Consenso Argentino de toxoplasmosis congénita 2020 / Argentine Consensus of congenital toxoplasmosis

Resumen La transmisión vertical de la infección por Toxoplasma gondii ocurre cuando la madre se infecta por primera vez en el transcurso del embarazo. El diagnóstico de la infección materna y la del re cién nacido se logra con el conjunto de pruebas serológicas, hallazgos clínicos y ecográficos. El reconocimiento temprano de la infección materna permite un tratamiento que reduce la tasa de transmisión y el riesgo de daño en el producto de la concepción. El objetivo de este consenso de expertos fue revisar la literatura científica para actualizar las recomendaciones de práctica clínica respecto de la prevención, el diagnóstico y el tratamiento de la toxoplasmosis congénita en nuestro país.

Abstract Mother-to-child transmission in Toxoplasma gondii infection occurs only when the infection is acquired for the first time during pregnancy. Diag nosis of maternal infection and the newborn is achieved by a combination of serological tests, clinical features and ultrasound images. An early diagnosis of maternal infection allows treatment that offers a reduction both in transmission rate and risk of congenital damage. The aim of this expert consensus was to review the scientific literature which would enable an update of the clinical practice guideline of prevention, diagnosis and treatment of congenital toxoplasmosis in our country.

[Argentine consensus of congenital toxoplasmosis]. / Consenso argentino de toxoplasmosis congénita 2020.

Mother-to-child transmission in Toxoplasma gondii infection occurs only when the infection is acquired for the first time during pregnancy. Diagnosis of maternal infection and the newborn is achieved by a combination of serological tests, clinical features and ultrasound images. An early diagnosis of maternal infection allows treatment that offers a reduction both in transmission rate and risk of congenital damage. The aim of this expert consensus was to review the scientific literature which would enable an update of the clinical practice guideline of prevention, diagnosis and treatment of congenital toxoplasmosis in our country.

La transmisión vertical de la infección por Toxoplasma gondii ocurre cuando la madre se infecta por primera vez en el transcurso del embarazo. El diagnóstico de la infección materna y la del recién nacido se logra con el conjunto de pruebas serológicas, hallazgos clínicos y ecográficos. El reconocimiento temprano de la infección materna permite un tratamiento que reduce la tasa de transmisión y el riesgo de daño en el producto de la concepción. El objetivo de este consenso de expertos fue revisar la literatura científica para actualizar las recomendaciones de práctica clínica respecto de la prevención, el diagnóstico y el tratamiento de la toxoplasmosis congénita en nuestro país.

Cystic echinococcosis in South America: a call for action.

Cystic echinococcosis (CE) or hydatidosis, a parasitic zoonosis caused by a cestode of the family Taeniidae, species Echinococcus granulosus, is endemic in Argentina, Chile, Peru, Uruguay, and southern Brazil. This report presents CE figures for these five countries in 2009 - 2014 and proposes indicators to measure national control programs. Nearly 5 000 new CE cases were diagnosed annually in the five countries during the study period. The average case fatality rate was 2.9%, which suggests that CE led to approximately 880 deaths in these countries during the 6-year period. CE cases that required secondary or tertiary health care had average hospital stays of 10.6 days, causing a significant burden to health systems. The proportion of new cases (15%) in children less than 15 years of age suggests ongoing transmission. Despite figures showing that CE is not under control in South America, the long-standing implementation of national and local control programs in three of the five countries has achieved reductions in some of the indicators. The Regional Initiative for the Control of CE, which includes the five countries and provides a framework for networking and collaboration, must intensify its efforts.

[Contribution of the PCR assay to the diagnosis of Mansonella ozzardi in endemic areas of Argentina]. / Aporte de la técnica de PCR en el diagnóstico de Mansonella ozzardi en zonas endémicas de la Argentina.

Mansonella ozzardi is a tissue-dwelling parasitic nematode, the causative agent of mansonelliasis in almost all Latin American countries. It has been described along the Argentine Yungas region. The microscopic diagnosis can yield false-negative test results at low microfilaremia levels. The aim of this study was to optimize the molecular diagnostic technique and compare it with the Knott's method and standard blood smear procedures (thin blood films and thick smears) in 92 blood samples of individuals from an endemic area. The PCR technique followed by the sequencing of the amplified product yielded 100 % sensitivity compared to the Knott's test, which is considered a reference method. Seven more cases of this parasitosis could only be identified with the molecular technique.

