Food and Environmental Virology

https://doi.org/10.1007/s12560-018-9357-1

ORIGINAL PAPER

Hepatitis A Virus, Hepatitis E Virus, and Rotavirus in Foods of Animal

Origin Traded at the Borders of Brazil, Argentina, and Uruguay
Juliano Gonçalves Pereira1,2   · Vanessa Mendonça Soares3 · Fernanda Gil de Souza4 · Leonardo Ereno Tadielo3 ·
Emanoelli Aparecida Rodrigues dos Santos3 · Mário Celso Sperotto Brum3 · Andreia Henzel4 ·
Eduarda Hallal Duval2 · Fernando Rosado Spilki4 · Wladimir Padilha da Silva2

Received: 30 July 2018 / Accepted: 6 September 2018

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Abstract
The aim of this study was to investigate hepatitis A virus (HAV), hepatitis E (HEV), and rotavirus (RV) in fresh and pro-
cessed meat traded on the border of Brazil with Argentina and Uruguay. In total, 159 samples of raw and processed foods of
animal origin were collected in Paso de los Libres, Argentina (n = 53 raw meat, n = 24 processed meat) and Rivera, Uruguay
(n = 55 raw meat, n = 18 processed meat), or were seized by the Brazilian International Agricultural Surveillance System—
VIGIAGRO (Brazil–Argentina border) (n = 8 raw meat, n = 1 bush meat). All samples were tested for the presence of HAV,
HEV, and RV genomes. HAV genes were detected in 18.23% of samples and RV genes in 23.89%. No HEV-positive samples
were detected. HAV was also detected in two of the VIGIAGRO samples. Processed meats from Argentina and Uruguay had
a higher rate of HAV and RV than raw meat (P > 0.05). The median HAV in the Argentinian and Uruguayan samples was
6.9 × 104 and 3.5 × 103 copies/g, respectively. The presence of RV viral genes in raw meats from Argentina was significant,
and this was not observed in processed meats. The presence of HAV and RV genes in a significant portion of products from
Argentina and Uruguay is a potential source of human infection. This also indicates precarious conditions of acquisition,
processing, and manipulation, which could be improved by improved regulation of food across borders.

Keywords  Contamination · HAV · HEV · RV · Transboundary disease

Introduction

Foodborne diseases are an important public health problem

* Juliano Gonçalves Pereira
juliano.pereira@unesp.br
* Wladimir Padilha da Silva
wladimir@pq.cnpq.br
1
Departamento de Higiene Veterinária e Saúde Pública,
Faculdade de Medicina Veterinária e Zootecnia,
Universidade Estadual Paulista “Júlio de Mesquita Filho”,
Campus de Botucatu, Rua Prof. Walter Maurício Correa, SN,
Botucatu, SP CEP 18618681, Brazil
2
Universidade Federal de Pelotas, Campus Capão do
Leão. Avenida Eliseu Maciel, SN, Capão do Leão,
RS CEP 96010900, Brazil
3
Universidade Federal do Pampa, Campus Uruguaiana. BR
472, Km 585, Uruguaiana, RS CEP 97501970, Brazil
4
Laboratório de Microbiologia Molecular, Instituto de
Ciências da Saúde, Universidade Feevale, Rodovia ERS‑239,
2755, Novo Hamburgo, RS CEP 93525075, Brazil
worldwide. From 2000 until 2017, 12,503 outbreaks were
reported to the Brazilian Health Ministry. These involved
at least 236,403 individuals and resulted in 182 deaths. The
etiologic agents associated with these outbreaks included
6.0% Hepatitis A (HAV), Rotavirus (RV), and Norovirus
(NV) (Brasil 2018). However, these numbers may be under-
estimated, owing to difficulties related to diagnosis, notifi-
cation, and identification of the agents. In the United States
of America, between 2000 and 2015, there were 5165 out-
breaks caused by these pathogens, involving 131,682 indi-
viduals and causing 23 deaths (Centers for Disease Control
and Prevention 2017).
HAV and hepatitis E (HEV) viruses are the major causa-
tive agents of acute hepatitis in humans, and RV is one of
the most important agent associated with infantile diarrhea
worldwide. Besides the differences observed in the clini-
cal manifestations of these viruses, both are transmitted

