BACKGROUND
hepatitis b virus  is a dual polarity and partially double-stranded enveloped dna virus of hepadnaviridae family  <cit> .
the agent can be transmitted by sexual, perinatal and percutaneous means  <cit> , and is considered a major cause of acute and chronic liver disease, that may progress to cirrhosis and hepatocellular carcinoma  <cit> .
beyond the prototype member that infects humans and non-human primates , hepadnaviridae family also houses hbv-related viruses circulating in mammalian hosts, like woodchuck  and squirrels , and avian hosts, like ducks , geese , herons , and storks   <cit> .
a virus similar to hbv has been diagnosed by serology  in swine herds  <cit>  and chickens flocks  <cit>  from china, and in domestic pig herds in brazil  <cit> .
moreover, positive molecular diagnosis was demonstrated for the first time in swine from brazil  <cit>  and in chickens flocks from china  <cit> .
similarity with human hbv  was confirmed for swine strains by phylogenetic analysis and by cross reactivity in non-host specific commercial serological assays.
however, the partial nucleotide sequencing  was equivalent to about  <dig> % of full-length hbv genome.
and even for chickens, despite the high percentage of similarity with hbv , the short length of amplified product in both cases limits a conclusion.
therefore, we performed the current study to improve the molecular characterization of hepadnavirus circulating in swine from abattoirs in brazil, revealing if there is evidence that pigs destined for human consumption might act as a potential new reservoir or host for a virus hbv-similar.
methods
this study was approved by the institutional committee for ethics in the use of research animals .
on december  <dig>  a total of  <dig> bile and liver samples were collected from domestic pigs sus scrofa , breed large white, from three slaughterhouses located in petrópolis , itaocara  and itaperuna  .
commercial establishments are submitted to controlled inspection by an official agency of rio de janeiro state.
all animals from the three abattoirs were classified healthy and approved for slaughter and further commercialization according to inspect evaluation criteria.
swine from each abattoir were acquired from distinct pig farms suppliers.
the samples were collected during the evisceration process under sanitary requisites determined by regulations of animal sanitary protection agency in rio de janeiro state .
five milliliters of bile were collected by vaccum-punction through the gallbladder wall with sterile syringe; and 500 mg of liver samples were collected with medblade® bisturi blades.
all samples were stored in nalgene® cryogenic vials and immediately frozen in dry ice.
at the laboratory, bile samples were stored at -80°c and liver samples in liquid nitrogen until analysis.
viral dna was extracted from bile and liver samples using dna purification kit  according to moricz et al,  <dig>  <cit>  and was concentrated to a final volume of 25 ul.
extracted dna was analyzed by semi-nested pcr  specific for open reading frame  of hbv - as previously established for the first round of amplification  <cit> , and for the second round  <cit>  - and by pcr specific for core gene of hbv – as previously established  <cit> .
direct sequencing of amplicons was performed to identify the sequence amplified.
after molecular diagnosis, in order to quantify the viral load dna samples were evaluated in duplicate by real time pcr using taqman® method.
the real time pcr assay was performed for pre-s2/s region as previously established  <cit> , using the following primers pair and fluorescent probe: forward primer , reverse primer , and probe  .
hbv plasmid serially diluted from 5 ×  <dig> to 5 ×  <dig> copies/ μl was used as hbv standard pcr template.
the amplification was analyzed using the software applied biosystems  <dig>
indirect immunofluorescence assay was also performed in samples of liver tissue cryosectioned at 4 μm.
liver biopsies were stained with anti-hbs mouse monoclonal primary antibody specific for human hbsag  at the dilution 1: <dig> overnight at 4°c, followed by alexa fluor  <dig> conjugated goat anti-mouse secondary antibodies  at the dilution 1: <dig> for 1 h at 37°c.
liver sections were counterstained with evans blue  and dapi .
images of the green fluorescent hbsag-positive liver cells were observed and photographed using a confocal scanning laser microscope equipped with camera .
RESULTS
hepadnavirus-dna was detected in bile and liver samples from about  <dig> % pigs .
all samples from petrópolis  were tested negative.
hepadnavirus-dna was detected in both samples types from  <dig> animal in itaocara  and from  <dig> animals in itaperuna .
these samples showed a positive result for the semi-nested pcr specific for hbv orf s  , and  <dig> of them also for the pcr specific for core gene  .
the sequences found in bile and liver of the same animal matched.
phylogenetic reconstruction using partial nucleotide sequences of orf s  showed a close relationship of hepadnavirus strains from pigs and hepadnavirus nucleotide sequences from non-human primates , bats , rodents  and birds  .figure  <dig>
phylogenetic analysis of swine hepadnavirus strains and
hepadnaviridae
family.
the bayesian phylogenetic tree was constructed by using partial nucleotide sequence of open reading frame s  of hbv and related viruses.
for each sequence used, the genbank accession number and country of isolation are shown.
multiple nucleotide sequence alignment was analyzed by using the markov chain monte carlo method implemented in the program mrbayes version  <dig> .
