KPC-2-producing Pseudomonas aeruginosa isolated from wild animals in Brazil.

Pseudomonas aeruginosa an opportunistic pathogen that causes infections in hospitals and has high morbidity and mortality rates. In addition, it is a widely distributed environmental bacterium that can colonise a variety of habitats. Although wild animals do not have access to antibiotics, antibacterial resistance in these animals has increasingly been reported worldwide. Although the presence of Klebsiella pneumoniae carbapenemase (KPC) is uncommon in P. aeruginosa, it has been increasingly reported. This study examined KPC-2-producing P. aeruginosa in wild animals. A total of 27 P. aeruginosa isolates were obtained from clinical cases treated at the Microbiology Laboratory of the Veterinary Hospital of UFMT, Brazil. P. aeruginosa and blaKPC-2 carbapenemase resistance genes were identified using PCR. Antimicrobial susceptibility of KPC-producing P. aeruginosa was evaluated using the disk diffusion method. The blaKPC-2 gene was detected in 40.7% of the isolates (11/27). The rates of antimicrobial resistance and intermediate sensitivity were as follows: piperacillin/tazobactam (44.4%), imipenem (29.6%), meropenem (51.8%), amikacin (77.8%), cefepime (85.2%), and ciprofloxacin (70.4%). Twelve isolates were classified as Multidrug-resistant (MDR). This study presents the first report of P. aeruginosa with the blaKPC-2 gene in wild animals in Brazil, highlighting the importance of molecular research on resistance genes in P. aeruginosa from a One-Health perspective.

Molecular prevalence and factors associated with Ehrlichia canis infection in dogs from the North Pantanal wetland, Brazil.

Background and Aim: Canine monocytic ehrlichiosis is a vector-borne disease caused by the obligatory intracellular bacterium Ehrlichia canis, which is distributed across tropical and subtropical regions worldwide. Its prevalence within dog populations is high in municipalities located across the Pantanal biome, but it remains unknown in Barão de Melgaço, Mato Grosso, Brazil. This study aimed to determine the molecular prevalence and factors associated with E. canis infection in dogs domiciled in Barão de Melgaço. Materials and Methods: A cross-sectional study was carried out to investigate the prevalence of E. canis infection in 369 dogs from urban and rural areas in Barão de Melgaço, North Pantanal wetland, Brazil. Initially, the dogs were examined, and, through a questionnaire, the risk factors were investigated. Blood samples were subjected to DNA extraction and PCR was performed to estimate the prevalence of E. canis infection. Results: The molecular prevalence of E. canis infection in dogs was 42.5% and none of the studied variables were significantly associated with polymerase chain reaction (PCR) positivity (p > 0.05). Conclusion: The high molecular prevalence demonstrates an increased transmission of the agent across the city. This also indicates that attention needs to be paid to E. canis infection and control measures should be introduced to prevent its transmission. The demographic and clinical risk factors commonly associated with E. canis infection in this study were not associated with PCR positivity.

Pleural effusion-related Nocardia otitidiscaviarum, Anaplasma platys and Ehrlichia canis coinfection in a dog.

The coinfections by some microorganisms have been related to severe diseases in humans and animals, where immunosuppressive agents favor opportunistic behavior of other pathogens. A 4-month-old, female mixed-breed dog with a two-week history of inappetence, prostration, emaciation, and respiratory distress was admitted at a veterinary hospital in Brazil. Tachycardia, pale mucous membranes, severe respiratory distress, and a large number of ticks (Rhipicephalus sanguineus s.l.) in different body regions were observed at clinical examination. Hematological examination of dog showed leukocytosis, neutrophilia, mild anemia, and thrombocytopenia, whereas unremarkable values in biochemical tests. Thoracic radiography revealed a pleural effusion image. Blood and the pleural fluid (purulent aspect) samples were subjected to qPCR (16S rRNA and dsb genes) and sequencing, which identified Ehrlichia canis and Anaplasma platys coinfection. An aggregate of coccoid-to-branching or long filamentous microorganisms, surrounded by pyogranulomatous inflammatory reaction was seen at the cytology of the pleural fluid. Bacteriological culture of pleural effusion showed colonies compatible with the genus Nocardia, which revealed gram-positive filamentous organisms with a tendency of fragmentation and were identified as Nocardia otitidiscaviarum in mass spectrometry (MALDI-TOF MS). Therapy of N. otitidiscaviarum isolate using levofloxacin (supported by a previous in vitro susceptibility testing) and doxycycline for E. canis and A. platys resulted in complete resolution of the clinical picture. Here, we report for the first time a triple coinfection by Nocardia otitidiscaviarum, A. platys, and E. canis in a dog with pleural effusion, where debilitating or immunosuppressive conditions induced by A. platys and E. canis coinfection probably contributed to the opportunistic behavior of N. otitidiscaviarum.