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through the oral–fecal route, person-to-person contact, or already confirmed the presence of pathogenic bacteria (Melo
contaminated food. Therefore, hygienic procedures during et al. 2014a, 2015). However, no studies have investigated
processing and handling are an important barrier to viral the presence of a virus of public health relevance in products
contamination of foods (White et al. 2016). that transit in border areas of the country. Therefore, the aim
Globalization of the food market, ease of international of this study was to evaluate the presence of HAV, HEV, and
travels, and the illegal trade of animal products all contrib- RV in animal products, both raw and processed, traded in the
ute to the increased illnesses caused by food-borne patho- border areas of Brazil, Argentina, and Uruguay.
gens (Käferstein et al. 1997). To reduce the risk of pathogen
dissemination, the World Organization for Animal Health
(OIE) advocates that animals and foods of animal origin Materials and Methods
must be forbidden to flow among countries without approval
and the fulfillment of rigid national and international sani- Sample Collection
tary guidelines. This is because they can quickly disseminate
pathogens, directly affecting consumer and animal health This study was carried in Rio Grande do Sul (RS), a state
(World Organization for Animal Health 2016). Since the located in southern Brazil that has an international bor-
OIE’s recommendations, it became the responsibility of der with Argentina and Uruguay. Samples were collected
countries to set suitable measures for food and animal move- between September 2015 and November 2016, totaling 159
ment control. However, a failure in the surveillance services products of animal origin (raw and processed meat), with 77
at borders may compromise the population’s health (Hueston of these originating from Argentina, 73 from Uruguay, and
et al. 2011; Noordhuizen et al. 2013). nine seized by VIGIAGRO-MAPA (Table 1).
The border surveillance in Brazil is in charge of the Inter- The samples obtained from VIGIAGRO (n = 9) were col-
national Agricultural Surveillance (VIGIAGRO), a Ministry lected during an inspection of baggage at the International
of Agriculture, Livestock, and Food Supply (MAPA) that Bridge Getúlio Vargas—Augustín Pedro Justo (29°45′18″S,
operates in more than a hundred units located in ports, air- 57°05′16″O). This bridge connects the cities of Uruguai-
ports, and borders posts. The Brazilian border surveillance ana (Brazil) and Paso de los Libres (Argentina). Among
inspects live animals and animal products as well as checks these samples, eight were raw meat and one was bush meat
travelers’ baggage (Brasil 2006a). (Hydrochoerus hydrochaeris—capybara) transported by
Currently, MAPA has authorized the entry of animal travelers who were entering Brazil.
products bought abroad and internalized via airports or
border posts, provided that some requirements are met: the
amount per person must characterize personal consumption
(10 kg of processed meat products and 5 kg of dairy prod- Table 1  Foods evaluated at the Brazil–Argentina–Uruguay border for
the presence of hepatitis A virus (HAV), hepatitis E virus (HEV), and
ucts, egg, or fish processed products); and original package, rotavirus (RV)
sealed and with no signs of leakage or violations. These
requirements must be checked the moment the travelers Foods From Total
arrive in Brazil, although many times the surveillance does Argentina Uruguay
not happen or, when it does, it is not efficient in detecting
Raw meats
irregularities. This allows the illegal entry of foods that do
 Bovinea 30 27 57
not meet the requirements, such as fresh meat or higher
 Swine 13 17 30
amounts than recommended (Brasil 2016).
 Chicken 18 11 29
The entry of non-inspected food into the country is
 Bush ­meatb 1 0 1
alarming because there is no guarantee that foods are in
 Total 62 55 117
conditions fit for consumption. Furthermore, such foods
Processed meats
can carry exotic pathogens or introduce new variants due
 Mortadela 5 3 8
to many unknown factors such as the origin, the location
 Sausage 8 4 12
and method of slaughter, the industrialization type, whether
 Salami 9 5 14
they meet international sanitary requirements, or whether the
 Ham 2 2 4
food product was subjected to any kind of official sanitary
 Pâté 0 4 4
inspection in its originating country (Brasil 2006b).
 Total 24 18 42
Bacterial and viral pathogens have been detected in ani-
Overall Total 86 73 159
mal products detained from travelers in airports around the
world (Beutlich et al. 2015; Rodríguez-Lázaro et al. 2015). a
 Eight samples were seized at VIGIAGRO
In Brazil, studies on products seized in the airports have b
 Sample seized next to VIGIAGRO

