Proposal of a pathway for enteric virus groups detection as indicators of faecal contamination to enhance the evaluation of microbiological quality in freshwater in Argentina.

An environmental survey was conducted in order to assess the frequency of detection of picobirnavirus (PBV), human adenovirus (HAdV) and infective enterovirus (iEV) as indicators of faecal contamination in freshwater, and to determine their potential as reporters of the presence of other enteric viruses, such as group A rotavirus (RVA). The study was carried out over a three-year period (2013-2015) in the San Roque Dam, Córdoba, Argentina. The overall frequency detection was 62.9% for PBV, 64.2% for HAdV and 70.4% for iEV. No significant differences were observed in the rates of detection for any of these viruses through the years studied, and a seasonal pattern was not present. Whenever there was RVA detection in the samples analyzed, there was also detection of iEV and/or HAdV and/or PBV. At least one of the viral groups analyzed was demonstrated in the 100% of the samples with faecal coliforms values within the guideline limits. In this setting, especially in those samples which reveal faecal indicator bacteria within the guideline limit, we propose to carry out a pathway, involving PBV, HAdV and iEV detection in order to enhance the evaluation of microbiological quality in freshwater in Argentina. The proposed methodological strategy could report faecal contamination in water, mainly of human origin, and the condition of the matrix to maintain viral viability. In addition, the viral groups selected could report the presence of RV.

Tracking enteric viruses in green vegetables from central Argentina: potential association with viral contamination of irrigation waters.

Consumption of green vegetable products is commonly viewed as a potential risk factor for infection with enteric viruses. The link between vegetable crops and fecally contaminated irrigation water establishes an environmental scenario that can result in a risk to human health. The aim of this work was to analyze the enteric viral quality in leafy green vegetables from Córdoba (Argentina) and its potential association with viral contamination of irrigation waters. During July-December 2012, vegetables were collected from peri-urban green farms (n = 19) and its corresponding urban river irrigation waters (n = 12). Also, urban sewage samples (n = 6) were collected to analyze the viral variants circulating in the community. Viruses were eluted and concentrated by polyethylene glycol precipitation and then were subject to Reverse Transcription Polymerase Chain Reaction to assess the genome presence of norovirus, rotavirus and human astrovirus. The concentrates were also inoculated in HEp-2 (Human Epidermoid carcinoma strain #2) cells to monitor the occurrence of infective enterovirus. The frequency of detection of the viral groups in sewage, irrigation water and crops was: norovirus 100%, 67% and 58%, rotavirus 100%, 75% and 5%, astrovirus 83%, 75% and 32% and infective enterovirus 50%, 33% and 79%, respectively. A similar profile in sewage, irrigation water and green vegetables was observed for norovirus genogroups (I and II) distribution as well as for rotavirus and astrovirus G-types. These results provide the first data for Argentina pointing out that green leafy vegetables are contaminated with a broad range of enteric viruses and that the irrigation water would be a source of contamination. The presence of viral genomes and infective particles in food that in general suffer minimal treatment before consumption underlines that green crops can act as potential sources of enteric virus transmission. Public intervention in the use of the river waters as irrigation source is needed.

Enteric Viruses in Surface Waters from Argentina: Molecular and Viable-Virus Detection.

Water resources contaminated with wastewater are an important source for the dissemination of enteric viruses with an impact on the health of the population. The aim of the study was to assess the viral contamination of freshwater from a dam in Argentina by using infectious enterovirus detection, viral RNA amplification, and a genetic characterization of five enteric viruses associated with diarrhea and hepatitis. Enterovirus infectivity (iEV) was evaluated by cell culture and direct immunofluorescence. The detection of the viral genome of rotavirus (RV), human astrovirus (HAstV), norovirus (NoV), hepatitis A virus (HAV), and hepatitis E virus (HEV) was performed by reverse transcriptase PCR (RT-PCR). A total of 48 water samples from 4 monitoring points on the body of the dam from January to December 2012 and 66 water samples from 3 tourist beaches on the edge of the dam from October 2013 to October 2015 were collected monthly. During the first period, the overall viral frequency detection was 52.1% for group A RV, 50% for HAstV, 60.4% for NoV, 22.9% for HAV, 2.1% for HEV, and 64.6% for iEV. The overall frequency detection for the second sampling was 18.2% for RV and HAstV, 31.8% for NoV, 7.57% for HEV, and 66.7% for iEV. There was no detection of HAV during this period. The genotypes and genogroups detected through the study correlated with the most common genomic variants associated with human gastrointestinal and hepatitis illnesses. The results obtained could alert the health systems and environmental sanitation to make decisions for viral control and prevention in our environment.IMPORTANCE The study shows the impact of anthropic contamination of one of the most important tourist water resources in Argentina. This course of recreational water would be a favorable scenario for infection, as well as a reservoir for the enteric viruses, creating a risk for the population exposed to these waters. The results obtained could alert the health systems and environmental sanitation to make decisions for the control and prevention of viral diseases in this environment.