Aporte de la técnica de PCR en el diagnóstico de Mansonella ozzardi en zonas endémicas de la Argentina / Contribution of the PCR assay to the diagnosis of Mansonella ozzardi in endemic areas of Argentina

Mansonella ozzardi es un nematode parásito tisular, agente etiológico de mansonellosis en casi la totalidad de los países latinoamericanos. En Argentina la mansonellosis ha sido descrita a lo largo de la región de las yungas. Su diagnóstico microscópico puede dar resultados falsos negativos en microfilaremias bajas. El objetivo del presente estudio fue optimizar su diagnóstico molecular y comparar los resultados con los obtenidos mediante las pruebas microscópicas de Knott, de gota gruesa y de extendido hemático fino, en 92 muestras de sangre de pacientes de zona endémica. La técnica de PCR seguida de la secuenciación del producto amplificado presentó una sensibilidad del 100 % frente al método de Knott, considerado como referencia, e incluso permitió identificar 7 casos más de la parasitosis.

Mansonella ozzardi is a tissue-dwelling parasitic nematode, the causative agent of mansonelliasis in almost all Latin American countries. It has been described along the Argentine Yungas region. The microscopic diagnosis can yield false-negative test results at low microfilaremia levels. The aim of this study was to optimize the molecular diagnostic technique and compare it with the Knott's method and standard blood smear procedures (thin blood films and thick smears) in 92 blood samples of individuals from an endemic area. The PCR technique followed by the sequencing of the amplified product yielded 100 % sensitivity compared to the Knott's test, which is considered a reference method. Seven more cases of this parasitosis could only be identified with the molecular technique.

Primer acuerdo nacional de referentes para la prevención de las complicaciones y del diagnóstico y tratamiendo de la toxoplasmosis / First national agreement for prevention of complications, diagnosis and treatment of toxoplasmosis

El presente trabajo es un relato de experiencia de la reuniónrealizada en la ciudad de Alta Gracia, Córdoba, donde se llevocabo el “1er Acuerdo Nacional de Referentes para la Prevenciónde las Complicaciones y del Diagnóstico y Tratamiento de laToxoplasmosis”. Se discutieron conocimientos validados porla literatura científica como base para propuestas racionalesde diagnóstico.Los objetivos del acuerdo fueron: discutir y reflexionarsobre las prácticas y diagnóstico de la Toxoplasmosis enel sistema público nacional, consensuar y proponer unalgoritmo de diagnóstico para el sistema público, analizarla brecha entre el nivel de conocimiento y la aplicación depracticas y procedimientos en la población general y en losgrupos de riesgo, discutir y proponer actividades destinadasa identificar el riesgo de transmisión vertical con el fin deproponer estrategias de tamizaje adecuados a la prevalencianacional, proponer actividades para reducir la brechaconocimiento/aplicación de ese conocimiento en un marcode equidad.Los profesionales referentes acordaron la firma del Actade Alta Gracia que contempla para la red pública de atenciónmédica el diagnóstico de Toxoplasmosis con equidad, calidad,acceso a metodologías complejas, cooperación y asesoríatécnica y disponibilidad oportuna de insumos en casos deemergencias jurisdiccionales.

Impacto de la hidatidosis: Influencia de factores ambientales y socioculturales en Tucumán, Argentina / Impact of the Hydatidosis: Influence of Environment and Social-Culture Factors. Tucumán - Argentina