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Food and Environmental Virology
The remaining samples (n = 150) were obtained
through sampling in establishments located in Paso de Los
Libres, Province of Corrientes, Argentina (29°43′00″S,
57°05′00″O), and Rivera Department, Uruguay (30°54′09″S,
55°33′02″O). The samples were authorized and inspected
by VIGIAGRO. The products were acquired from 16 com-
mercial establishments (supermarkets, grocery stores, and
butcheries), eight in each city (Argentina – establishments
A to H; Uruguay—establishments I to P). In each location,
three samples were collected at distinct times. All samples
were packed in plastic bags and sent to the laboratory in
isothermal boxes containing recyclable ice. Upon arrival at
the laboratory, they were maintained at 4 °C until analysis.
HAV, HEV, and RV Detection
Samples Preparation, RNA Extraction, and cDNA Synthesis
The samples (1 g) were minced with 1 mL of 1X Mini-
mum Essential Medium (MEM, pH 7.0) and homogenized
by vortexing for 10 s and submitted to RNA extraction by
the TRIzol™ reagent method (Invitrogen) according to the
manufacturer´s instructions with minor modifications. After
homogenization with 1X MEM, 250 µL was suspended in
750  µL of TRIzol™ reagent (Invitrogen), incubated for
5 min, and centrifuged at 11,000×g for 10 min at 4 °C. The
supernatant was transferred to a tube containing 200 µL of
chloroform, incubated at 21 °C for 5 min, and centrifuged at
11,000×g for 10 min at 4 °C, with separation of the aqueous
phase to a new tube. The proteins were precipitated with
500 µL of isopropanol, and then incubated at room tempera-
ture for 5 min. Subsequently, the samples were centrifuged
on 12,000×g for 8 min and the supernatant was disposed. At
the end of the process, 1 mL of 75% ethanol (v/v) was added,
with a final centrifugation at 9000×g for 5 min. The ethanol
layer was removed by inversion and dried for 3 min. The
RNA pellets were suspended in 60 µL of TE buffer solution
and stored at − 80 °C.
For cDNA preparation, 10 µL of total RNA was added
to 10 µL of master mix (High Capacity cDNA Synthesis—
Applied Biosciences), following the manufacturer’s instruc-
tions. Each solution mix consisted of 3.2 µL of DNase/
RNase free water, 2 µL of buffer, 0.8 µL of dNTP, 2 µL of
random primers, 1 µL of RNase inhibitor, and 1 µL of RT
enzyme. The samples were amplified in a thermal cycler
(10 min at 20 °C, 120 min at 37 °C, 5 min at 85 °C) and then
refrigerated at 4 °C.
RT‑qPCR for HAV Detection
In real-time quantitative reverse transcription PCR (RT-
qPCR), primers to the non-coding region (5′-UTR) of the
viral genome were used, in accordance with the study of
Jothikumar et al. (2005). The qPCR was performed using the
TaqMan Universal PCR Master Mix Kit (Applied Biosys-
tems), following the manufacturer’s instructions. The reac-
tion volume was 20 µL and 100 nM of each primer and probe
were used. The thermal cycle involved initial denaturation
at 95 °C for 15 min, followed by 40 cycles of 95 °C for 10 s,
55 °C for 20 s, and 70 °C for 15 s.
The qPCR was carried out in 96-well plates with posi-
tive and negative controls (ultrapure water free from DNase/
RNase). The standard curve was prepared from serial dilu-
tions from plasmid PCR products (positive control). Master
Mix (15 µL) and cDNA (5 µL) were added, and the plate
was then sealed and placed in the Multicolor Real-Time PCR
Detection System instrument (Bio-Rad). All the controls and
samples were tested in duplicate to establish the efficiency
and limit of the technique. A standard curve was obtained
with serial dilutions of the controls at concentrations of
­101–1010. From this curve, the efficiency obtained was of
103% with an R2 of 0.99.
RT‑Nested PCR for HEV Detection
HEV was detected using primers for the HEV ORF1 region
(Table 2), according to Heldt et al. (2016). RNA isolated
from monkey’s feces positive for HEV served as the posi-
tive control. The reaction had a final volume of 50  µL,
containing 25 µL of GoTaq Green Master Mix (Promega),
18 µL of water free from DNase/RNase, 1 µL of each primer
(100 nM), and 5 µL of cDNA. Amplification conditions were
as follows: an initial temperature of 95 °C for 5 min, fol-
lowed by 45 cycles of 95 °C for 30 s, 59 °C for 1 min, 72 °C
for 1 min, and 72 °C for 7 min for final elongation. The
amplified products were resolved with electrophoresis and
observed under UV light.
RT‑PCR for RV Detection
For RV detection, primers to the VP6 gene region (Spilki
et al. 2013) were used at the same volume and concentration
as the reagents used for HEV detection. The positive control
was RNA extracted from cell culture infected with the RV
vaccine. The amplification conditions were initial denatura-
tion at 94 °C for 5 min, followed by 35 cycles of 94 °C for
1 min, 54 °C for 1 min, 72 °C for 1 min, and a final elonga-
tion at 72.8 °C for 7 min. After the reactions, the amplified
products were exposed to electrophoresis and then evaluated
under UV light.
Data Analysis
The data are tabulated, and the results are expressed as the
frequency of positives for each pathogen analyzed. They
were then analyzed by the Chi-square test to verify the
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 Food and Environmental Virology

