Improved SARS-CoV-2 RNA recovery in wastewater matrices using a CTAB-based extraction method.

Wastewater-based epidemiology has allowed tracking the magnitude and distribution of SARS-CoV-2 in communities, allowing public health officials to prepare for impending outbreaks. While many factors influence recovery of SARS-CoV-2 from wastewater, proper extraction, concentration, and purification of RNA are key steps to ensure accurate detection of viral particles. The aim of this study was to compare the efficiency of four commonly used RNA extraction methods for detection of the SARS-CoV-2 RNA genome in sewage samples artificially inoculated with the virus, in order to identify a protocol that improves viral recovery. These methods included CTAB-based, TRIzol-based, and guanidinium thiocyanate (GTC)-based extraction procedures coupled with silica spin column-based purification, and an automated extraction/purification protocol using paramagnetic particles. Following RNA extraction, virus recovery rates were compared using RT-qPCR-based detection. The CTAB-based approach yielded the highest recovery rates and was the only method to consistently demonstrate stable virus recovery percentages regardless of the specific physicochemical characteristics of the samples tested. The TRIzol method proved to be the second most effective, yielding significantly higher recovery rates compared to both the GTC-based and the automated extraction methods. These results suggest that the CTAB-based approach could be a useful tool for the recovery of viral RNA from complex wastewater matrices.

Surveillance of Shiga toxin-producing Escherichia coli associated bloody diarrhea in Argentina / Vigilancia de diarrea sanguinolenta asociada a Escherichia coli productor de toxina Shiga en Argentina

Abstract In Argentina, hemolytic uremic syndrome (HUS) caused by Shiga toxin-producing Escherichia coli (STEC-HUS) infection is endemic, and reliable data about prevalence and risk factors have been available since 2000. However, information about STEC-associated bloody diarrhea (BD) is limited. A prospective study was performed during the period October Surveillance; 2018-June 2019 in seven tertiary-hospitals and 18 referral units from different regions, aiming of STEC-HUS cases in the same hospitals and during the same period were also assessed. Twenty-nine (4.1%) of the BD patients were STEC-positive, as determined by the Shiga Toxin Quik Chek (STQC) test and/or the multiplex polymerase chain reaction (mPCR) assay. The highest fre-quencies were found in the Southern region (Neuquén, 8.7%; Bahía Blanca, 7.9%), in children between 12 and 23 month of age (8.8%), during summertime. Four (13.8%) cases progressed to HUS, three to nine days after diarrhea onset. Twenty-seven STEC-HUS in children under 5 years of age (77.8%) were enrolled, 51.9% were female; 44% were Stx-positive by STQC and all by mPCR. The most common serotypes were O157:H7 and O145:H28 and the prevalent geno-types, both among BD and HUS cases, were sfx2a-only or -associated. Considering the endemic behavior of HUS and its high incidence, these data show that the rate of STEC-positive cases is low among BD patients. However, the early recognition of STEC-positive cases is important for patient monitoring and initiation of supportive treatment.

Resumen En Argentina, el síndrome urémico hemolítico asociado a Escherichia coli productor de toxina Shiga (STEC-SUH) es endémico y, desde 2000, de notificación obligatoria. Sin embargo, la información sobre diarrea sanguinolenta (DS) asociada a STEC (DS-STEC) es limitada. Se realizó un estudio prospectivo desde octubre de 2018 hasta junio de 2019 en siete hospitales de tercer nivel y 18 unidades de referencia de diferentes provincias argentinas, con el objetivo de determinar la frecuencia de casos de DS-STEC en 714 niños de 1 a 9 años que tuvieron DS (I) y la tasa de progresión de DS a SUH en dicha cohorte (II). También se evaluó el número y distribución regional de casos de STEC-SUH en los mismos hospitales en dicho período. Veintinueve casos de DS (4,1%) fueron STEC-positivos, determinados por Shiga Toxin Quik Chek (STQC) o PCR múltiple (mPCR). Las frecuencias más altas se encontraron en el sur del área relevada (Neuquén, 8,7%; Bahía Blanca, 7,9%), en niños de 12 a 23 meses (8,8%), en verano. Cuatro casos de DS-STEC (13,8%) progresaron a SUH, de tres a nueve días después del inicio de la diarrea. Se registraron 27 niños con STEC-SUH, estos fueron mayoritariamente <5 anos (77,8%) del sexo femenino (51,9%). El 44% de estos casos fueron Stx-positivos por STQC y todos por mPCR. Los serotipos más comunes fueron O157:H7y O145:H28, y el genotipo predominante fue stx2a, solo o asociado, en DS y SUH. Considerando el comportamiento endémico del SUH y su alta incidencia, estos datos muestran que la tasa de casos de DS-STEC es baja. Sin embargo, su reconocimiento temprano es importante para el seguimiento e inicio del tratamiento de sostén.