Objetivo: Exponer experiencias de trabajo en terreno sobre la epidemiología y control de hidatidosis en la región de Alta Montaña de Tucumán, su impacto social, económico y sanitario. Material y Métodos: Mediante visitas programadas a la región endémica se estudiaron factores socioculturales establecidos en la población y su contribución para la permanencia de la enfermedad en la región de Alta Montaña de Tucumán. Se consultaron registros para medir el impacto de la patología en el hombre y en los sistemas de salud. Resultados: La comunidad conserva usos y costumbres que la identifican culturalmente. Presenta bajo nivel de instrucción y escasa accesibilidad a los servicios de salud y educación. La evaluación de usos, costumbres, actitudes y creencias demostró un gran desconocimiento de la población respecto a la patología hidatídica, con una interpretación distorsionada sobre ella. Existen prácticas que contribuyen a la persistencia de la hidatidosis como la costumbre de alimentar sus perros con vísceras crudas, estrecha convivencia con ellos, alta población canina con permanencia en el ámbito doméstico. Conclusiones: La hidatidosis en la región de Alta Montaña de Tucumán está asociada a características sociales y culturales que tienen una profunda implicancia en la perpetuación del ciclo parasitario. El entorno contaminado y ciertas actividades domésticas son factores de riesgo que determinan la presencia de la enfermedad, particularmente en la población infantil. No obstante el trabajo realizado para el control, se requieren propuestas para la atención integral de la salud que contribuyan al bienestar social y sanitario de estas comunidades.

Objective: To set out experiences of fieldwork on epidemiology and control of endemic hydatidosis in the Tucuman Mountain Range, its social, economic and sanitary impacts. Material and Methods: By means of programmed visits to the region, socio-cultural factors were studied in the population and their contribution to the endemic character of the disease in the Tucuman Mountain Range. The impact of the pathology on humans and the Health System was evaluated on the basis of existing records. Results: The populations have social habits and customs that define their cultural imprint. They present low level of education and little access to health and education services. The assessment of uses, customs, attitudes and beliefs hasshown their lack of knowledge about hydatidic disease andconsequences. Some habits such as feeding dogs with raw meat, live together with dogs, their high number in the house contribute to the persistence of the endemic hydatidosis. Conclusions: The hydatidosis in the region of High Mountain of Tucuman is associated to social and cultural characteristics that have deep consequences in the persistence of the parasitic cycle. The contaminated environment and certain domestic activities are factors of risk that determine the presence of the disease, especially among the children population. In spite of a tight control of the disease, proposals for an integral attention of the community health is required.

Hidatidosis en Argentina: carga de enfermedad

Con la finalidad de producir conocimiento sobre el impacto de la Echinococcosis quística (hidatidosis) en el sistema de salud de Argentina, se evaluarán indicadores relacionados con la muerte (mortalidad); con los años perdidos de vida, ajustados en función de la discapacidad y con los años de vida perdidos prematuramente en un período determinado. La suma de los dos indicadores es la llamada "Carga de la Enfermedad" que se expresa en la unidad tiempo, generalmente en años.

Molecular evidence for a novel encapsulated genotype of Trichinella from Patagonia, Argentina.

At present, Trichinella spiralis is the only species of this genus reported from South America. Herein, we detail a molecular analysis of a new encapsulated isolate of muscle larvae of Trichinella, found in a mountain lion (Puma concolor) coming from the Patagonia, Argentina. We studied three DNA regions previously probed to be useful for the identification of all eleven recognized Trichinella genotypes: expansion segment 5 (ES5), cytochrome c-oxidase subunit I (COI) and 5S ribosomal DNA intergenic spacer region (5S ISR). BLAST searches with these DNA sequences showed that the mitochondrial and nuclear ribosomal regions most closely resemble other Trichinella sequences available in GenBank. However, they did not exactly match any of the eleven recognized genotypes. The phylogenetic analysis from COI and 5S ISR sequences showed that the mountain lion isolate is grouped with encapsulated members, in concordance with morphological data. Furthermore, this new isolate was located at the base of the encapsulated genotypes, signifying that it is an old genotype that could have emerged earliest in this group. These data strongly suggest that this isolate from the Patagonia represents the twelfth genotype (T12) described in the genus Trichinella. Nevertheless, further studies are necessary to adequately establish this isolate as a unique genotype.

[Argentine Consensus of Congenital Toxoplasmosis]. / Consenso Argentino de Toxoplasmosis Congénita.