Table 2  Primers used for the detection of hepatitis A virus (HAV), hepatitis E virus (HEV), and rotavirus (RV)
Virus Name Sequence Size (pb) References

HAV 5′-UTR F 5′ GGT​AGG​CTA​CGG​GTG​AAA​C 3′ 100 Jothikumar et al. (2005)

5′-UTR R 5′ GCG​GAT​ATT​GGT​GAG​TTG​TT 5′
Probe (position 413–441) 5′ CTT​AGG​CTA​ATA​CTT​CTA​TGA​AGA​GATGC 5′
HEVa HEVORF1con-s1 5′ CTG​GCA​TYA​CYC​TAC​TGC​YAT​TGA​GC 3′ 800 Heldt et al. (2016)
HEVORF1con-a1 5′ CCA​TCR​ARR​CAG​TAA​GTG​CGGTC 3′
HEVORF1con-s2 5′ CTG​CCY​TKGCG​AAT​GCT​GTG​G 3′ 287 Heldt et al. (2016)
HEVORF1con-a2 5′ GGC​AGW​RTA​CCA​RCG​CTG​AAC​ATC​ 3′
RV ROTAFEEVALE F 5′ GAT​GTC​CTG​TAC​TCC​TTG​T 3′ 160 Spilki et al. (2013)
ROTAFEEVALE R 5′ GGT​AGA​TTA​CCA​ATT​CCT​CC 3′
a
 Degenerate primers: Y = C or T; R = A or G; K = G or T; and W = A or T

association between the type of product and the detected
pathogen. The odds ratio (OR) was calculated to estimate the
chances of the virus’ genetic material being present in the
products analyzed. All statistical analyses were performed
using a statistic program (IBM SPSS Statistics Version 20)
using a significance level of 0.05.
Results
Among the 159 evaluated samples, 29 (18.23%) were
positive for the HAV genome and 38 (23.89%) for RV. No
HEV genetic material was detected in the samples ana-
lyzed (Table 3). The processed meats presented a higher
frequency of HAV and RV than raw meat. However, the
association was significant only for HAV (P < 0.05). The
OR for HAV detection in processed meat was 2.353 (IC95%
1.012–5.473).
Approximately 23% of products from Argentina were
positive for HAV and 36.04% for RV. Among the raw meat
samples that were positive for HAV, two were obtained
from VIGIAGRO. A higher frequency of HAV and RV was
detected in processed meats than in raw meats; however, this
association was not significant (P > 0.05). Among the fresh
meat samples, 19.35% were positive for HAV. However, in
processed meat samples, the frequency was 33.33%. The
frequency of RV in raw meat samples was 33.87% and in
processed meats 41.66%.
In the products originating from Uruguay, 12.32%
contained HAV and 9.58% had RV. In addition, 9.09% of