Surveillance of Shiga toxin-producing Escherichia coli associated bloody diarrhea in Argentina.

In Argentina, hemolytic uremic syndrome (HUS) caused by Shiga toxin-producing Escherichia coli (STEC-HUS) infection is endemic, and reliable data about prevalence and risk factors have been available since 2000. However, information about STEC-associated bloody diarrhea (BD) is limited. A prospective study was performed during the period October 2018-June 2019 in seven tertiary-hospitals and 18 referral units from different regions, aiming to determine (i) the frequency of STEC-positive BD cases in 714 children aged 1-9 years of age and (ii) the rate of progression of bloody diarrhea to HUS. The number and regional distribution of STEC-HUS cases in the same hospitals and during the same period were also assessed. Twenty-nine (4.1%) of the BD patients were STEC-positive, as determined by the Shiga Toxin Quik Chek (STQC) test and/or the multiplex polymerase chain reaction (mPCR) assay. The highest frequencies were found in the Southern region (Neuquén, 8.7%; Bahía Blanca, 7.9%), in children between 12 and 23 month of age (8.8%), during summertime. Four (13.8%) cases progressed to HUS, three to nine days after diarrhea onset. Twenty-seven STEC-HUS in children under 5 years of age (77.8%) were enrolled, 51.9% were female; 44% were Stx-positive by STQC and all by mPCR. The most common serotypes were O157:H7 and O145:H28 and the prevalent genotypes, both among BD and HUS cases, were stx2a-only or -associated. Considering the endemic behavior of HUS and its high incidence, these data show that the rate of STEC-positive cases is low among BD patients. However, the early recognition of STEC-positive cases is important for patient monitoring and initiation of supportive treatment.

The Lambda Variant in Argentina: Analyzing the Evolution and Spread of SARS-CoV-2 Lineage C.37.

The second wave of COVID-19 occurred in South America in early 2021 and was mainly driven by Gamma and Lambda variants. In this study, we aimed to describe the emergence and local genomic diversity of the SARS-CoV-2 Lambda variant in Argentina, from its initial entry into the country until its detection ceased. Molecular surveillance was conducted on 9356 samples from Argentina between October 2020 and April 2022, and sequencing, phylogenetic, and phylogeographic analyses were performed. Our findings revealed that the Lambda variant was first detected in Argentina in January 2021 and steadily increased in frequency until it peaked in April 2021, with continued detection throughout the year. Phylodynamic analyses showed that at least 18 introductions of the Lambda variant into the country occurred, with nine of them having evidence of onward local transmission. The spatial--temporal reconstruction showed that Argentine clades were associated with Lambda sequences from Latin America and suggested an initial diversification in the Metropolitan Area of Buenos Aires before spreading to other regions in Argentina. Genetic analyses of genome sequences allowed us to describe the mutational patterns of the Argentine Lambda sequences and detect the emergence of rare mutations in an immunocompromised patient. Our study highlights the importance of genomic surveillance in identifying the introduction and geographical distribution of the SARS-CoV-2 Lambda variant, as well as in monitoring the emergence of mutations that could be involved in the evolutionary leaps that characterize variants of concern.

Assessing the hidden diversity underlying consensus sequences of SARS-CoV-2 using VICOS, a novel bioinformatic pipeline for identification of mixed viral populations.