Quantification of human infection risk caused by rotavirus in surface waters from Córdoba, Argentina.

Fecal contamination of water is a worrying problem because it is associated with the transmission of enteric pathogenic microorganisms that can cause many infectious diseases. In this study, an environmental survey was conducted to assess the level of viral contamination by viable enterovirus and rotavirus genome in two recreational rivers (Suquía and Xanaes) of Córdoba, Argentina. Quantitative microbial risk assessment (QMRA) was calculated to estimate the risk of rotavirus infection. Water sampling was carried out during a one-year period, the presence of total and fecal coliforms was determined and water samples were then concentrated for viral determination. Cell culture and indirect immunofluorescence were applied for enterovirus detection and RT-qPCR for rotavirus quantification. Coliform bacteria levels found in Suquía River often far exceeded the guideline limits for recreational waters. The Xanaes exhibited a lower level of bacterial contamination, frequently within the guideline limits. Enterovirus and rotavirus were frequently detected in the monitoring rivers (percentage of positive samples in Suquía: 78.6% enterovirus, 100% rotavirus; in Xanaes: 87.5% enterovirus, 18.7% rotavirus). Rotavirus was detected at a media concentration of 5.7×10(5) genome copies/L (gc/L) in the Suquía and 8.5×10(0)gc/L in the Xanaes. QMRA revealed high risk of rotavirus infection in the Suquía, at sampling points with acceptable and non-acceptable bacteria numbers. The Xanaes showed significantly lower health risk of rotavirus infection but it proved to be a public health hazard. The viral occurrence was not readily explained by the levels of bacteria indicators, thus viral monitoring should be included to determine microbiological water quality. These findings provide the first data of QMRA for recreational waters in Argentina and reveal the need for public awareness of the health implications of the use of the river waters.

Rotavirus seasonality in urban sewage from Argentina: effect of meteorological variables on the viral load and the genetic diversity.

In Argentina, the rotavirus disease exhibits seasonal variations, being most prevalent in the fall and winter months. To deepen the understanding of rotavirus seasonality in our community, the influence of meteorological factors on the rotavirus load and the genetic diversity in urban raw sewage from Córdoba city, Argentina were evaluated. Wastewater samples were collected monthly during a three-year study period and viral particles were concentrated by polyethylene glycol precipitation. RT-nested PCR was applied for rotavirus detection, and VP7/VP4 characterization and real-time PCR for rotavirus quantification. Both molecular techniques showed relatively similar sensitivity rates and revealed rotavirus presence in urban wastewater in cold and warm seasons, indicating its circulation in the local community all year round. However, a slight trend for rotavirus circulation was noted by real-time PCR in the fall and winter seasons, showing a significantly higher peak of rotavirus concentration at mean temperatures lower than 18°C and also higher, although not statistically different during drier weather. VP7 and VP4 gene characterization showed that G1 and P[8] genotypes were dominant, and temporal variations in genotype distribution were not observed. Rotavirus spread is complex and our results point out that weather factors alone cannot explain the seasonal quantitative pattern of the rotavirus disease. Therefore, alternative transmission routes, changes in human behavior and susceptibility, and the stability and survivability of the virus might all together contribute to the seasonality of rotavirus. The results obtained here provide evidence regarding the dynamics of rotavirus circulation and maintenance in Argentina.

Distribution of genetic polymorphisms associated with hepatitis C virus (HCV) antiviral response in a multiethnic and admixed population.

The prevalence of genetic polymorphisms identified as predictors of therapeutic-induced hepatitis C virus (HCV) clearance differs among ethnic groups. However, there is a paucity of information about their prevalence in South American populations, whose genetic background is highly admixed. Hence, single-nucleotide polymorphisms rs12979860, rs1127354 and rs7270101 were characterized in 1350 healthy individuals, and ethnicity was assessed in 259 randomly selected samples. The frequency of rs12979860CC, associated to HCV treatment response, and rs1127354nonCC, related to protection against hemolytic anemia, were significantly higher among individuals with maternal and paternal Non-native American haplogroups (64.5% and 24.2%), intermediate among admixed samples (44.1% and 20.4%) and the lowest for individuals with Native American ancestry (30.4% and 6.5%). This is the first systematic study focused on analyzing HCV predictors of antiviral response and ethnicity in South American populations. The characterization of these variants is critical to evaluate the risk-benefit of antiviral treatment according to the patient ancestry in admixed populations.