Table 3  Frequency and odds ratio (OR) for the presence of hepatitis A virus (HAV), hepatitis E virus (HEV), and rotavirus (RV) in products of
animal origin obtained from Argentina and Uruguay
Virus/food From Total P ­valuea OR (IC 95%)a
Argentina Uruguay

HEV 0/86 (0%) 0/73 (0%) 0/159 (0%)

HAV
 Raw meat 12/62 (19.35%) 5/55 (9.09%) 17/117 (14.52%) 0.116 0.417 (0.137–1.127)
 Processed meat 8/24 (33.33%) 4/18 (22.22%) 12/42 (28.57%) 0.506 0.571 (0.141–2.131)
 Total 20/86 (23.25%) 9/73 (12.32%) 29/159 (18.23%) 0.075 0.464 (0.197–1.095)
 P ­valueb 0.169 0.141 0.043
 OR (IC 95%)b 2.083 (0.724–5.995) 2.857 (0.675–12.085) 2.353 (1.012–5.473)
RV
 Raw meat 21/62 (33.87%) 4/55 (7.27%) 25/117 (21.36%) 0.001 0.153 (0.049–0.481)
 Processed meat 10/24 (41.66%) 3/18 (16.66%) 13/42 (30.23%) 0.083 0.280 (0.064–1.132)
 Total 31/86 (36.04%) 7/73 (9.58%) 38/159 (23.89%) 0.001 0.118 (0.77 − 0.460)
 P ­valueb 0.499 0.240 0.212
 OR (IC 95%)b 1.395 (0.530–3.668) 2.55 (0.513–12.679) 1.65 (0.749–3.633)
a
 P value and OR for comparison of the same type of product of different origin where P < 0.05 indicates association of the presence of the patho-
gen
b
 P value and OR for comparison of different products from the same origin where P < 0.05 indicates association of the presence of the pathogen