<dig> under gtr + g nucleotide substitution model, selected using the jmodeltest program.
the tree was rooted at midpoint.
posterior probabilities are shown at the branch label.
newly described swine hepadnavirus sequences are indicated.
scale bar indicates evolutionary distance.
nucleotide identity between hepadnavirus strains from pigs and other species
species
accession number genbank
swine hepadnavirus
kc832935
kc832936
kc832937
kc832938
legend: the nucleotide identity matrix was constructed by using partial nucleotide sequence of open reading frame s  of hbv and related viruses.
for each sequence used, the genbank accession number and specie infected are shown.
newly described swine hepadnaviruses sequences are indicated.
genetic distances calculated with nucleotide sequences of orf s  showed filogenetic distance between the virus detected in brazilian domestic pigs and human samples from significant alignment listed by ncbi blast  ranging from  <dig> - <dig> %.
these data also revealed that distinct strains might be responsible for the infection in domestic pigs .
comparing to human hbv strains previously genotyped, swine samples were assigned to two different genotypes groups.
three of them were close to samples from genotype a, the most prevalent genotype in brazil  <cit> .
and, another was similar to samples that belong to genotype e, which is common in western africa  <cit> .
genotype a was found circulating in itaocara and itaperuna, while genotype e was found exclusively in itaperuna.figure  <dig>
phylogenetic analysis of swine hepadnavirus strains and hbv from human samples.
the bayesian phylogenetic tree was constructed by using partial nucleotide sequence of open reading frame s  of hbv.
for each sequence used, the genbank accession number and country of isolation are shown.
multiple nucleotide sequence alignment was analyzed by using the markov chain monte carlo method implemented in the program mrbayes version  <dig> .
<dig> under gtr + g nucleotide substitution model, selected using the jmodeltest program.
the tree was rooted at midpoint.
posterior probabilities are shown at the branch label.
newly described swine hepadnavirus sequences are indicated.
scale bar indicates evolutionary distance.
concerning viral load quantification, a linear relationship was obtained between the cycle threshold  values and the log <dig> concentration of the hbv dna.
the regression analysis yielded a correlation coefficient of  <dig> .
all four bile samples could be quantified.
the viral loads ranged from  <dig>  ×  <dig> to 1 ×  <dig> copies/ml.
further, we investigated viral hepatotropism by indirect immunofluorescence analysis using monoclonal anti-hbs antibody in liver biopsies from hepdnavirus-dna positive animals, and compared the results to the control animals.
hbsag was observed in cell membrane and cytoplasm of sinusoidal lining cells in liver parenchyma and in hepatocytes .
no staining was observed for controls.
molecular data and antigenic detection in situ confirm that swine were infected by a hepadnavirus with hepatotropic behavior, similar to human hbv.figure  <dig>
detection of hepatitis b virus surface antigen in liver of swine by immunofluorescence staining.
different frozen sections of hbsag-positive cells  from liver of pigs stained in green using alexa fluor 488-conjugated anti-mouse igg as secondary antibody.
evans blue  and dapi  counterstained all liver sections.
images were examined under a confocal immunofluorescent microscope .
liver sections from negative hepadnavirus-dna animals were used as negative controls .
discussion
our challenge in this study was to refine the molecular characterization of hepadnavirus circulating in swine from brazil, disclosing how close porcine strains are to hbv and to hbv-like viruses sequences described for different hosts, including human.
data presented here showed different strategies for viral detection, including the molecular characterization of complete orf c and orf s in bile and liver samples, the genotype classification based on the genotypic classification of hbv, the viral load quantification and antigenic detection in hepatic parenchyma.
this is the first record of molecular characterization of swine hepadnavirus from bile and liver samples.
there are no previous data available in the literature concerning genotypic classification based on complete sequencing of pre-s/s region and viral load evaluation in swine from slaughterhouses.
according to results described previously for brazilian domestic swine  <cit> , the similarity between pigs and human hepadnavirus strains was supposed through superficial and preliminary estimates, and cross reactivity in non-host specific commercial serological assays.
