Sandra Frabasile

Genotypes of respiratory syncytial virus group B identified in Uruguay

Summary.Genotypes of Human respiratory syncytial virus (HRSV) of group B from Uruguay were assigned to strains isolated during 1999 and 2001 outbreaks and others formerly reported isolated in the period 1989–1996. The nucleotide sequences of the C-terminal portion of the G protein were compared to sequences representative of previously defined HRSV genotypes. Most Uruguayan strains clustered into five of the previously identified genotypes. Nine isolates clustered in two genotypes named URU1 and URU2 which were not described up to present. Two of the analyzed sequences isolated in 2001 have a six nucleotide duplication that is discussed in terms of HRSV variability.

Molecular epidemiology of human respiratory syncytial virus in Uruguay: 1985-2001 - A review

The variability of the G glycoprotein from human respiratory syncytial viruses (HRSV) (groups A and B) isolated during 17 consecutive epidemics in Montevideo, Uruguay have been analyzed. Several annual epidemics were studied, where strains from groups A and B circulated together throughout the epidemics with predominance of one of them. Usually, group A predominates, but in some epidemics group B is more frequently detected. To analyse the antigenic diversity of the strains, extracts of cells infected with different viruses of group A were tested with a panel of anti-G monoclonal antibodies (MAbs). The genetic variability of both groups was analyzed by sequencing the C-terminal third of the G protein gene. The sequences obtained together with previously published sequences were used to perform phylogenetic analyses. The data from Uruguayan isolates, together with those from the rest of the world provide information regarding worldwide strain circulation. Phylogenetic analyses of HRSV from groups A and B show a model of evolution analogous to the one proposed for influenza B viruses providing information that would be beneficial for future immunization programs and to design safe vaccines.

Cytotoxic, Virucidal, and Antiviral Activity of South American Plant and Algae Extracts

Herpes simplex virus type 1 (HSV-1) infection has a prevalence of 70% in the human population. Treatment is based on acyclovir, valacyclovir, and foscarnet, three drugs that share the same mechanism of action and of which resistant strains have been isolated from patients. In this aspect, innovative drug therapies are required. Natural products offer unlimited opportunities for the discovery of antiviral compounds. In this study, 28 extracts corresponding to 24 plant species and 4 alga species were assayed in vitro to detect antiviral activity against HSV-1. Six of the methanolic extracts inactivated viral particles by direct interaction and 14 presented antiviral activity when incubated with cells already infected. Most interesting antiviral activity values obtained are those of Limonium brasiliense, Psidium guajava, and Phyllanthus niruri, which inhibit HSV-1 replication in vitro with 50% effective concentration (EC50) values of 185, 118, and 60 μg/mL, respectively. For these extracts toxicity values were calculated and therefore selectivity indexes (SI) obtained. Further characterization of the bioactive components of antiviral plants will pave the way for the discovery of new compounds against HSV-1.

Molecular detection and genetic variability of human metapneumovirus in Uruguay.

Human metapneumovirus (hMPV) has been described as circulating among the Uruguayan population at least since 1998 based on serologic evidence. However, no isolation attempts, molecular detection, or genetic studies have been carried out so far in the country. In the present study, molecular detection of circulating hMPV in children hospitalized with acute respiratory tract infection in Montevideo–Uruguay was carried out by reverse transcription‐polymerase chain reaction (RT‐PCR) amplification of the hMPV nucleoprotein (N) gene from 217 nasopharyngeal aspirates. Genetic variability analysis of the positive samples was performed by amplification and sequencing of both N and attachment glycoprotein (G) genes. Eighteen of the 217 samples tested positive for hMPV, with tachypnea, chest indrawing, and wheezing being the main clinical symptoms recorded. Phylogenetic analysis of N and G genes showed that Uruguayan samples clustered in genotypes described previously as A2, B1, and B2, with bootstrap values ≥98%. Sublineages A2a and A2b could also be distinguished within the samples that belong to A2. This is the first molecular report on the circulation of hMPV in Uruguay. The pattern of circulation of this virus, analyzed for both N and G genes independently, resembles the complex evolutionary pattern of respiratory syncytial virus (RSV). J. Med. Virol. 82:861–865, 2010.

Selection and characterization of human respiratory syncytial virus escape mutants resistant to a polyclonal antiserum raised against the F protein

A human respiratory syncytial virus (HRSV) neutralization escape mutant was obtained after 56 serial passages in the presence of a polyclonal antiserum raised against the F protein. Nucleotide sequence analysis of this escape mutant virus revealed two amino acid substitutions: Asn268Ile and Val533Met. When this virus was allowed to grow in the absence of the anti-F polyclonal serum, only the mutation Asn268Ile was stably maintained. Both the double and single escape mutant viruses lost reactivity with mAbs belonging to antigenic site II of the fusion protein of RSV. Mutation Asn268Ile has already been reported in RS viruses that are resistant to mAbs 47F and 11 and palivizumab (PZ). We have thus identified a novel mutation (Val533Met) in the transmembrane domain of the F protein that was selected under immune pressure.

Genomic Characterization and Seroprevalence Studies on Alphaviruses in Uruguay

Alphaviruses (Togaviridae) are arboviruses frequently associated with emerging infectious diseases. In this study, we aimed to investigate the presence of alphaviruses in Uruguay by detecting the viral genome in mosquitoes and neutralizing antibodies in equines. A total of 3,575 mosquitoes were analyzed for alphavirus genome detection. Serologic studies were performed on 425 horse sera by plaque reduction neutralization test (PRNT80) against Venezuelan equine encephalitis virus (VEEV) subtype IAB, Pixuna virus (PIXV), Rio Negro virus (RNV), western equine encephalitis virus (WEEV), and Madariaga virus (MADV). Mosquitoes belonging to six genera were captured and 82.9% were identified as Culex pipiens. Two Cx. pipiens pools collected in Fray Bentos and Las Toscas localities were alphavirus positive, and phylogenetic analyses showed that the sequences grouped into two different clusters: the lineage I of eastern equine encephalitis virus and RNV (VEEV complex), respectively. Plaque reduction neutralization test assays showed antibodies against strains of the VEEV complex, MADV, and WEEV. Rio Negro virus was the most geographically widespread virus, showing higher seroprevalences (up to 20%). Seroprevalences against VEEV IAB ranged between 4.6% and 13%; antibodies against PIXV, WEEV, and MADV were less frequent (3-4%). In conclusion, RNV exhibited the highest seroprevalence in horses, a wide geographical distribution, and viral genome was detected in Cx. pipiens mosquitoes. Madariaga virus had a low seroprevalence in equines, but an epizootic lineage typical of North America was detected in Cx. pipiens mosquitoes. Taken together, our results show that alphaviruses are present in Uruguay with variable occurrence and geographical distribution being a potential threat for human and equine health.