The mother-to-child transmission in Toxoplasma gondii infection occurs only when the infection is acquired for the first time during pregnancy. The prenatal and early postnatal diagnosis can only be achieved by serological testing. Serologic tests have different sensitivities, specificities and complexities, so that different tests in more than one blood sample are necessary for the diagnosis. Serological follow-up of the infants should be conducted during the first year of life or until the diagnosis of congenital toxoplasmosis can be ruled out. Treatment recommendations try to reduce the transmission rate and the risk of congenital damage. Congenital toxoplasmosis incidence rate is approximately 5 per 1000 births, but can be reduced to 0.5 per 1000 with an active screening program. The aim of this consensus group was to review the scientific literature on congenital toxoplasmosis and prepare a statement on prevention, diagnosis and treatment that should be implemented in our country.

Consenso argentino de toxoplasmosis congénita / Argentine Consensus of Congenital Toxoplasmosis

La transmisión de la infección por Toxoplasma gondii de la madre al hijo ocurre cuando la madre se infecta por primera vez en el transcurso del embarazo. Tanto el diagnóstico prenatal, como el del primer año de vida se basa en pruebas serológicas; y la mayoría de las veces es necesario realizar más de una de estas pruebas ya que tienen distintos porcentajes de sensibilidad y/o especificidad así como distintos niveles de complejidad. El recién nacido requiere seguimiento serológico en el primer año de vida o hasta que se descarte el diagnóstico de toxoplasmosis congénita. El diagnóstico temprano de la infección, en la mujer embarazada, permite un tratamiento oportuno y se indica con el propósito de reducir la tasa de transmisión y el daño congénito. Es posible que con un programa activo, de prevención y tratamiento temprano, se pueda reducir la tasa de incidencia de la toxoplasmosis congénita de alrededor del 5 por mil nacimientos a 0.5 por mil. El objetivo de este consenso fue revisar la literatura científica para la prevención, el diagnóstico y el tratamiento de la toxoplasmosis congénita, para que se pueda implementar en nuestro país.

The mother-to-child transmission in Toxoplasma gondii infection occurs only when the infection is acquired for the first time during pregnancy. The prenatal and early postnatal diagnosis can only be achieved by serological testing. Serologic tests have different sensitivities, specificities and complexities, so that different tests in more than one blood sample are necessary for the diagnosis. Serological follow-up of the infants should be conducted during the first year of life or until the diagnosis of congenital toxoplasmosis can be ruled out. Treatment recommendations try to reduce the transmission rate and the risk of congenital damage. Congenital toxoplasmosis incidence rate is approximately 5 per 1000 births, but can be reduced to 0.5 per 1000 with an active screening program. The aim of this consensus group was to review the scientific literature on congenital toxoplasmosis and prepare a statement on prevention, diagnosis and treatment that should be implemented in our country.

Echinococcus granulosus antigen B gene family: further studies of strain polymorphism at the genomic and transcriptional levels.

Echinococcus granulosus AgB gene family is constituted by five gene loci. In a previous study, we analyzed the strain variation of EgAgB2/B4 sequences. Here, we have analyzed, by SSCP and sequencing, 250 genomic clones of EgAgB1/B3/B5 gene cluster from five E. granulosus strains. Several new EgAgB genomic variants were found. EgAgB1 and EgAgB3 genomic sequences grouped E. granulosus strains by phylogenetic tools in two clusters: one formed by G1/G2 and the other by G5, G6/G7 strains, in accordance with other molecular markers. EgAgB5 genomic and cDNA sequences were only found in G1/G2 cluster. Reverse transcription-PCR analysis using subunit specific primers revealed that all the EgAgB genes were transcribed in G1 and G7 strains with the exception of EgAgB5 transcripts that were not detected in G7 strain. Interestingly, AgB2 transcripts that were probably processed by an aberrant splicing mechanism leading to a non-functional B2 protein were found in G7 strain.

Identification of the first strain of swine hepatitis E virus in South America and prevalence of anti-HEV antibodies in swine in Argentina.

