Hepatitis A virus and environmental quality indicators in aquatic ecosystems for oyster farming in the Northeast of the State of Pará, Brazil

Authors: José Raul Rocha de Araújo Júnior; Denise Suéllen Amorim de Sousa Santos; Mônica Cristina de Moraes Silva; Dielle Monteiro Teixeira; Elivam Rodrigues Vale; Walber Victor de Moraes Pinto
DOI
10.5281/zenodo.3264045
DIN
IJOEAR-JUN-2019-10
Abstract

Research into the occurrence of microbiological contaminants, including hepatitis A virus (HAV), in river waters intended for oyster farming is of extreme importance for public health. This study aimed to detect the occurrence of HAV in the aquatic environment for ostreiculture in northeastern Pará, Brazil, and correlate with microbiological, physico-chemical and climatological variables. The HAV research was based on the method of water concentration by filtration membrane adsorption-elution and in the organic flocculation method with skim milk, followed by Nested-PCR. Quantification of coliforms, Enterococci and heterotrophic bacteria was performed. The physico-chemical variables were measured with multiparametric probe and spectrophotometry. Positive samples were purified and submitted to sequencing. From March 2017 to December 2018, 203 samples of river water were collected and analyzed in the municipalities of Augusto Corrêa, Curuçá, Salinópolis and São Caetano de Odivelas. In 10.8% of the analyzed samples the HAV RNA was detected, in all localities the HAV was classified in genotype IB. There was no significant difference between the concentration methods of the water samples. The only physicochemical variable that most influenced HAV detection was dissolved oxygen. Regarding the bacterial indicators, the highest statistical significance occurred with thermotolerant coliforms and Escherichia coli. The detection of HAV in the four municipalities studied shows that the virus is circulating in the aquatic environment and, therefore, in the community. In this context, more effective hygienic-sanitary measures are necessary in these communities dedicated to oyster farming.

Keywords
Hepatitis A Virus Quality Indicators Water and Ostreiculture.
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Introduction

The hepatitis A virus (HAV) belongs to the order of Picornavirales, family Picornaviridae, genus Hepatovirus, species Hepatovirus A [1]. It is an icosahedral symmetry virus, not enveloped, with a diameter of 27 to 32 nanometers. Classified in six genotypes, according to the phylogenetic analysis of the complete VP1 protein sequence. Genotypes I, II, and III are found in humans, being subdivided into subgenotypes IA and IB; IIA and IIB; IIIA and IIIB [2].

HAV is transmitted enterally by the ingestion of contaminated food and water, causing Hepatitis A, a self-limiting, infectious, symptomatic or asymptomatic disease of benign evolution, with the occurrence of fulminant cases. HAV infection is prevalent throughout the world, but with different epidemiology according to age of exposure and immunization [3].

With increasing pollution of the aquatic system, microorganisms that cause water-borne diseases can occur in oysters due to contamination by human waste or chemical pollution [4]. The quality of seawater where oysters are grown, as well as the animal itself, is of extreme importance for public health, since food and / or water contaminated by pathogens are the main causes of the occurrence of gastrointestinal diseases in Brazil [5]. Due to the various contaminants and forms of contamination of the aquatic environment, it is necessary to use quality micro-organisms that ensure the absence of other pathogens. The most commonly used indicators are coliform bacteria, total coliforms, thermotolerant bacteria, and Escherichia coli. However, depending on the environment studied, heterotrophic bacteria and Enterococci can be used to evaluate water quality [6].

One of the most widely used methods in environmental virology is based on the adsorption of virus particles to the filter media by loading interaction and subsequent elution of the virus by a pH adjusted solution [7]. Another method of viral concentration is organic flocculation, where an alkaline proteinaceous solution containing glycine with meat extracts or skimmed milk is used, promoting the recovery of virus particles adsorbed on the flakes [8].

After the concentration processes, molecular tests are carried out by PCR techniques, which has allowed advances in the detection of enteric viruses. However, the assay is susceptible to inhibitors found in aquatic environments [9]. The higher the level of water pollution, the lower the detection efficiency of the viral genome, the presence of particulate matter or suspended solids in water [10].

The investigation of the contamination of water in relation to enteric viruses using molecular techniques may be impaired due to the large volume of water in the environment in relation to the low concentration of viruses in these environments, and also by the possible presence of inhibitors of enzymatic reactions that can be found in these samples that compromise the detection of viral genomes [9].

This study aimed to detect the occurrence of HAV in the aquatic environment for ostreiculture in northeastern Pará, Brazil, and correlate with microbiological, physico-chemical and climatological variables.

Conclusion

The detection of HAV, genotype IB, in the four municipalities studied shows that the virus is circulating in the aquatic environment and, therefore, in the community. We suggest the use of more than one concentration method with a view to reducing false-negative results, since even though no significant difference was observed between the concentration methods of the water samples for HAV detection, in only two the results were concordant.

The results obtained in this research are important to evaluate the occurrence and concentration of microbiological and physicochemical contaminants in oyster farming areas in the Northeast of Para. In this context, it is necessary to make more hygienic-sanitary measures in the communities dedicated to the cultivation of this food so that it meets the minimum standards established in current legislation, aiming at the expansion of oyster farming in the region and greater sanitary security for consumers.
 

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