13
Food and Environmental Virology
raw meats and 22.22% of processed meats were positive
for HAV. However, there was no significant association
regarding the presence of this pathogen among the samples
(P > 0.05). Regarding RV, 7.27% of raw meats and 16.66%
of processed meats were positive (P > 0.05).
A significant association between the products from
Argentina and the presence of RV was observed, compared
to products from Uruguay. This association was noted
in the sum of the samples (P < 0.05; OR 0.118; IC95%
0.77–0.460), as in the raw meat products (P < 0.05; OR
0.153; IC95% 0.049–0.481). For HAV, there was no associa-
tion between the pathogen presence and the product source.
Among the 16 commercial establishment analyzed, 11
were positive for HAV or RV in at least one of the col-
lections. In Argentinian samples, the HAV and/or RV
genome was detected in samples from seven out of the eight
established samples evaluated: establishment A (n = 15),
B (n = 8), C (n = 7), D (n = 6), E (n = 3), F (n = 2), and G
(n = 1). In Uruguay, four establishments presented positive
samples (I = 6; J = 4; K = 3, and L = 2). Of the 159 samples
analyzed, eight presented simultaneous contamination by
HAV and RV, seven of them being from Argentina (three
from establishment A, three from B, and one from E) and
one from Uruguay (establishment J).
The median of HAV, as assessed by RT-qPCR, in sam-
ples from Argentina was 6.9 × 104  copies of genome/g,
with a minimum of 1.3 × 102 copies/g and a maximum of
3.2 × 10 13  copies/g. The HAV-positive samples (n = 2)
that were seized by VIGIAGRO presented scores of
3.2 × 1013 copies/g and 3.2 × 102 copies/g. In samples from
Uruguay, the median was 3.5 × 103 copies/g with a minimum
of 1.3 × 101 copies/g and a maximum of 2.9 × 109 copies/g.
Discussion
The presence of HAV and RV in a large number of animal
food products from various commercial establishments in
Argentina and Uruguay demonstrates the risk associated
with the consumption of these products that transit through
the international borders of the RS state of Brazil. The fact
that HAV and RV were detected in various establishments,
at different product sampling time points, indicates that
inadequate practices of obtaining/manipulating are routine
at those places.
The presence of viruses in food and their involvement
in food-borne disease outbreaks are associated with fecal
contamination (White et al. 2016). Therefore, food and water
used in many stages of processing may be contaminated
when sanitary–hygienic measures are not carefully imple-
mented (Victoria et al. 2014, Sa-Nguanmoo et al. 2015). In
this context, the informal trade of food (including illegal
import) is important in disseminating enteropathogens, such
as HAV and RV. Many activities involved in this practice
occur illegally, without the presence of sanitary inspection
authorities that would regulate the hygienic process of pro-
duction, trading, and transport of these products.
Currently, the import of some kinds of food is allowed
in Brazil (Brasil 2016). However, sanitary surveillance at
border posts must be rigorous to guarantee that food that has
the potential to disseminate pathogens is not introduced into
the country. Yet, even with sanitary surveillance at borders,
illegal import of animal products is a common practice in
many regions of Brazil, such as land borders (Pereira et al.
2017, 2018) or airports (Melo et al. 2014a, b, 2015). Specifi-
cally, in the border region of RS, it is estimated that more
than 60% of the travelers who go to Argentina or Uruguay
return to Brazil carrying some food of animal origin (fresh
or processed). Moreover, less than 10% of these travelers
have, at some point, passed by baggage inspection when
returning to Brazil (Pereira et al. 2017).
In the RS state, the main point of entry for these foods
is by land borders. However, in Brazil, illegal food is also
introduced through airports. In 2004, more than 65 tons of
illegal animal or plant products were seized in passengers’
luggage at three of the biggest Brazilian international air-
ports (Confins, in Belo Horizonte; Guarulhos, in São Paulo
and Salgado Filho, in Porto Alegre). The most seized prod-
ucts were of animal origin, such as dairy, sausages, fish,
honey, and meats. Seeds, seedlings, and fruits were the
most seized among the products derived from plants (Brasil
2015).
According to Melo et al. (2014b), 657.4 kg of animal
products that were illegally imported through the Guarul-
hos (São Paulo) and Galeão (Rio de Janeiro) airports were
seized by sanitary surveillance. This same study pointed
out that, between 2006 and 2009, around 40 tons of animal
products were seized in Guarulhos, and, between 2008 and
2009, VIGIAGRO seized 19 tons of these products in the
Galeão airport. Other studies show that this type of food
may contain pathogenic bacteria (Melo et al. 2014a, 2015);
however, no data in the literature describe the presence of
viruses in animal products in Brazil.
The HAV and RV are commonly involved in food-borne
outbreaks (Brasil 2018; Centers for Disease Control and
Prevention 2017). In Argentina and Uruguay, HAV is con-
sidered the causative agent of an endemic disease (Mon-
tano et al. 2001; Blánco Fernandez et al. 2012) and RV is
responsible for 40% of hospitalizations due to acute infant
diarrhea in Argentina and Uruguay (Degiuseppe et al. 2013;
Tort et al. 2015). In Brazil and Argentina, vaccination for RV
in children was introduced into the public health system in
the middle of 2000s, in addition, the two countries also vac-
cinate children and some adult groups for HAV. In Uruguay,
vaccination for HAV is universal (all ages), but for RV the
public health system does not yet provide the vaccine. In
13