INTRODUCTION: Coinfection with two SARS-CoV-2 viruses is still a very understudied phenomenon. Although next generation sequencing methods are very sensitive to detect heterogeneous viral populations in a sample, there is no standardized method for their characterization, so their clinical and epidemiological importance is unknown. MATERIAL AND METHODS: We developed VICOS (Viral COinfection Surveillance), a new bioinformatic algorithm for variant calling, filtering and statistical analysis to identify samples suspected of being mixed SARS-CoV-2 populations from a large dataset in the framework of a community genomic surveillance. VICOS was used to detect SARS-CoV-2 coinfections in a dataset of 1,097 complete genomes collected between March 2020 and August 2021 in Argentina. RESULTS: We detected 23 cases (2%) of SARS-CoV-2 coinfections. Detailed study of VICOS's results together with additional phylogenetic analysis revealed 3 cases of coinfections by two viruses of the same lineage, 2 cases by viruses of different genetic lineages, 13 were compatible with both coinfection and intra-host evolution, and 5 cases were likely a product of laboratory contamination. DISCUSSION: Intra-sample viral diversity provides important information to understand the transmission dynamics of SARS-CoV-2. Advanced bioinformatics tools, such as VICOS, are a necessary resource to help unveil the hidden diversity of SARS-CoV-2.

Evaluation of viral concentration methods for SARS-CoV-2 recovery from wastewaters.

Wastewater-based epidemiology (WBE) is a useful tool that has the potential to act as a complementary approach to monitor the presence of SARS-CoV-2 in the community and as an early alarm system for COVID-19 outbreak. Many studies reported low concentrations of SARS-CoV-2 in sewage and also revealed the need for methodological validation for enveloped viruses concentration in wastewater. The aim of this study was to evaluate different methodologies for the concentration of viruses in wastewaters and to select and improve an option that maximizes the recovery of SARS-CoV-2. A total of 11 concentration techniques based on different principles were evaluated: adsorption-elution protocols with negatively charged membranes followed by polyethylene glycol (PEG) precipitation (Methods 1-2), PEG precipitation (Methods 3-7), aluminum polychloride (PAC) flocculation (Method 8), ultrafiltration (Method 9), skim milk flocculation (Method 10) and adsorption-elution with negatively charged membrane followed by ultrafiltration (Method 11). To evaluate the performance of these concentration techniques, feline calicivirus (FCV) was used as a process control in order to avoid the risk associated with handling SARS-CoV-2. Two protocols, one based on PEG precipitation and the other on PAC flocculation, showed high efficiency for FCV recovery from wastewater (62.2% and 45.0%, respectively). These two methods were then tested for the specific recovery of SARS-CoV-2. Both techniques could recover SARS-CoV-2 from wastewater, PAC flocculation showed a lower limit of detection (4.3 × 102 GC/mL) than PEG precipitation (4.3 × 103 GC/mL). This work provides a critical overview of current methods used for virus concentration in wastewaters and the analysis of sensitivity for the specific recovery of SARS-CoV-2 in sewage. The data obtained here highlights the viability of WBE for the surveillance of COVID-19 infections in the community.

Genetic characterization of human hydatid cysts shows coinfection by Echinococcus canadensis G7 and Echinococcus granulosus sensu stricto G1 in Argentina.