Cocirculation of Rio Negro Virus (RNV) and Pixuna Virus (PIXV) in Tucumán province, Argentina.

Venezuelan equine encephalitis complex includes viruses considered emerging pathogens for humans and animals in the Americas. Two members of this complex have been detected previously in Argentina: Rio Negro Virus (RNV), detected in mosquitoes from Chaco province and rodents from Formosa province, and Pixuna Virus (PIXV), detected in mosquitoes from Chaco province. To carry out surveillance studies in other parts of the country, detection of a 195-bp fragment of alphaviruses by RT-nested PCR was performed in mosquito samples from San Miguel de Tucumán city. Four pools resulted positive and three were sequenced. Two amplicons grouped with RNV and one with PIXV. This is the first report of viral activity of members of the Venezuelan equine encephalitis complex in north-eastern Argentina.

An indirect immunofluorescence assay to detect antibodies against St. Louis Encephalitis virus.

An in house indirect immmunofluorescence assay ( IFA ) in relation to neutralization (NT) reference test, was assessed as a fast and cheap method to carry out serological surveys for St. Louis Encephalitis virus (SLE). Sera obtained from 213 blood donors were analyzed by both tests. The prevalence of seropositivity obtained with IFA was lower than (30.98%) that observed on NT (41.78%). The relative specificity rate of IFA was 96.77% whereas its relative sensitivity rate was 69.66%. Kappa index showed a good correlation between both tests. The results indicate that neutralization assay is still the serological test with the highest sensitivity and specificity relative rates for detecting antibodies against SLE virus. Nevertheless, the IFA could be useful as an alternative test in order to learn the circulation of the Flavivirus genus in a certain area.

[Seroprevalence of infection by St. Louis encephalitis virus in the Province of Formosa]. / Seroprevalencia de infección por el virus encefalitis San Luis en la Provincia de Formosa.

The aim of this work was to determine the prevalence of antibodies against St. Louis encephalitis virus (SLE) in human sera provided by the Laboratory of Epidemiological Surveillance from Formosa District (Province of Formosa, Argentina) in 1995 and 1997. The tests used for this study were hemagglutination inhibition (HI) and neutralization (NT). The screening performed by NT test showed prevalences of 21% (60/284) and 32% (50/157) of antibodies in samples obtained in 1995 and 1997 respectively. In 1995, 14% of tested sera showed low titer of neutralizing antibodies (NT) (1/20 and 1/40) whereas in 1997, 19% of the sera presented titers of NT antibodies equal or greater than 1/80. It was observed that sera with low titers of NT antibodies (1/20-1/40) resulted negative in HI in the simultaneous titration of antibodies by NT and HI whereas other sera presented high titers for both tests. This relation between the low and the high titers of antibodies indicates the presence of past and recent infections and the continuous circulation of this virus. Moreover, the prevalence of NT antibodies in the surveyed population increased significantly in 2 years (p < 0.0075) confirming the endemicity of this agent in this area and showing the need to perform studies of non-confirmed viral etiology febrile diseases to determine its importance in human pathogenicity.

Seroprevalencia de infección por el virus encefalitis San Luis en la Provincia de Formosa. / [Seroprevalence of infection by St. Louis encephalitis virus in the Province of Formosa]

The aim of this work was to determine the prevalence of antibodies against St. Louis encephalitis virus (SLE) in human sera provided by the Laboratory of Epidemiological Surveillance from Formosa District (Province of Formosa, Argentina) in 1995 and 1997. The tests used for this study were hemagglutination inhibition (HI) and neutralization (NT). The screening performed by NT test showed prevalences of 21

(60/284) and 32

(50/157) of antibodies in samples obtained in 1995 and 1997 respectively. In 1995, 14

of tested sera showed low titer of neutralizing antibodies (NT) (1/20 and 1/40) whereas in 1997, 19

of the sera presented titers of NT antibodies equal or greater than 1/80. It was observed that sera with low titers of NT antibodies (1/20-1/40) resulted negative in HI in the simultaneous titration of antibodies by NT and HI whereas other sera presented high titers for both tests. This relation between the low and the high titers of antibodies indicates the presence of past and recent infections and the continuous circulation of this virus. Moreover, the prevalence of NT antibodies in the surveyed population increased significantly in 2 years (p < 0.0075) confirming the endemicity of this agent in this area and showing the need to perform studies of non-confirmed viral etiology febrile diseases to determine its importance in human pathogenicity.