in this study, molecular results from a larger and more representative fragment of the genome allowed us to calculate an approximate identity not only with hbv from human host, as well as other viruses that comprise the hepdnaviridae family:  <dig> - <dig> % to human hbv,  <dig> - <dig> % to chimpanzee hbv,  <dig> - <dig> % to gibbon hbv,  <dig> - <dig> % to gorilla hbv,  <dig> - <dig> % to orangutan hbv,  <dig> - <dig> % to woolly monkey wmhbv,  <dig> %- <dig> % to bat hepadnavirus,  <dig> - <dig> % to woodchuck whv,  <dig> - <dig> % to arctic/ ground squirrels ashv/ gshv,  <dig> - <dig> % to heron hhbv,  <dig> - <dig> % to stork sthbv,  <dig> - <dig> % to snow goose ghbv,  <dig> - <dig> % to duck dhbv.
genetic distances data are relevant and need to be better evaluated once despite the diversity of hosts, no reservoirs are described for hbv so far  <cit> .
at the same time, as commercial swine are slaughtered at a young age , the identification of any clinical signs of disease may be hindered notably considering that health inspections are based only in visible lesions present and post-mortem gross macroscopic examinations in animal from abattoirs.
no molecular tests are executed.
due to these factors, a follow-up study for longer periods should be considered.
it is noteworthy that investigation of the population of animals from three different commercial herds was performed after a single collection, and  <dig> % of swine showed positive molecular diagnosis, in which hepatotropism was also confirmed.
these positive animals were distributed in two slaughterhouses which do not share the same pig farm supplier.
the possibility of an eventual vertical transmission can not be ruled out for positive animals from the same slaughterhouse since they came from the same herd.
however, as there are distinct breeding sows in a single pig farm, animals destined to the same abattoir may be born from different sows.
to consider the risk of vertical transmission, pigs should also share the same genotype in addition to sharing the same geographical origin.
in this study, three positive animals came from itaperuna.
one animal was assigned to genotype e, and two animals shared genotype a. due to the circulation of both genotypes in swine from the same abattoir, at least two different breeding sows were involved in its respective pig farm supplier.
even though, the risk of vertical transmission for two animals of genotype a cannot be confirmed.
since the role of bats as reservoirs of zoonotic viruses is now being investigated, including for bat hepadnaviruses  <cit> , further studies are also necessary to extrapolate this reasoning to the pigs analogously.
whether there is a zoonotic ability of swine hepadnavirus strains and if this virus is able to induce cross-infection in other species are remaining issues.
considering the main routes of hbv transmission, and since personal protective equipment is not often used in slaughterhouses, novel studies are necessary to assess a potential occupational risk of infection by hepadnaviruses during the management of pigs and their derivatives.
CONCLUSIONS
the present study demonstrated the circulation of an emerging virus in swine, similar to hepadnavirus that causes human infection.
further studies are necessary to disclose the actual role of swine in the viral cycle.
abbreviations
hbvhuman hepatitis b virus
orf sopen reading frame s
orf copen reading frame c
whvhbv-related virus of woodchuck
gshvhbv-related virus of ground squirrels
ashvhbv-related virus of arctic squirrels
dhbvhbv-related virus of ducks
ghbvhbv-related virus of geese
hhbvhbv-related virus of herons
sthbvhbv-related virus of storks
hbsagsurface antigen of hbv
anti-hbshbv surface antibody
anti-hbcantibody to hbv core antigen
bpbase pair
ceuacommittee for ethics in the use of research animals
aspaanimal sanitary protection agency in rio de janeiro state
ntnucleotide
competing interests
the authors declare that they have no competing interests.
authors’ contributions
this study was designed by yrv, drls, map and vsp.
yrv performed qualitative molecular assays, sequence analysis, interpreted the data, collaborated to quantitative molecular assays, to immunofluorescence assay and wrote the manuscript.
drls contributed to the sample collection and helped to write the manuscript.
mmp performed quantitative molecular assays.
cepv contributed to the immunofluorescence assay.
ma contributed with immunological reagents.
lmv contributed with molecular reagents, materials and analysis tools.
map contributed to the sample collection, coordinated the study and revised the manuscript.
vsp coordinated the study and revised the manuscript.
all authors read, corrected and approved the final manuscript.
this study was supported by oswaldo cruz institute , coordenação de aperfeiçoamento de pessoal de nível superior  and fundação de amparo à pesquisa do estado do rio de janeiro .
the authors appreciated the help from gilberto vaughan  and leandro layter xavier .