Human cystic echinococcosis caused by the larval stage of Echinococcus granulosus sensu lato (s.l.) is a highly endemic disease in the province of Neuquén, Patagonia, Argentina. Human infections with E. granulosus sensu stricto (s.s.) G1 and Echinococcus canadensis G6 were reported in Neuquén in previous studies, whereas four genotypes were identified in livestock: G1, G3, G6, and G7. The aim of this study was to identify the genotypes of E. granulosus s.l. isolates from humans of Neuquén province, Patagonia, Argentina, through the 2005-2014 period. Twenty six hydatid cysts were obtained from 21 patients. The most frequent locations were the liver and lungs. Single cysts were observed in 81.0% of patients, and combined infection of liver and lungs was detected in 9.5% of cases. Partial sequencing of mitochondrial cytochrome c oxidase subunit 1 (cox1) and NADH dehydrogenase subunit 1 (nad1) genes identified the presence of E. granulosus s.s. G1 (n = 11; 42.3%) including three different partial sequences; E. canadensis G6 (n = 14; 53.8%) and E. canadensis G7 (n = 1; 3.9%). Coinfection with G1 and G7 genotypes was detected in one patient who harbored three liver cysts. Most of the liver cysts corresponded to G1 and G6 genotypes. This study presents the first report in the Americas of a human infection with E. canadensis G7 and the second worldwide report of a coinfection with two different species and genotypes of E. granulosus s.l in humans. The molecular diversity of this parasite should be considered to redesign or improve the control program strategies in endemic regions.

First study about the development of adult Echinococcus canadensis G6 genotype of goat origin in experimentally infected dogs.

Echinococcus granulosus sensu lato (E. granulosus sl) must be considered as a species complex, comprising Echinococcus granulosus sensu stricto (E. granulosus ss, genotypes G1-G3), Echinococcus equinus (G4), Echinococcus ortleppi (G5) and Echinococcus canadensis (G6-G10) although the species status of E. canadensis is still controversial. These genotypes closely match the intermediate hosts associated strains described in earlier times among which E. canadensis G6 corresponds to the camel strain. As there are no studies concerning the development of adult stages of the G6 genotype from non-camel origin, the aims of the present study were: to characterize for the first time the development of E. canadensis G6 in dogs experimentally infected with protoscoleces derived from goats, to describe the resultant adult morphology, to evaluate the growth of their rostellar hooks from larval to adult stages and to determine the prepatent period of the strobilar stage of E. canadensis G6 derived from goats. The development of the strobilar stage of E. canadensis G6 genotype of goat origin was examined by studying the growth (variation of the total worm length) and segmentation in experimentally infected dogs at 14, 25, 35 and 56days post infection. A morphological characterization of 35-day-old worms as well as of larval and adult rostellar hooks was also carried out by conventional optical microscopic observations and/or by scanning electron microscopy. The prepatent period of the strobilar stage was assessed by microscopic examination of faeces from 2 infected dogs. Our results were compared with published data from the camel and other strains. The roles of the host, genotype and species in morphological and developmental features as well as the taxonomic position of E. canadensis G6 were discussed. The prepatent period of E. canadensis G6 genotype of goat origin was determined as at least, 41days. The obtained results contribute to increase the knowledge about the biology and genetics of E. granulosus sl complex and are also of practical usefulness for the design of disease control strategies.

Genotypic characterization of Escherichia coli O157:H7 strains that cause diarrhea and hemolytic uremic syndrome in Neuquén, Argentina.

Shiga toxin-producing Escherichia coli (STEC) are important food-borne pathogens associated with cases of diarrhea, hemorrhagic colitis and hemolytic uremic syndrome (HUS). E. coli O157:H7 is the dominant serotype in Argentina and also in Neuquén Province, in which HUS incidence is above the national average, with a maximum of 28.6 cases per 100,000 children less than 5 years old reported in 1998. The aim of this study was to characterize a collection of 70 STEC O157 strains isolated from patients with diarrhea and HUS treated in the province of Neuquén, Argentina, between 1998 and 2011. All strains harbored eae, ehxA, rfbO157, and fliCH7 genes, and stx2a/stx2c (78.7%) was the predominant genotype. A total of 64 (91.4%) STEC O157 strains belonged to the hypervirulent clade 8 tested using both 4 and 32 SNP typing schemes. The strains showed the highest values reported in the literature for 6 of the 7 virulence determinants described in the TW14359 O157 strain associated with the raw spinach outbreak in the U.S. in 2006. Clade 8 strains were strongly associated with two of them: ECSP_3286, factor encoding an outer membrane protein that facilitates the transport of the heme complex (P=0.001), and in particular extracellular factor ECSP_2870/2872, coding proteins related to adaptation to plant hosts (P=0.000004). The q933 allele, which has been related to high toxin production, was present in 97.1% of the strains studied for the anti-terminator Q gene. In summary, this study describes, for the first time in Argentina, the almost exclusive circulation of strains belonging to the hypervirulent clade 8, and also the presence of putative virulence factors in higher frequencies than those reported worldwide. These data may help to understand the causes of the particular epidemiological situation related to HUS in Neuquén Province.

