Santiago Mirazo

Transmission, diagnosis, and management of hepatitis E: an update

Hepatitis E virus (HEV) infection is an important public health concern in many developing countries, causing waterborne outbreaks as well as sporadic autochthonous hepatitis. HEV is mainly transmitted by the fecal–oral route in endemic areas through drinking of contaminated water. However, zoonotic transmission from animal reservoirs to humans has also been suggested. Three additional routes of HEV transmission have been proposed to occur: blood borne, human to human, and vertical transmission from mother to child. Acute HEV infection is usually diagnosed by detecting specific anti-HEV antibodies. However, the performance of the available assays in different settings is not optimal. Analysis of HEV ribonucleic acid in biologic specimens such as stools, serum, and liver biopsy by using nucleic acid amplification techniques is also employed. Nonetheless, additional consensus regarding the best technologies suitable for serosurveys and diagnosis of acute HEV infection is also needed. This review article summarizes the current status of HEV infection end epidemiology with particular emphasis in transmission, diagnosis, and clinical management.

Molecular analysis of Porcine Circovirus Type 2 strains from Uruguay: evidence for natural occurring recombination.

Porcine Circovirus Type 2 (PCV2) is a worldwide distributed virus and is considered an important emerging pathogen related to several distinct disease syndromes in pigs. Genomic structure consists of three major open reading frames (ORFs). ORF1 (rep gene) encodes replication-related proteins, ORF2 (cap gene) encodes the capsid protein and ORF3 encodes a protein putatively involved in virus-induced apoptosis. Based on cap gene sequences, PCV2 strains are classified into two main genotypes, PCV2a with five clusters (2A-2E) and PCV2b with three clusters (1A-1C). According to previous theoretical studies, PCV2 strains can eventually undergo intra and inter-genotype recombination, mainly within the rep gene. Ever since, several evidences of recombination in the field have been reported and confirmed this hypothesis. In South America, data regarding molecular characterization of PCV2 strains is still scant. Genotyping studies in the region have concluded that PCV2b is the predominant circulating genotype in the region and till now, no recombinant strains have ever been reported. In this work we thoroughly characterized at the molecular level Uruguayan PCV2 strains by extensive sequence data analysis. Moreover, recombination software tools were applied to explore and characterize eventual occurrence of natural recombination events. Two recombinant PCV2 strains were detected in this study, as a consequence of an inter-genotype recombination event between PCV2b-1A and PCV2a-2D, as the major and minor parent, respectively. According to recombination software analysis, in both cases the event occurred within the ORF1. Herein, extensive viral sequence dataset is provided, including the characterization of the first PCV2 recombinant strains ever reported in South America. Additionally, our results suggested a multi-centered source of PCV2 infection in Uruguay, which probably involved Brazilian and European origins.

Genetic heterogeneity and subtyping of human Hepatitis E virus isolates from Uruguay

Hepatitis E virus (HEV) infection is an important public health concern in many developing countries causing waterborne outbreaks, as well as sporadic autochthonous hepatitis. It is transmitted primarily by the fecal-oral route. However, zoonotic transmission from animal reservoirs to human has also been suggested. Genotype 3 is the most frequent genotype found in South America and the HEV epidemiology in this region seems to be very complex. However, data about the molecular characterization of HEV isolates of the region is still lacking and further investigation is needed. Our study characterized human HEV strains detected in a 1-year period in Uruguay, by extensive sequence analysis of three regions of the HEV genome. Uruguayan strains were closely related to a set of European strains and in turn, were dissimilar to Brazilian, Argentinean and Bolivian isolates. Additionally, the co-circulation of viral subtypes 3i and 3h was observed. Circulation of subtype 3i had been reported in Argentina and Bolivia whereas sequences of subtype 3h are rare and had never been reported in Latin America. In order to contribute to shedding light over the molecular epidemiology of this emergent infection in the region, we thoroughly analyzed the genetic variability of HEV strains detected in Uruguay, providing the largest dataset of sequences of HEV ever reported in a country in South America.