 hepatitis b virus is a dual polarity and partially double-stranded enveloped dna virus of hepadnaviridae family. the agent can be transmitted by sexual, perinatal and percutaneous means, and is considered a major cause of acute and chronic liver disease, that may progress to cirrhosis and hepatocellular carcinoma. beyond the prototype member that infects humans and non-human primates, hepadnaviridae family also houses hbv-related viruses circulating in mammalian hosts, like woodchuck and squirrels, and avian hosts, like ducks, geese, herons, and storks. a virus similar to hbv has been diagnosed by serology in swine herds and chickens flocks from china, and in domestic pig herds in brazil. moreover, positive molecular diagnosis was demonstrated for the first time in swine from brazil and in chickens flocks from china. similarity with human hbv was confirmed for swine strains by phylogenetic analysis and by cross reactivity in non-host specific commercial serological assays. however, the partial nucleotide sequencing was equivalent to about % of full-length hbv genome. and even for chickens, despite the high percentage of similarity with hbv, the short length of amplified product in both cases limits a conclusion. therefore, we performed the current study to improve the molecular characterization of hepadnavirus circulating in swine from abattoirs in brazil, revealing if there is evidence that pigs destined for human consumption might act as a potential new reservoir or host for a virus hbv-similar. methods this study was approved by the institutional committee for ethics in the use of research animals. on december a total of bile and liver samples were collected from domestic pigs sus scrofa, breed large white, from three slaughterhouses located in petrópolis, itaocara and itaperuna. commercial establishments are submitted to controlled inspection by an official agency of rio de janeiro state. all animals from the three abattoirs were classified healthy and approved for slaughter and further commercialization according to inspect evaluation criteria. swine from each abattoir were acquired from distinct pig farms suppliers. the samples were collected during the evisceration process under sanitary requisites determined by regulations of animal sanitary protection agency in rio de janeiro state. five milliliters of bile were collected by vaccum-punction through the gallbladder wall with sterile syringe; and 500 mg of liver samples were collected with medblade® bisturi blades. all samples were stored in nalgene® cryogenic vials and immediately frozen in dry ice. at the laboratory, bile samples were stored at -80°c and liver samples in liquid nitrogen until analysis. viral dna was extracted from bile and liver samples using dna purification kit according to moricz et al, and was concentrated to a final volume of 25 ul. extracted dna was analyzed by semi-nested pcr specific for open reading frame of hbv - as previously established for the first round of amplification, and for the second round - and by pcr specific for core gene of hbv – as previously established. direct sequencing of amplicons was performed to identify the sequence amplified. after molecular diagnosis, in order to quantify the viral load dna samples were evaluated in duplicate by real time pcr using taqman® method. the real time pcr assay was performed for pre-s2/s region as previously established, using the following primers pair and fluorescent probe: forward primer, reverse primer, and probe. hbv plasmid serially diluted from 5 ×  to 5 ×  copies/ μl was used as hbv standard pcr template. the amplification was analyzed using the software applied biosystems indirect immunofluorescence assay was also performed in samples of liver tissue cryosectioned at 4 μm. liver biopsies were stained with anti-hbs mouse monoclonal primary antibody specific for human hbsag at the dilution 1: overnight at 4°c, followed by alexa fluor conjugated goat anti-mouse secondary antibodies at the dilution 1: for 1 h at 37°c. liver sections were counterstained with evans blue and dapi. images of the green fluorescent hbsag-positive liver cells were observed and photographed using a confocal scanning laser microscope equipped with camera.  hepadnavirus-dna was detected in bile and liver samples from about % pigs. all samples from petrópolis were tested negative. hepadnavirus-dna was detected in both samples types from animal in itaocara and from animals in itaperuna. these samples showed a positive result for the semi-nested pcr specific for hbv orf s, and of them also for the pcr specific for core gene. the sequences found in bile and liver of the same animal matched. phylogenetic reconstruction using partial nucleotide sequences of orf s showed a close relationship of hepadnavirus strains from pigs and hepadnavirus nucleotide sequences from non-human primates, bats, rodents and birds.figure phylogenetic analysis of swine hepadnavirus strains and hepadnaviridae family. the bayesian phylogenetic tree was constructed by using partial nucleotide sequence of open reading frame s of hbv and related viruses. for each sequence used, the genbank accession number and country of isolation are shown. multiple nucleotide sequence alignment was analyzed by using the markov chain monte carlo method implemented in the program mrbayes version. under gtr + g nucleotide substitution model, selected