Desarrollo y validación de una PCR múltiple para el diagnóstico de Bordetella spp / Development and validation of a multiplex PCR for diagnosis of Bordetella spp / Desenvolvimento e validação de uma PCR múltipla para o diagnóstico de Bordetella spp

El objetivo del trabajo consistió en diseñar y validar una PCR en formato convencional que permita confirmar la presencia o ausencia de Bordetella pertussis y detectar otras especies del género, como Bordetella parapertussis y Bordetella bronchiseptica, que pudieran estar involucradas en el cuadro clínico de coqueluche. A tal fin se diseñó una reacción en cadena de la polimerasa (PCR) múltiple que amplifica una secuencia del promotor del gen de Toxina Pertussis y otra del gen de la Toxina Adenilato Ciclasa-Hemolisina. Se validó la metodología siguiendo esquemas publicados anteriormente. Se optimizaron las condiciones de la PCR. Se validó la metodología obteniéndose un límite de detección para ambas secuencias de 0,5 bacterias por reacción. Se validó, además, la especificidad y robustez de la técnica. Se presenta una nueva herramienta diagnóstica optimizada y validada, que permite detectar la presencia de las especies de Bordetella más frecuentemente involucradas en el cuadro clínico de coqueluche. Su uso combinado con alguna de las PCR habituales en diagnóstico, como la PCR IS481, permite aumentar la sensibilidad del diagnóstico de esta en­fermedad, la especificidad del mismo discriminando los resultados falsos positivos/negativos y aumentar el conocimiento sobre los agentes etiológicos implicados en esta patología.

The aim of the present work was to design and validate a conventional PCR that enables to confirm the presence or absence of Bordetella pertussis and to detect other Bordetella species, such as Bordetella parapertussis metoand B. bronchiseptica, that may be involved in this pathology. To this aim, a multiplex PCR that amplifies a sequence of the promoter of the Pertussis Toxin gene and a sequence of the Adenylate Cyclase Toxin-Hemolysin gene were designed. The PCR was validated following previously published schemes. PCR conditions were optimized. The methodology was validated obtaining a detection limit of 0.5 bacteria per reaction, for both sequences. Specificity and robustness of the technique were also validated. A new optimized and validated tool to detect the presence of the Bordetella species most frequently responsible of pertussis was presented. The combined use with some of the usual PCR, such as IS 481, may increase the sensitivity of the diagnosis of this disease, its specificity discriminating false positive/negative results and increase awareness of the etiologic agents involved in this pathology.

O objetivo do trabalho foi desenhar e validar uma reação em cadeia da polimerase (PCR) em formato convencional que permita confirmar a presença ou ausência de Bordetella pertussis e detectar outras espécies do gênero, como Bordetella parapertussis e Bordetella bronchiseptica, que pudessem estar envolvidas no quadro clínico de coqueluche. Para tal, foi desenhada uma PCR múltipla que amplifica uma sequência do promotor do gene de Toxina Pertussis e outra do gene da Toxina Adenilato Ciclase-Hemolisina. A metodologia foi validada seguindo esquemas publicados anteriormente. Foram otimizadas as condições da PCR. Validou-se a metodologia obtendo-se um limite de detecção para ambas as sequências de 0,5 bactérias por reação. Validou-se também a especificidade e robustez da técnica. Apresenta-se uma nova ferramenta diagnóstica otimizada e validada, que permite detectar a presença das espécies de Bordetella mais frequentemente envolvidas no quadro clínico de coqueluche. Seu uso combinado com alguma das PCR habituais em diagnóstico, como a PCR IS481, permite aumentar a sensibilidade do diagnóstico desta doença, a especificidade do mesmo discriminando os resultados falsos positivos/negativos e aumentar o conhecimento sobre os agentes etiológicos envolvidos nesta patologia.