In Argentina, a country considered non-endemic for hepatitis E virus (HEV) infection, serologic evidence of HEV infection has been observed in different human population groups. In other countries, a high degree of genetic relatedness has been observed between human and swine HEV genotype 3 sequences, suggesting zoonosis as one probable route of infection. This is the first identification of swine HEV in South America. HEV RNA was detected and sequenced in the ORF 1 and ORF 2 regions from swine fecal samples from a herd located in Pergamino, in the province of Buenos Aires. These strains all group into genotype 3 and exhibit a close relationship to two novel HEV variants previously identified in Argentina from sporadic acute cases of non-A to -C hepatitis in humans. In addition, using a modified commercial ELISA, the presence of anti-HEV antibodies was surveyed in five provinces across the country and all five showed a prevalence of HEV antibodies, ranging from 4% to 58%. The results suggest that swine could be an important reservoir for virus transmission in Argentina as has been suggested for other non-endemic areas. The Argentine human strains and swine strain described in this article seem to be closely related to a human Austrian strain, suggesting a potential European origin of HEV infection in these cases.

Patent and pre-patent detection of Echinococcus granulosus genotypes in the definitive host.

The detection of Echinococcus granulosus in dogs is important for epidemiological surveillance and evaluation of cystic hydatic disease control programs. We report the efficacy of two PCR-based methods to detect patent and pre-patent infection in dogs experimentally infected with E. granulosus. The detection is based on amplification of a fragment of a mitochondrial gene (Mit-PCR) and a DNA repetitive element (Rep-PCR) of E. granulosus. We tested the ability of both methods to detect several genotypes of the parasite. Both PCR methods could detect E. granulosus in pre-patent and patent periods, even when microscopical observation of eggs resulted negative in fecal samples. The Mit-PCR produced the same amplification pattern for all the parasite genotypes tested while the amplification patterns with the Rep-PCR differed among groups of strains. Fecal samples collected from dogs of an endemic area were diagnosed with more sensitivity than arecoline hydrobromide purgation. These molecular methods could be applied in the confirmation of coproantigen-positive fecal samples and to verify the success of control programs.

Diagnosis of cystic echinococcosis on sheep farms in the south of Argentina: areas with a control program.

In 2000 Guarnera et al. proposed using ELISA in canine faeces collected from the ground to detect dogs infected with Echinococcus granulosus, thus determining sheep farms with active transmission. The objective was to evaluate the prevalence of E. granulosus infection in sheep farms of the Patagonia. Sheep farms were randomly selected in the Provinces of Río Negro, Chubut, Neuquén, Santa Cruz and Tierra del Fuego (areas with control programs) and La Pampa (comparison area). From one to three samples of fecal matter were obtained for each sheep farm, which were processed by means of copro-ELISA test with confirmation of positive samples by copro-Western blot. A total of 1042 samples were obtained from 352 sheep farms, 26 (7.3%) proving positive. Of these 5 (6.3%) were from La Pampa, 9 (13.8%) from Neuquén, 4 (4.7%) from Río Negro, 2 (2.9%) from Chubut, 1 (5.9%) from Santa Cruz and 5 (13.9%) from Tierra del Fuego. The identification of parasitized dogs is an essential activity upon which rests the strategy of control and surveillance. Arecoline tests or coproantigen test with fecal matter obtained directly from the dog contribute information on individual prevalence, while the use of coproantigens detected in ground-collected samples transfers the dog unit of observation to units of greater epidemiological value. In the present experience, the technique employed seems promising for its application in systems of epidemiological surveillance of cystic echinococcosis and in drawing a baseline on which to measure the progress of control programs in the Argentine Patagonia in subsequent years.

Identification and characterization of an interspersed repetitive DNA fragment in Plasmodium vivax with potential use for specific parasite detection.

We cloned and characterized a Plasmodium vivax repeat element of 7872bp named PvRE7.8. Several internal tandem repeats were found along the sequence. The repetitive nature of the PvRE7.8 element was confirmed by hybridization of a P. vivax YAC library. Based on the data bank analysis and the presence of two contiguous putative genes that may encode proteins related to DNA metabolism, PvRE7.8 could be considered an inactivated transposon-LINE element. By using Pv79 as probe or primers derived from Pv79-flanking sequences, P. vivax DNA Could be detected from whole blood and mosquito samples. We consider that the repeat element described here has potential for P. vivax malaria diagnosis and for epidemiological analysis of P. vivax transmission areas.