using the jmodeltest program. the tree was rooted at midpoint. posterior probabilities are shown at the branch label. newly described swine hepadnavirus sequences are indicated. scale bar indicates evolutionary distance. nucleotide identity between hepadnavirus strains from pigs and other species species accession number genbank swine hepadnavirus kc832935 kc832936 kc832937 kc832938 legend: the nucleotide identity matrix was constructed by using partial nucleotide sequence of open reading frame s of hbv and related viruses. for each sequence used, the genbank accession number and specie infected are shown. newly described swine hepadnaviruses sequences are indicated. genetic distances calculated with nucleotide sequences of orf s showed filogenetic distance between the virus detected in brazilian domestic pigs and human samples from significant alignment listed by ncbi blast ranging from - %. these data also revealed that distinct strains might be responsible for the infection in domestic pigs. comparing to human hbv strains previously genotyped, swine samples were assigned to two different genotypes groups. three of them were close to samples from genotype a, the most prevalent genotype in brazil. and, another was similar to samples that belong to genotype e, which is common in western africa. genotype a was found circulating in itaocara and itaperuna, while genotype e was found exclusively in itaperuna.figure phylogenetic analysis of swine hepadnavirus strains and hbv from human samples. the bayesian phylogenetic tree was constructed by using partial nucleotide sequence of open reading frame s of hbv. for each sequence used, the genbank accession number and country of isolation are shown. multiple nucleotide sequence alignment was analyzed by using the markov chain monte carlo method implemented in the program mrbayes version. under gtr + g nucleotide substitution model, selected using the jmodeltest program. the tree was rooted at midpoint. posterior probabilities are shown at the branch label. newly described swine hepadnavirus sequences are indicated. scale bar indicates evolutionary distance. concerning viral load quantification, a linear relationship was obtained between the cycle threshold values and the log concentration of the hbv dna. the regression analysis yielded a correlation coefficient of. all four bile samples could be quantified. the viral loads ranged from  ×  to 1 ×  copies/ml. further, we investigated viral hepatotropism by indirect immunofluorescence analysis using monoclonal anti-hbs antibody in liver biopsies from hepdnavirus-dna positive animals, and compared the results to the control animals. hbsag was observed in cell membrane and cytoplasm of sinusoidal lining cells in liver parenchyma and in hepatocytes. no staining was observed for controls. molecular data and antigenic detection in situ confirm that swine were infected by a hepadnavirus with hepatotropic behavior, similar to human hbv.figure detection of hepatitis b virus surface antigen in liver of swine by immunofluorescence staining. different frozen sections of hbsag-positive cells from liver of pigs stained in green using alexa fluor 488-conjugated anti-mouse igg as secondary antibody. evans blue and dapi counterstained all liver sections. images were examined under a confocal immunofluorescent microscope. liver sections from negative hepadnavirus-dna animals were used as negative controls. discussion our challenge in this study was to refine the molecular characterization of hepadnavirus circulating in swine from brazil, disclosing how close porcine strains are to hbv and to hbv-like viruses sequences described for different hosts, including human. data presented here showed different strategies for viral detection, including the molecular characterization of complete orf c and orf s in bile and liver samples, the genotype classification based on the genotypic classification of hbv, the viral load quantification and antigenic detection in hepatic parenchyma. this is the first record of molecular characterization of swine hepadnavirus from bile and liver samples. there are no previous data available in the literature concerning genotypic classification based on complete sequencing of pre-s/s region and viral load evaluation in swine from slaughterhouses. according to results described previously for brazilian domestic swine, the similarity between pigs and human hepadnavirus strains was supposed through superficial and preliminary estimates, and cross reactivity in non-host specific commercial serological assays. in this study, molecular results from a larger and more representative fragment of the genome allowed us to calculate an approximate identity not only with hbv from human host, as well as other viruses that comprise the hepdnaviridae family: - % to human hbv, - % to chimpanzee hbv, - % to gibbon hbv, - % to gorilla hbv, - % to orangutan hbv, - % to woolly monkey wmhbv, %- % to bat hepadnavirus, - % to woodchuck whv, - % to arctic/ ground squirrels ashv/ gshv, - % to heron hhbv, - % to stork sthbv, - % to snow goose ghbv, - % to duck dhbv. genetic distances data are relevant and need to be better evaluated once despite the diversity of hosts, no reservoirs are described for hbv so far. at the same time, as commercial swine are slaughtered at a young age, the identification of any clinical signs of disease may