Desarrollo y validación de una PCR múltiple para el diagnóstico de Bordetella spp. / Development and validation of a multiplex PCR for diagnosis of Bordetella spp. / Desenvolvimento e validaþÒo de uma PCR múltipla para o diagnóstico de Bordetella spp.

El objetivo del trabajo consistió en diseñar y validar una PCR en formato convencional que permita confirmar la presencia o ausencia de Bordetella pertussis y detectar otras especies del género, como Bordetella parapertussis y Bordetella bronchiseptica, que pudieran estar involucradas en el cuadro clínico de coqueluche. A tal fin se diseñó una reacción en cadena de la polimerasa (PCR) múltiple que amplifica una secuencia del promotor del gen de Toxina Pertussis y otra del gen de la Toxina Adenilato Ciclasa-Hemolisina. Se validó la metodología siguiendo esquemas publicados anteriormente. Se optimizaron las condiciones de la PCR. Se validó la metodología obteniéndose un límite de detección para ambas secuencias de 0,5 bacterias por reacción. Se validó, además, la especificidad y robustez de la técnica. Se presenta una nueva herramienta diagnóstica optimizada y validada, que permite detectar la presencia de las especies de Bordetella más frecuentemente involucradas en el cuadro clínico de coqueluche. Su uso combinado con alguna de las PCR habituales en diagnóstico, como la PCR IS481, permite aumentar la sensibilidad del diagnóstico de esta en¡fermedad, la especificidad del mismo discriminando los resultados falsos positivos/negativos y aumentar el conocimiento sobre los agentes etiológicos implicados en esta patología.(AU)

The aim of the present work was to design and validate a conventional PCR that enables to confirm the presence or absence of Bordetella pertussis and to detect other Bordetella species, such as Bordetella parapertussis metoand B. bronchiseptica, that may be involved in this pathology. To this aim, a multiplex PCR that amplifies a sequence of the promoter of the Pertussis Toxin gene and a sequence of the Adenylate Cyclase Toxin-Hemolysin gene were designed. The PCR was validated following previously published schemes. PCR conditions were optimized. The methodology was validated obtaining a detection limit of 0.5 bacteria per reaction, for both sequences. Specificity and robustness of the technique were also validated. A new optimized and validated tool to detect the presence of the Bordetella species most frequently responsible of pertussis was presented. The combined use with some of the usual PCR, such as IS 481, may increase the sensitivity of the diagnosis of this disease, its specificity discriminating false positive/negative results and increase awareness of the etiologic agents involved in this pathology.(AU)

O objetivo do trabalho foi desenhar e validar uma reaþÒo em cadeia da polimerase (PCR) em formato convencional que permita confirmar a presenþa ou ausÛncia de Bordetella pertussis e detectar outras espécies do gÛnero, como Bordetella parapertussis e Bordetella bronchiseptica, que pudessem estar envolvidas no quadro clínico de coqueluche. Para tal, foi desenhada uma PCR múltipla que amplifica uma sequÛncia do promotor do gene de Toxina Pertussis e outra do gene da Toxina Adenilato Ciclase-Hemolisina. A metodologia foi validada seguindo esquemas publicados anteriormente. Foram otimizadas as condiþ§es da PCR. Validou-se a metodologia obtendo-se um limite de detecþÒo para ambas as sequÛncias de 0,5 bactérias por reaþÒo. Validou-se também a especificidade e robustez da técnica. Apresenta-se uma nova ferramenta diagnóstica otimizada e validada, que permite detectar a presenþa das espécies de Bordetella mais frequentemente envolvidas no quadro clínico de coqueluche. Seu uso combinado com alguma das PCR habituais em diagnóstico, como a PCR IS481, permite aumentar a sensibilidade do diagnóstico desta doenþa, a especificidade do mesmo discriminando os resultados falsos positivos/negativos e aumentar o conhecimento sobre os agentes etiológicos envolvidos nesta patologia.(AU)

[Direct immunofluorescence assay performance in diagnosis of the Influenza A(H1N1) virus]. / Desempeño de la prueba de inmunofluorescencia directa en el diagnóstico del virus Influenza A(H1N1).