Brazilian porcupinepox virus infection in a free-ranging neotropical porcupine in Mato Grosso, Brazil.

Although the novel Brazilian porcupinepox virus (BPoPV) can infect wild porcupines, its lethality and zoonotic potential are not well-established. In May 2021, a free-ranging neotropical porcupine (Coendou longicaudatus boliviensis) rescued from the natural savanna in the Brazilian Cerrado, Mato Grosso State, was presented with a lethal poxviral infection. Clinical signs and PCR detection of BPoPV supported the diagnosis. Poxviral lesions included erythema, exfoliative dermatitis, and erosions involving mainly the face, hindlimb, and vulva. Histologically, the lesions consisted of proliferative and necrotic dermatitis, intraepidermal and follicular pustules, and intracytoplasmic inclusion bodies in keratinocytes. Phylogenetic analysis revealed BPoPV strains closely related to other rodent-infecting poxviruses. This poxviral infection resulted in the death of a specimen of C. l. boliviensis; the effect on species conservation, and the potential of spillover into humans and other vertebrates remain unknown.

Molecular detection and genetic characterization of Ehrlichia canis and Ehrlichia sp. in neotropical primates from Brazil.

The Anaplasmataceae family includes obligate, arthropod-transmitted intracellular bacteria that can be zoonotic and potentially fatal. Studies focusing on the interaction between neotropical primates and the agents of this family are scarce. The present study aimed to identify agents of the Anaplasmataceae family in the whole blood of free-living and captive neotropical primates in the State of Mato Grosso, Central-West Brazil. Thirty-eight samples of six nonhuman primate (NHP) species were collected in seven municipalities and analysed through polymerase chain reaction (PCR), nucleotide sequencing, and phylogenetic analysis of the dsb, groEL, 16S rRNA, and gltA genes. DNA fragments similar to those of Ehrlichia canis were detected in Sapajus apella and Ehrlichia chaffeensis from Mico melanurus. The sequences generated in this study and homologous sequences retrieved from GenBank® were used for phylogenetic analyses to characterize the Ehrlichial agents detected in NHPs. The agents were then grouped into clades corresponding to different isolates from the NHP species. In addition, an Anaplasma sp. closely related to Anaplasma marginale was identified in two S. apella individuals. These findings shed light on the susceptibility of neotropical NHPs to Anaplasmataceae agents. These bacteria are known to be transmitted by ticks, which can also serve as possible sources of infection for other animals, including humans.

SARS-CoV-2 antibodies in dogs and cats in a highly infected area of Brazil during the pandemic.

SARS-CoV-2 was a worldwide threat during the COVID-19 pandemic, and the state of Mato Grosso had the second highest mortality rate in Brazil, with 427. 4 deaths/100,000 inhabitants. However, no large-scale study among dogs and cats in such highly infected areas of Brazil has so far been conducted. Accordingly, the present study reports on a serosurvey among dogs and cats in Cuiabá, capital of Mato Grosso from November 2020 to July 2021, where the human mortality rate was 605/100,000 at that time. Overall, 33/762 dogs (4.3%) and 4/182 cats (2.2%) were found to be seropositive for SARS-CoV-2 through ELISA, and 3/762 dogs (0.4%) and 3/182 cats (1.6%) were seropositive through the serum neutralization test. Cats presented higher seroprevalence with higher titers of neutralizing antibodies. Although N-protein based ELISA may be a good screening test, cross-reactivity with other canine coronaviruses may impair its diagnostic use among dogs.