these countries, regressive epidemiological analyzes have
shown a reduction in the incidence of these diseases after the
establishment of vaccination programs (Romero et al. 2012;
Baay et al. 2017; Santos et al. 2017).
Detecting the viral genome in the food evaluated is
indicative of its presence; however, this does not indicate
its infectious capacity. Our study detected high counts of
HAV by RT-qPCR in food obtained in Argentina and Uru-
guay, with samples containing 1­ 010 copies of the genome.
Although the infectious dose of the viruses evaluated in this
study is not yet established, studies indicate that the incuba-
tion period is shortened with a higher amount of inoculum.
Besides, it is estimated that 0.1 g of feces contaminated by
HAV may be considered as the infectious dose for humans
(White et al. 2016). The food analyzed in this study could
serve as sources of infection to exposed consumers. Natu-
rally, samples with a higher number of genome copies have
a higher probability of being infectious.
There was a significant association between HAV and
processed meats, compared to raw meats from Uruguay.
However, although the results were not significant, pro-
cessed meats presented a higher frequency of HAV and RV
than raw meats. Because these products pass through more
stages during processing, the chances of contamination by
enteropathogens are higher, as indicated by the significant
OR compared to HAV contamination in processed and raw
meats for all samples (OR 2.353; IC95% 1.012–5.473).
The introduction of these microorganisms in the pro-
duction chain occurs through contaminated raw material,
non-treated water, cross-contamination by feces, inadequate
manipulation of food or any ambient source containing viral
particles (Shukla et al. 2016). Therefore, it is possible that
the higher the number of stages in the production chain, the
higher the manipulation, and consequently, the probability
of contamination.
The presence of viable pathogens in ready-for-consump-
tion food may increase the risk of food-borne outbreaks, as
these products are not exposed to any processes to elimi-
nate microorganisms such as thermic treatment. The fact
that HAV and RV were detected does not indicate that these
foods will serve as a source of infection. However, the pres-
ence of genetic material is indicative of virus presence and
points to hygienic deficiencies that allow contamination by
HAV and RV during the acquisition or processing of the
animal products analyzed. Hygienic deficiencies in Argen-
tinian and Uruguayan establishments have been reported by
consumers that illegally import foods made in the border
area of RS, Brazil (Pereira et al. 2017). This was confirmed
in this study by the high number of establishments trading
animal products containing HAV and RV in Argentina (7/8)
and Uruguay (4/8).
The absence of HEV indicates that the virus is not circu-
lating in the studied area or it has a limited circulation and
13
Food and Environmental Virology
low undetectable levels. However, some studies show that
animal products are an important vehicle of this pathogen,
including the ones that circulate in border areas. Rodríguez-
Lázaro et al. (2015) detected HEV in 53.3% of the food
samples seized at the Bilbao airport, in Spain, including
meat products imported from South America. The highest
frequency of HEV was detected in products from Colom-
bia, Peru, Brazil, and Argentina, with pork being the main
contaminated product. Swine, as well as their meat and
byproducts, are the main vehicles of HEV (Wilhelm et al.
2014; Vasconcelos et al. 2015), which has been detected in
fecal and carcass samples obtained from slaughterhouses
and retail stores (Intharasongkroh et al. 2016), as well as in
samples of meat products. Heldt et al. (2016) detected the
HEV genome in 36% of samples of pork pâté commercial-
ized in Novo Hamburgo, Brazil.
The results of this study emphasize the need for enhanced
monitoring of food transport in the borders of Brazil, since
HAV and RV viruses were detected in fresh and processed
meats obtained from Argentina and Uruguay. Considering
that at the studied area, the practice of illegal transport is
widespread, and that the products entering Brazil (pro-
cessed) may contain important pathogens to public health,
it is necessary to implement more effective border inspec-
tion programs. In addition, border agricultural surveillance
should be more active and operational to stop the entry of
food containing potentially pathogenic agents that may cause
food-borne diseases, thereby protecting the health of Brazil-
ians living in this area. Otherwise, the possibility of illegal
transportation of animal products carrying pathogens such
as Porcine Reproductive & Respiratory Syndrome and Foot-
and-mouth disease cannot be estimated.
Acknowledgements  We thank the VIGIAGRO-MAPA-Uruguaiana
for supplying the seized samples, for supervision during sample col-
lection, and introduction of food obtained in Argentina and Uruguay.
We also thank Prof. Dr. Célia Barardi of the Federal University of
Santa Catarina for the yield of the plasmids. We also thank the Federal
University of Pampa (UNIPAMPA) for providing a scholarship to Ema-
noelli Aparecida Rodrigues dos Santos (Programa de Desenvolvimento
à Pesquisa PDA 2016) and for financial support (Programa de Apoio
aos Grupos de Pesquisa).
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