By 25 April 2009, less than one month after the first human with Influenza A(H1N1) virus was detected in Mexico, the disease had already spread to more than 40 countries, with over 10,000 cases reported. Due to its unpredictability, this type of virus requires appropriate, reliable, and safe diagnostic methods that are also accessible to clinical laboratories. Through the analysis of 291 samples taken from patients with suspected Influenza A(H1N1) virus infection in Neuquén, Argentina, this study compares the two diagnostic methods used simultaneously: direct immunofluorescence assay (DFA) and real-time polymerase chain reaction (RT-PCR). DFA had a sensitivity of 44.4%, a specificity of 99.6%, a positive predictive value of 95.2%, and a negative predictive value of 90.7%. Positive results obtained with this method can be considered true positives. A negative result does not rule out the presence of the virus. In this case, the sample should be examined by RT-PCR. Out of a total of 291 samples, there were 45 positive results with RT-PCR and 21 positive results with DFA.

Desempeño de la prueba de inmunofluorescencia directa en el diagnóstico del virus Influenza A(H1N1) / Direct immunofluorescence assay performance in diagnosis of the Influenza A(H1N1) virus

El 25 de abril de 2009, a menos de un mes de la detección en México del primer humano con virus Influenza A(H1N1), la enfermedad ya se había propagado a más de 40 países superando los 10 000 casos notificados. Dada su naturaleza impredecible, este tipo de virus requiere métodos diagnósticos apropiados, confiables y seguros, pero que también estén al alcance de los laboratorios clínicos. Mediante el estudio de 291 muestras de pacientes con sospecha de infección por virus Influenza A(H1N1) en Neuquén, Argentina, el presente trabajo compara los dos métodos de diagnóstico utilizados simultáneamente: la prueba de inmunofluorescencia directa (DFA) y la de reacción en cadena de la polimerasa en tiempo real (RT-PCR). La DFA dio una sensibilidad de 44,4 por ciento, especificidad de 99,6 por ciento, valor predictivo positivo de 95,2 por ciento y valor predictivo negativo de 90,7 por ciento. Los resultados positivos de la metodología pueden considerarse verdaderos positivos. Un resultado negativo no excluye la presencia del virus y la muestra debe examinarse mediante RT-PCR. Del total de 291 muestras, 45 resultaron positivas por RT-PCR y 21 por DFA.

By 25 April 2009, less than one month after the first human with Influenza A(H1N1) virus was detected in Mexico, the disease had already spread to more than 40 countries, with over 10 000 cases reported. Due to its unpredictability, this type of virus requires appropriate, reliable, and safe diagnostic methods that are also accessible to clinical laboratories. Through the analysis of 291 samples taken from patients with suspected Influenza A(H1N1) virus infection in Neuquén, Argentina, this study compares the two diagnostic methods used simultaneously: direct immunofluorescence assay (DFA) and real-time polymerase chain reaction (RT-PCR). DFA had a sensitivity of 44.4 percent, a specificity of 99.6 percent, a positive predictive value of 95.2 percent, and a negative predictive value of 90.7 percent. Positive results obtained with this method can be considered true positives. A negative result does not rule out the presence of the virus. In this case, the sample should be examined by RT-PCR. Out of a total of 291 samples, there were 45 positive results with RT-PCR and 21 positive results with DFA.

Articulo original: desempeño de la prueba de inmunofluorescencia directa en el diagnostico del virus influenza A(H1N1)

El 25 de abril de 2009, a menos de un mes de la detección en México del primer humano con virus InfluenzaA(H1N1), la enfermedad ya se había propagado a más de 40 países superando los 10 000 casos notificados. Dada su naturaleza impredecible, este tipo de virus requiere métodos diagnósticos apropiados, confiables y seguros, pero que también estén al alcance de los laboratorios clínicos.