Is there a link between the human TRIM21 and Trypanosoma cruzi Clone 36 genes in Chagas' disease?

The homology between TRIM21 and Trypanosoma cruzi Antigen Clone 36 nucleotide sequence was discovered in 1998 (Winkler et al., Parasite 5, 94-95) prior to the functions of Ro52, the TRIM21 protein product, being understood. Ro52 has since been shown to be an Ubiquitin ligase targeting transcription factors, Interferon Regulatory Factor 1 (IRF1) and Interferon Regulatory Factor 3 (IRF3), in immune cells. This communication explores the possibility that there is a connection between the Clone 36 homologous sequence and perturbation of the host immune system in Chagas' disease. RNA interference by the Clone 36 transcripts is hypothesized as a mechanism for host immune suppression during acute Chagas' disease and/or for autoimmunity in chronic Chagas' disease through down-regulation of Ro52. In the chronic forms of the disease such as Chagas Cardiomyopathy, Clone 36 RNA containing its repetitive motif may down-regulate levels of Ro52 in monocytes, fibroblasts, or T cells, allowing IRF 1 and IRF3 to continuously stimulate transcription of interferons alpha and beta, a pro-inflammatory state favoring autoimmunity. In acute Chagas' disease, messenger RNA from Clone 36 could interfere with host macrophage Ro52 RNA, down-regulating the level of Ro52 so that it would stimulate less cytokine production, including IL-12/p40. This theory is thought to help the parasite avoid attack by the innate immune system early in the acute phase of the disease. Experiments with transgenic mice and genetically modified T. cruzi are discussed which may provide insight to addressing these questions.

Development of a high-throughput Alamar blue assay for the determination of influenza virus infectious dose, serum antivirus neutralization titer and virus ca/ts phenotype.

FluMist is an intranasal influenza live vaccine containing two Influenza A strains (currently H1N1 and H3N2) and one B strain (Yamagata or Victoria lineage). Characterization of the vaccine requires determination of the median tissue culture infectious dose (TCID(50)) titer, serum antivirus neutralization titer and vaccine cold adapted/temperature sensitive (ca/ts) phenotype. Visual cytopathic effect (CPE) readings are used widely in viral assays, but these are subjective and labor intensive. In response to the need for an efficient, inexpensive and high-throughput assay, a 96-well microplate assay was developed that uses Alamar blue dye staining as a replacement for CPE observation in the determination of influenza virus infectious dose, serum antivirus neutralization titer and virus ca/ts phenotype. Relative operating characteristic curves verified that there was a clear distinction between the fluorescence readings of the Alamar blue stained CPE positive and CPE negative wells. Virus titer was determined by use of both Alamar blue staining and CPE-based TCID(50) assays for wild-type and FluMist influenza vaccine strains as well as a plasmid-rescued influenza FluMist A strain containing a H5N1 derived hemmaglutinin and neuramidinase. Correlation of the two assays was measured by regression analysis and resulted in R(2) values of 0.814 (Influenza A), 0.983 (Influenza B) and 1.000 (H5N1), respectively. Serum microneutralization as well as virus ca/ts phenotype assays also showed a high concordance between readings based on CPE observation and Alamar blue staining. The Alamar blue dye assay is user friendly, environmentally safe and sensitive. Also, it is adaptable to automation, which could provide a high-throughput platform for analysis of pre-clinical and clinical samples.

Biophysical characterization of influenza virus subpopulations using field flow fractionation and multiangle light scattering: correlation of particle counts, size distribution and infectivity.

Adequate biophysical characterization of influenza virions is important for vaccine development. The influenza virus vaccines are produced from the allantoic fluid of developing chicken embryos. The process of viral replication produces a heterogeneous mixture of infectious and non-infectious viral particles with varying states of aggregation. The study of the relative distribution and behavior of different subpopulations and their inter-correlation can assist in the development of a robust process for a live virus vaccine. This report describes a field flow fractionation and multiangle light scattering (FFF-MALS) method optimized for the analysis of size distribution and total particle counts. The FFF-MALS method was compared with several other methods such as transmission electron microscopy (TEM), atomic force microscopy (AFM), size exclusion chromatography followed by MALS (SEC-MALS), quantitative reverse transcription polymerase chain reaction (RT Q-PCR), median tissue culture dose (TCID(50)), and the fluorescent focus assay (FFA). The correlation between the various methods for determining total particle counts, infectivity and size distribution is reported. The pros and cons of each of the analytical methods are discussed.

La amenaza de la enfermedad de chagas en la medicina transfunsional

La enfermedad de chagas es una infección causada por el parásito protozoo Trypanosoma cruzi. La enfermedad es endémica en los países tropicales de américa latina, incluidos centroamérica y mexico, y es la mayor causa de enfermedad y muerte en la clase social baja. desde mediados de los años 70, se viene aumentado el desplazamiento de inmigrantes de estos países hacia los estados unidos. Investigadores, incluyendo un grupo de la cruz roja americana, hicieron estudios epimiológicos que sugiren que esas personas tienen anticuerpos contra este parásito. Este artículo se enfoca hacia la biología la transmisión y la prevalencia en los Estados Unidos. También hacia las pruebas diagnósticas. el potencial y transmisión y los cambios que que esta enfermedad representa para los bancos de sangre de Estados Unidos y las comunidades de salud pública.