Pulmonary Granuloma Is Not Always the Tuberculosis Hallmark: Pathology of Tuberculosis Stages in New World and Old World Monkeys Naturally Infected with the Mycobacterium tuberculosis Complex.

We present the pathology of monkeys naturally infected with Mycobacterium tuberculosis complex from five different colonies in Rio de Janeiro, Brazil. On the basis of gross and histopathological findings, the lesions were classified into chronic-active, extrapulmonary, early-activation or latent-reactivation stages. Typical granulomatous pneumonia was seen in 46.6% of cases (six rhesus monkeys [Macaca mulatta] and one Uta Hick's bearded saki [Chiropotes utahickae]). The absence of pulmonary granulomas did not preclude a diagnosis of tuberculosis (TB): classical granulomatous pneumonia was observed in the chronic-active and latent-reactivation stages but not in the extrapulmonary and early-activation stages. The early-activation stage was characterized by interstitial pneumonia with a predominance of foamy macrophages and molecular and immunohistochemical evidence of M. tuberculosis complex infection. TB should be considered as a cause of interstitial pneumonia in New World Monkeys. We recommend the use of immunohistochemistry and molecular analysis for diagnosis of TB, even when typical macroscopic or histological changes are not observed.

Determination of multidrug-resistant populations and molecular characterization of complex Klebsiella spp. in wild animals by multilocus sequence typing.

Background and Aim: One of the most significant public health concerns is multidrug-resistant (MDR) microorganisms. Klebsiella spp. have been at the forefront of causing different types of infections such as bacteremia, urinary tract infections, pneumonia, enteritis, and sepsis in humans as well as animals. This study aimed to determine the genomic similarity between Klebsiella spp. isolated from wild animal samples and those described in the Institut Pasteur genomic database to verify the spread of resistant clones regionally in the state of Mato Grosso, and to compare the epidemiological data in different regions of Brazil and the world. Materials and Methods: Isolates from various sites of injury in wild animals were identified by sequencing the 16S rRNA gene. Antimicrobial susceptibility testing was performed using the disk diffusion method to verify the resistance profile, and then, multilocus sequence typing was performed to verify the population structure and compare the isolates from other regions of Brazil and the world. Results: Twenty-three sequence types (STs) were observed; of these, 11 were new STs, as new alleles were detected. There was no predominant ST among the isolates. All isolates were MDR, with high rates of resistance to sulfonamides, ampicillin, amoxicillin, and nitrofurantoin and low resistance to meropenem, imipenem, and amikacin. Conclusion: Improving our understanding of the population structure of Klebsiella spp. in wild animals may help determine the source of infection during outbreaks in humans or animals, as the One Health concept emphasizes the interlinks between humans, animals, and environmental health.

Clinical and parasitological impact of short-term treatment using miltefosine and allopurinol monotherapy or combination therapy in canine visceral leishmaniasis.

Canine visceral leishmaniasis is an endemic zoonosis in Brazil. Dogs are the main hosts in urban environments. The treatment has gained popularity since the Brazilian government authorized miltefosine for canine treatment. The aim of this study was to investigate the clinical and parasitological impact of short-term treatment with miltefosine and allopurinol, alone and in combination. We evaluated the ability of pharmacotherapy to reduce clinical signs of disease, antibody levels using the indirect fluorescence antibody test (IFAT) and skin parasite load via qPCR after 28 days of treatment. The therapeutic protocols promoted a significant decline in clinical signs and in the skin parasite load in dogs (p < 0.01). We observed a moderate correlation between the skin parasite load and the clinical score in all three treatment groups (r > 0.5) Antibody levels did not decrease in this short period. It was concluded that the treatment with allopurinol reduced the number of parasites in the skin of dogs with visceral leishmaniasis in the short term. However, its efficiency is potentiated when associated with miltefosine.

Genotyping of rabies positive samples isolated from animals in Mato Grosso and Rondônia - Brazil.