Cystic echinococcosis in Argentina: evolution of metacestode and clinical expression in various Echinococcus granulosus strains.

Echinococcus granulosus hydatid cysts were examined in 41 patients from Neuquén and Tucumán provinces in Argentina. Sequencing of the mitochondrial cytochrome c oxidase subunit 1 (CO1) revealed in 19 patients common sheep strain (G1), in 6 patients Tasmania sheep strain (G2), in 1 patient cattle strain (G5), and in 15 patients camel strain (G6). In Argentina the only known is the domestic cycle that affects dogs and herbivorous, including ovine, swine, cattle and goats. These strains produced a total of 58.6% of primary liver infections, 29.2% primary in lung, 2.4% primary in spleen and 9.8% were multiorgan abdominal infections. The metacestode was classified using the evolutive stages proposed by WHO-IWGE (from CE1 to CE5). We estimated that CE1 cyst has a duration of about 22 years, CE2 of 14 years, CE3 of 10 years, CE4 of 19 years and CE5 was not determined. The active types CE1 and CE2 reached 75% of all cases from all strains. In 36 patients with cysts from G1, G5 and G6 strain, there were only two asymptomatic cases. The strains of the E. granulosus complex do not present important clinical differences; only G6 seems to have higher growth rate.

Recombinant GRA4 or ROP2 protein combined with alum or the gra4 gene provides partial protection in chronic murine models of toxoplasmosis.

The efficacy of vaccination with Toxoplasma gondii recombinant GRA4 (rGRA4) and ROP2 (rRPO2) proteins and a mix of both combined with alum were evaluated in C57BL/6 and C3H mice. In C57BL/6 mice, rGRA4 and rGRA4-rROP2 immunizations generated similar levels of immunoglobulin G1 (IgG1) and IgG2a isotypes against GRA4, whereas immunizations with rROP2 and the mix induced a predominant IgG1 production against ROP2. All groups of C3H vaccinated mice exhibited higher levels of IgG1 than IgG2a. rGRA4-stimulated splenocytes from vaccinated mice produced primarily gamma interferon while those stimulated with rROP2 produced interleukin-4. Challenge of rGRA4- or rGRA4-rROP2-vaccinated mice from both strains with ME49 cysts resulted in fewer brain cysts than the controls, whereas vaccination with rROP2 alone only conferred protection to C3H mice. Immunization with a plasmid carrying the entire open reading frame of GRA4 showed a protective level similar to that of rGRA4 combined with alum. These results suggest that GRA4 can be a good candidate for a multiantigen anti-T. gondii vaccine based on the use of alum as an adjuvant.

Identification and characterization of serine proteinase inhibitors from Neospora caninum.

Two cDNA clones obtained from the Neospora caninum Expressed Sequence Tag project were selected by their homology with the Toxoplasma gondii serine proteinase inhibitor (serpin) gene, TgPI-1 and TgPI-2. One of them, named NcPI-H, showed several premature stop codons. The other cDNA, named NcPI-S, encoded a 79 amino acid protein containing a putative signal peptide and only one non-classical Kazal domain. Two other N. caninum EST sequences (NcEST1 and NcEST2) and one from Eimeria tenella (EtPI-S) were retrieved from the database. Amino acid sequence analysis suggested that NcEST1 and NcEST2 might be the N. caninum counterparts of TgPI-1 and TgPI-2, respectively. EtEST-S, as NcPI-S, is a single domain serpin. The open reading frame encoding the mature version of NcPI-S was expressed as recombinant protein, fused to a 6 histidine tag in Escherichia coli. Specific rabbit antiserum generated against the recombinant NcPI-S was used in immunoblot assays. Bands of 20, 30, 40, and 66-kDa were detected by SDS-PAGE of whole parasite homogenate. In addition, when an anti-TgPI-1 serum was used, bands of 25 and 35-kDa were detected indicating that there is no cross-reactivity between both serpins, and showing as well, the presence of another putative serpin in N. caninum. The recombinant protein NcPI-S, inhibited bacterial subtilisin completely, and showed lower inhibitory capacity on human neutrophil elastase, animal trypsin, and chymotrypsin, suggesting differences in effectiveness.