Etapas tempranas en la replicación del virus de la fiebre aftosa. (VFA). Cinética de la degradación parcial y sitios de estabilización del virus penetrante residual. / [Early stages in the replication of aphthous fever virus. Kinetics of partial degradation and sites for the establishment of penetrating residual virus]

At input multiplicities between 5-10 UFP/cell FMDV O1 Caseros adsorb in BHK21 Clon 13 cells at a rate of 5-20

depending whether cells are in suspension or in monolayers. At least four washings with medium are required to eliminate non specifically adsorbed virions. The remaining attached virus appears to be in contact with [quot ]specific[quot ] receptors of the cytoplasmic membrane. After penetration, 80

of virus became degraded to slow molecular weight material which is detected at first in the subcellular fraction and is gradually excreted out of the cells. The degradation process occurs from 0 to 15 minutes and it is demonstrated by a parallel decrease in infectivity and stability of purified labelled FMDV O1 Caseros. The 20

remaining virus is found firmly attached to a subcellular fraction precipitable at 15,000 xg. Parental residual infectivity remains remarkably stable for 90 min at which time newly synthetized RNA appears. Fraction of 15,000 xg from infected cultures was obtained at 60 min PI and incubated in vitro at 37 C. The infectivity remains unaltered for a period of 120 min even if the preparation is treated with 0.2

of triton x 100. Fraction of 15,000 xg from infected and control cultures obtained at different time PI were seeded on a linear sucrose gradient. A sharp peak of acid phosphatase associated with the lower mobility bands indicates the presence of intact lysosome structures. A shoulder of the enzyme activity is detected between the two main protein bands. Associated with the region rich in unbroken lysosomes, a very sharp peak of infectivity and/or 3H uridine or 32P labelled incoming virus can be detected. A different pattern of the residual incoming infectivity is found associated with the more rapid sedimentation protein band. 3H uridine incorporation at the beginning of viral RNA synthesis initiation shows that an important amount of newly synthesized viral RNA is found in the lower mobility band of the 15,000 xg fraction. Some incorporation is also detected in the faster band. The results presented here suggest that some functional activity is associated with the residual virus present at the beginning of infection in the 15,000 xg fraction. It can also be accepted that this virus penetrates by pinocytosis of specific receptors at the cytoplasmic membrane.

In vivo down regulation of HIV replication after hepatitis C superinfection.

There are increasing molecular and clinical evidences that the effects of human immunodeficiency virus (HIV) infection can be modified by coinfection with other viruses. The objective was to investigate the viral interaction between HIV and hepatitis C virus (HCV) after HCV superinfection. A 16 year-old pregnant woman was evaluated because of icteric acute hepatitis. Admission laboratory tests showed the following results: ALT 877 IU/L; AST 1822 IU/L; bilirubin 6.79 mg/dl. Diagnosis of acute HCV was based on detection of serum HCV RNA by PCR and anti-HCV seroconversion. ELISA for anti HIV testing was positive and confirmed by western blot. Serum markers for other viruses were negative. The patient was followed during 19 months; serum samples were taken monthly during this period for detection of plasma HIV and HCV RNA. Levels of plasma HIV-RNA were positive in all samples tested before and after the onset of acute hepatitis C. Six months later and a for two month period, and 13 months later for a period of one month HIV viremia was undetectable; then HIV-RNA in plasma was detectable again. In conclusion, HCV superinfection may have temporarily interfered with HIV replication in our patient. The following observations support our hypothesis: it has been demonstrated that HIV-1 replication is suppressed by HCV core protein which has transcriptional regulation properties of several viral and cellular promoters. Clinical implications of this event are not generally known and the interaction between these two viruses in dual infections is worth considering.

Relationship between diversity of hepatitis C quasispecies and histological severity of liver disease.

The aim of this work was to assess if the diversity of hepatitis C virus (HCV) quasispecies is related to histological severity and duration of infection in a cohort of untreated patients with an estimated onset of the disease. A total of 27 patients with diagnosis of chronic liver disease and history of blood transfusion (n = 16) or intravenous drug use (IDU) (n = 11) were included. All were anti-HCV positive and had detectable serum HCV-RNA. The onset and the duration of the disease were estimated from the time of the transfusion or the first drug injection. Patients who consumed drugs for more than 2 years, or were coinfected with HBV or HIV were excluded. History of alcohol intake (> 80 g/day), ALT level and age at infection were recorded. Histological assessment of grading and staging was performed according to Knodell score. The quasispecies diversity was investigated by single strand conformation polymorphism (SSCP) targeted to HVR-E2 region and SSCP pattern was evaluated as a single or multiple bands. The number of quasispecies did not correlate with the estimated duration of the disease. Patients who acquired hepatitis C by blood transfusion did not differ in number of bands from patients who were IDU. There was no correlation between the heterogeneity of HCV quasispecies and age, serum ALT, Knodell score, HAI and fibrosis. In conclusion the quasispecies diversity of E2 had no correlation with grade and stage of chronic HCV infection and the presence of quasispecies was independent of the duration of the disease.

Detección y caracterización de mutantes pre-core del virus de la hepatitis B (HBV) en pacientes cronicamente infectados / Detection and characterization of precore mutant of hepatitis B virus (HBV) in chronically infected patients

La infección por el virurs de la Hepatitis B (HBV) es causa importante de morbi-mortalidad en el mundo, hallándose asociada con afecciones hepáticas graves tales como cirrosis y/o carcinoma hepatocelular primario (HCC). Entre los diversos factores que influyen en la severidad del daño hepático pueden señalarse, la emergencia de mutantes pre-core (HBeAg defectuosa), la heterogeneidad genética del virus y del hospedador y la competencia del sistema inmune. En particular la presencia de mutantes pre-core en portadores crónicos se asocia, además, con una baja respuesta a la terapia con interferón alfa. En este trabajo presentamos evidencias de la presencia de dichas mutantes en pacientes cronicamente infectados HBeAg negativos (HB Agi positivo) en la Argentina. Al suero de estos pacientes se les extrajo el ADN viral que fue amplificado por PCR y caracterizado por análisis con enzimas de restricción. Finalmente uno de estos sueros fue secuenciado en la región pre-core evidenciando la presencia de mutaciones en los codones 15 y 28 de dicha región característicos de estas mutantes (AU)

Detección y caracterización de mutantes pre-core del virus de la hepatitis B (HBV) en pacientes crónicamente infectados. / [Detection and characterization of pre-core mutants of hepatitis B virus (HBV) in chronically infected patients]

Hepatitis B virus infection is responsible of an important number of world morbimortality. This is associated to severe liver outcome such as cirrhosis and hepatocellular carcinoma. Diverse factors influence the severity of liver injury produced by Hepatitis B Virus (HBV): the emergence of HBV mutants unable to secrete [quot ]e[quot ] antigen (pre-core mutants), the virus and host genetic heterogenicity and immune system competence. Particularly, the presence of pre-core mutants in chronic carriers is associated a low response to interferon therapy. We present paper is to present evidence as to the presence of these mutants in chronically infected, HBeAg negative patients (HBsAg positive) in Argentina. Viral DNAs were extracted from sera of nine patients, amplified by PCR and characterized by restriction enzyme assay. All of them appear to be pre-core mutants according with serological markers and a very low level of viral DNA detected in serum. Further genetic characterization of one of them by nucleotide sequence analysis of the pre-core region let allowed us to show modifications at codon 15 and 28 both of them previously described for pre-core mutants.