia, Chaco; Argentina.Fil: Bisio, Margarita. Laboratorio de Biología Molecular de la Enfermedad de Chagas, Instituto de Ingeniería Genética y Biología Molecular (INGEBI-CONICET), Vuelta de Obligado 2490, 2do piso, 1428, Ciudad de Buenos Aires; Argentina.Fil: Oshiro, E. ANLIS Dr.C.G.Malbrán. Centro Nacional de Diagnóstico e Investigación en Endemo-Epidemias; Argentina.Fil: Formichelli, L B. Instituto de Medicina Regional, Universidad Nacional del Nordeste, Av. Las Heras 727, 3500, Resistencia, Chaco; Argentina.Fil: Burgos, Juan M. Departamento de Microbiología, Inmunología y Parasitología, Facultad de Medicina, Universidad de Buenos Aires, Paraguay 2155, 1121, Ciudad de Buenos Aires; Argentina.Fil: Sergio, Lejona. Área de Biología Molecular, Centro de Especialidades Médicas Ambulatorias (CEMAR), San Luis 2020, 2000, Rosario, Santa Fe; Argentina.Fil: Brusés, B L. Instituto de Medicina Regional, Universidad Nacional del Nordeste, Av. Las Heras 727, 3500, Resistencia, Chaco; Argentina.Fil: Hernández, D. Consultorio de Chagas, Facultad de Medicina, Universidad Nacional del Nordeste, Moreno 1240, 3400, Corrientes; Argentina.Fil: Severini, G V. Consultorio de Chagas, Facultad de Medicina, Universidad Nacional del Nordeste, Moreno 1240, 3400, Corrientes; Argentina.Fil: Velazquez, Elsa. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Parasitología; Argentina.Fil: Duffy, Tomas. Laboratorio de Biología Molecular de la Enfermedad de Chagas, Instituto de Ingeniería Genética y Biología Molecular (INGEBI-CONICET), Vuelta de Obligado 2490, 2do piso, 1428, Ciudad de Buenos Aires; Argentina.Fil: Anchart, E. Área de Biología Molecular, Centro de Especialidades Médicas Ambulatorias (CEMAR), San Luis 2020, 2000, Rosario, Santa Fe; Argentina.Fil: Lattes, R. Instituto de Nefrología, Cabello 3889, 1425, Ciudad de Buenos Aires; Argentina.Fil: Altcheh, J. Servicio de Parasitología y Enfermedad de Chagas, Hospital de Niños Ricardo Gutiérrez, Gallo 1330, 1425, Buenos Aires; Argentina.Fil: Freilij, H. Servicio de Parasitología y Enfermedad de Chagas, Hospital de Niños Ricardo Gutiérrez, Gallo 1330, 1425, Buenos Aires; Argentina.Fil: Diez, M. Unidad de Trasplante Cardíaco, Hospital Universitario Fundación Favaloro, Av. Belgrano 1746, 1093, Ciudad de Buenos Aires; Argentina.Fil: Nagel, C. Unidad de Trasplante Cardíaco, Hospital Universitario Fundación Favaloro, Av. Belgrano 1746, 1093, Ciudad de Buenos Aires; Argentina.Fil: Vigliano, Carlos. Unidad de Trasplante Cardíaco, Hospital Universitario Fundación Favaloro, Av. Belgrano 1746, 1093, Ciudad de Buenos Aires; Argentina.Fil: Favaloro, Liliana E. Unidad de Trasplante Cardíaco, Hospital Universitario Fundación Favaloro, Av. Belgrano 1746, 1093, Ciudad de Buenos Aires; Argentina.Fil: Favaloro, R R. Unidad de Trasplante Cardíaco, Hospital Universitario Fundación Favaloro, Av. Belgrano 1746, 1093, Ciudad de Buenos Aires; Argentina.Fil: Merino, D E. Instituto de Medicina Regional, Universidad Nacional del Nordeste, Av. Las Heras 727, 3500, Resistencia, Chaco; Argentina.Fil: Sosa-Estani, Sergio. ANLIS Dr.C.G.Malbrán. Centro Nacional de Diagnóstico e Investigación en Endemo-Epidemias; Argentina.Fil: Schijman, A G. Laboratorio de Biología Molecular de la Enfermedad de Chagas, Instituto de Ingeniería Genética y Biología Molecular (INGEBI-CONICET), Vuelta de Obligado 2490, 2do piso, 1428, Ciudad de Buenos Aires; Argentina.Genetic diversity of Trypanosoma cruzi may play a role in pathogenesis of Chagas disease forms. Natural populations are classified into 6 Discrete Typing Units (DTUs) Tc I-VI with taxonomical status. This study aimed to identify T. cruzi DTUs in bloodstream and tissue samples of Argentinean patients with Chagas disease. PCR-based strategies allowed DTU identification in 256 clinical samples from 239 Argentinean patients. Tc V prevailed in blood from both asymptomatic and symptomatic cases and Tc I was more frequent in bloodstream, cardiac tissues and chagoma samples from immunosuppressed patients. Tc II and VI were identified in a minority of cases, while Tc III and Tc IV were not detected in the studied population. Interestingly, Tc I and Tc II/VI sequences were amplified from the same skin biopsy slice from a kidney transplant patient suffering Chagas disease reactivation. Further data also revealed the occurrence of mixed DTU populations in the human chronic infection. In conclusion, our findings provide evidence of the complexity of the dynamics of T. cruzi diversity in the natural history of human Chagas disease and allege the pathogenic role of DTUs I, II, V and VI in the studied population

<i>Burkholderia puraquae</i> sp. nov., a novel species of the <i>Burkholderia cepacia</i> complex isolated from hospital settings and agricultural soils

Bacteria from the Burkholderia cepacia complex (Bcc) are capable of causing severe infections in patients with cystic fibrosis (CF). These opportunistic pathogens are also widely distributed in natural and man-made environments. After a 12-year epidemiological surveillance involving Bcc bacteria from respiratory secretions of Argentinean patients with CF and from hospital settings, we found six isolates of the Bcc with a concatenated species-specific allele sequence that differed by more than 3% from those of the Bcc with validly published names. According to the multilocus sequence analysis (MLSA), these isolates clustered with the agricultural soil strain, Burkholderia sp. PBP 78, which was already deposited in the PubMLST database. The isolates were examined using a polyphasic approach, which included 16S rRNA, recA, Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), DNA base composition, average nucleotide identities (ANIs), fatty acid profiles, and biochemical characterizations. The results of the present study demonstrate that the seven isolates represent a single novel species within the Bcc, for which the name Burkholderia puraquae sp. nov. is proposed. Burkholderia puraquae sp. nov. CAMPA 1040T (=LMG 29660T=DSM 103137T) was designated the type strain of the novel species, which can be differentiated from other species of the Bcc mainly from recA gene sequence analysis, MLSA, ANIb, MALDI-TOF MS analysis, and some biochemical tests, including the ability to grow at 42 ºC, aesculin hydrolysis, and lysine decarboxylase and β-galactosidase activities.Facultad de Ciencias ExactasCentro de Investigación y Desarrollo en Fermentaciones IndustrialesInstituto de Biotecnologia y Biologia Molecula