be hindered notably considering that health inspections are based only in visible lesions present and post-mortem gross macroscopic examinations in animal from abattoirs. no molecular tests are executed. due to these factors, a follow-up study for longer periods should be considered. it is noteworthy that investigation of the population of animals from three different commercial herds was performed after a single collection, and % of swine showed positive molecular diagnosis, in which hepatotropism was also confirmed. these positive animals were distributed in two slaughterhouses which do not share the same pig farm supplier. the possibility of an eventual vertical transmission can not be ruled out for positive animals from the same slaughterhouse since they came from the same herd. however, as there are distinct breeding sows in a single pig farm, animals destined to the same abattoir may be born from different sows. to consider the risk of vertical transmission, pigs should also share the same genotype in addition to sharing the same geographical origin. in this study, three positive animals came from itaperuna. one animal was assigned to genotype e, and two animals shared genotype a. due to the circulation of both genotypes in swine from the same abattoir, at least two different breeding sows were involved in its respective pig farm supplier. even though, the risk of vertical transmission for two animals of genotype a cannot be confirmed. since the role of bats as reservoirs of zoonotic viruses is now being investigated, including for bat hepadnaviruses, further studies are also necessary to extrapolate this reasoning to the pigs analogously. whether there is a zoonotic ability of swine hepadnavirus strains and if this virus is able to induce cross-infection in other species are remaining issues. considering the main routes of hbv transmission, and since personal protective equipment is not often used in slaughterhouses, novel studies are necessary to assess a potential occupational risk of infection by hepadnaviruses during the management of pigs and their derivatives.  the present study demonstrated the circulation of an emerging virus in swine, similar to hepadnavirus that causes human infection. further studies are necessary to disclose the actual role of swine in the viral cycle. abbreviations hbvhuman hepatitis b virus orf sopen reading frame s orf copen reading frame c whvhbv-related virus of woodchuck gshvhbv-related virus of ground squirrels ashvhbv-related virus of arctic squirrels dhbvhbv-related virus of ducks ghbvhbv-related virus of geese hhbvhbv-related virus of herons sthbvhbv-related virus of storks hbsagsurface antigen of hbv anti-hbshbv surface antibody anti-hbcantibody to hbv core antigen bpbase pair ceuacommittee for ethics in the use of research animals aspaanimal sanitary protection agency in rio de janeiro state ntnucleotide competing interests the authors declare that they have no competing interests. authors’ contributions this study was designed by yrv, drls, map and vsp. yrv performed qualitative molecular assays, sequence analysis, interpreted the data, collaborated to quantitative molecular assays, to immunofluorescence assay and wrote the manuscript. drls contributed to the sample collection and helped to write the manuscript. mmp performed quantitative molecular assays. cepv contributed to the immunofluorescence assay. ma contributed with immunological reagents. lmv contributed with molecular reagents, materials and analysis tools. map contributed to the sample collection, coordinated the study and revised the manuscript. vsp coordinated the study and revised the manuscript. all authors read, corrected and approved the final manuscript. this study was supported by oswaldo cruz institute, coordenação de aperfeiçoamento de pessoal de nível superior and fundação de amparo à pesquisa do estado do rio de janeiro. the authors appreciated the help from gilberto vaughan and leandro layter xavier.
in this study, we genetically characterize the swine hepadnavirus strains in swine from slaughterhouses located in certified abattoirs from rio de janeiro state, brazil and evaluate its hepatotropic potential. bile and liver samples from swine were positive for partial genome amplification, direct sequencing and viral load quantification.
 preliminary studies showed the prevalence of a virus similar to human hepatitis b virus in swine from farms in china and the molecular evidence of hepadnavirus infection in domestic pigs herds in brazil. in this study, we genetically characterize the swine hepadnavirus strains in swine from slaughterhouses located in certified abattoirs from rio de janeiro state, brazil and evaluate its hepatotropic potential.  bile and liver samples from swine were positive for partial genome amplification, direct sequencing and viral load quantification. sequencing of the gene encoding the surface antigen allowed classification of hepadnavirus into genotypes, similar to hbv genotype classification. indirect immunofluorescence confirmed the presence of hbsag antigen in liver tissue sections.  so far our data suggest that commercial swine house an hbv-like virus and this relevant finding should be considered in studies on the origin and viral evolution. keywords hepadnavirusbrazilcommercial swinebilelivergenotypingimmunofluorescenceissue-copyright-statement© the author 2014 