Detection of all four human metapneumovirus subtypes in nasopharyngeal specimens from children with respiratory disease in Uberlândia, Brazil

The human metapneumovirus (hMPV) is a pathogen of the respiratory tract identified first in the Netherlands in 2001 and since then it has been detected worldwide. The purpose of this study was to identify and characterize hMPV in samples collected from children <5 years presenting with acute respiratory disease (ARD) seen at a public hospital in Uberlândia, in Southeastern Brazil. One hundred fourteen nasopharyngeal aspirates (NPAs) samples that were negative for the presence of nine other respiratory viruses were tested by reverse transcription polymerase chain reaction (RT‐PCR) for the presence of hMPV RNA. Fourteen out of 114 (12.3%) samples were positive for presence of hMPV RNA. PCR products, obtained by the amplification of partial nucleotide sequence of gene N, were sequenced and compared with sequences deposited in GenBank. Sequences from eight samples were obtained and all four subtypes were identified. Also, the recently proposed sublineages “a” and “b” of subtype A2 were found; mean age was 21 months old; upper respiratory tract infection (URTI) was the most common clinical symptom; the virus was detected in samples collected from March to November, a period that corresponds to late summer to mid‐spring in Brazil. This is the first study to describe the circulation of all hMPV subtypes in Minas Gerais state. J. Med. Virol. 81:1814–1818, 2009.

Detection and molecular characterization of sporadic cases of acute human hepatitis E virus infection in Uruguay.

In developing countries, hepatitis E virus (HEV) infection is a public-health concern because it causes epidemics and waterborne outbreaks. In South America, few HEV strains have been characterized at the molecular level. We report the detection and molecular analysis of the first set of sporadic cases of autochthonous human genotype 3 HEV infection in Uruguay.

Viral quasispecies profiles as the result of the interplay of competition and cooperation

BackgroundViral quasispecies can be regarded as a swarm of genetically related mutants. A common approach employed to describe viral quasispecies is by means of the quasispecies equation (QE). However, a main criticism of QE is its lack of frequency-dependent selection. This can be overcome by an alternative formulation for the evolutionary dynamics: the replicator-mutator equation (RME). In turn, a problem with the RME is how to quantify the interaction coefficients between viral variants. Here, this is addressed by adopting an ecological perspective and resorting to the niche theory of competing communities, which assumes that the utilization of resources primarily determines ecological segregation between competing individuals (the different viral variants that constitute the quasispecies). This provides a theoretical framework to estimate quantitatively the fitness landscape.ResultsUsing this novel combination of RME plus the ecological concept of niche overlapping for describing a quasispecies we explore the population distributions of viral variants that emerge, as well as the corresponding dynamics. We observe that the population distribution requires very long transients both to A) reach equilibrium and B) to show a clear dominating master sequence. Based on different independent and recent experimental evidence, we find that when some cooperation or facilitation between variants is included in appropriate doses we can solve both A) and B). We show that a useful quantity to calibrate the degree of cooperation is the Shannon entropy.ConclusionsIn order to get a typical quasispecies profile, at least within the considered mathematical approach, it seems that pure competition is not enough. Some dose of cooperation among viral variants is needed. This has several biological implications that might contribute to shed light on the mechanisms operating in quasispecies dynamics and to understand the quasispecies as a whole entity.

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.

Distribution of HPV-16 variants among isolates from Paraguayan women with different grades of cervical lesion

To determine the distribution of HPV‐16 variants among Paraguayan women with different grades of cervical lesions.

New insights into the hepatitis E virus genotype 3 phylodynamics and evolutionary history

Hepatitis E virus (HEV) is an emergent hepatotropic virus endemic mainly in Asia and other developing areas. However, in the last decade it has been increasingly reported in high-income countries. Human infecting HEV strains are currently classified into four genotypes (1-4). Genotype 3 (HEV-3) is the prevalent virus genotype and the mostly associated with autochthonous and sporadic cases of HEV in developed areas. The evolutionary history of HEV worldwide remains largely unknown. In this study we reconstructed the spatiotemporal and population dynamics of HEV-3 at global scale, but with particular emphasis in South America, where case reports have increased dramatically in the last years. To achieve this, we applied a Bayesian coalescent-based approach to a comprehensive data set comprising 97 GenBank HEV-3 sequences for which the location and sampling date was documented. Our phylogenetic analyses suggest that the worldwide genetic diversity of HEV-3 can be grouped into two main Clades (I and II) with a Ƭmrca dated in approximately 320years ago (95% HPD: 420-236years) and that a unique independent introduction of HEV-3 seems to have occurred in Uruguay, where most of the human HEV cases in South America have been described. The phylodynamic inference indicates that the population size of this virus suffered substantial temporal variations after the second half of the 20th century. In this sense and conversely to what is postulated to date, we suggest that the worldwide effective population size of HEV-3 is not decreasing and that frequently sources of error in its estimates stem from assumptions that the analyzed sequences are derived from a single panmictic population. Novel insights on the global population dynamics of HEV are given. Additionally, this work constitutes an attempt to further describe in a Bayesian coalescent framework, the phylodynamics and evolutionary history of HEV-3 in the South American region.