Lyssavirus is a genus that causes infectious disease transmit by bats transmit, which results in economic losses in livestock and public health problems. From 2005 to 2019, more than 49 thousand cases of the disease were registered in animals in Brazil, with 3418 registered in Mato Grosso (MT). The lack of information on the genetic diversity and distribution of the rabies virus in MT was the motivation for carrying out this study. A total of 117 samples of brain tissue from cattle, horses, donkeys, mules and sheep from 29 municipalities in the state of MT and one municipality in Rondônia were used. Direct immunofluorescence and/or biological tests performed from 2014 to 2021 indicated that all samples were positive for the disease. RNA was extracted and molecular analysis was performed using RT-PCR for the N gene. Of the 117 samples analyzed, 50 were amplified by RT-PCR, purified and sequenced. The samples showed 93.13%-100% identity with the rabies virus. The sequences were submitted to phylogenetic analysis that resulted in a tree of four clades; these were genetically grouped into distinct regions within the Desmodus rotundus lineage. The results of the geolocation of clades will be useful to guide monitoring, control and health surveillance programs in MT.

Natural SARS-CoV-2 Infection in a Free-Ranging Black-Tailed Marmoset (Mico melanurus) from an Urban Area in Mid-West Brazil.

The emergence of spillover pathogens such as SARS-CoV-2 is a risk to vulnerable human populations. We report natural SARS-CoV-2 infection in a free-ranging adult female black-tailed marmoset (Mico melanurus) from an urban area of Cuiabá, Mato Grosso State, Brazil. The animal was found after a motor vehicle collision without previous clinical history. Necropsy confirmed polytrauma. Severe multifocal to coalescent haemorrhage and mild multifocal peribronchial lymphocytic hyperplasia were seen in lung sections. The alveolar septa were multifocally expanded by a few lymphocytes. Mild lymphocytic periportal hepatitis and interstitial nephritis were found. The lymphoid nodules of the large intestine showed marked lymphocytic hyperplasia. Infection by SARS-CoV-2 was established by viral RNA detection in a pool of nasopharyngeal and oropharyngeal swabs and liver samples. Immunohistochemistry detected the viral nucleocapsid protein in sections of lung, liver, spleen, lymph nodes and large intestine, and spike protein antigen in lung tissue. This is the first report of naturally occurring SARS-CoV-2 infection in a New World monkey. Platyrrhine species should be included as potential hosts of natural infection of SARS-CoV-2.

Molecular detection of Brucella abortus in wild and captive felids.

PURPOSE: Brucellosis is a zoonotic disease of great public health importance. In wild animals, Brucella abortus is one of the most diagnosed species, mainly in enzootic environments where domestic animals share the same environment. B. abortus is common in environments shared by cattle, wild, and domestic animals. This study aimed to detect the presence of B. abortus DNA in free-ranging and captivity felids at Mato Grosso State, Brazil. METHOD: Polymerase chain reaction, based on the genetic element IS711, was performed in blood samples collected from 23 free-ranging and captive felids. The species represented include Leopardus colocolo, Leopardus pardalis, Leopardus wiedii, Panthera onca, Puma concolor, and Puma yagouaroundi. RESULTS: DNA amplification of B. abortus was observed in only one captive P. concolor (4.34%). CONCLUSION: The detection of this pathogen in captive animals using molecular tools demonstrates the importance of monitoring, as it raises concerns about the possibility of transmission between humans and wild and domestic animals, especially in regions of vast biodiversity, such as in the State of Mato Grosso, Brazil.

Molecular detection of Toxoplasma gondii and Neospora caninum in neotropical primates.

BACKGROUND: Toxoplasma gondii and Neospora caninum infections in primates are potentially fatal and directly impact the conservation of these animals and public health. MATERIALS AND METHODS: A total of 38 blood/clot samples collected from free-living and captive neotropical primates undergoing clinical care or found dead by environmental authorities in the Mato Grosso State, Brazil, were analyzed by PCR for DNA detection of T. gondii and N. caninum. Furthermore, eight animals were submitted to immunohistochemistry for the detection of T. gondii. RESULTS: DNA of T. gondii and N. caninum was amplified in 11 (28.95%) 10 (26.32%) of samples analyzed, respectively. Coinfection was observed in three individuals. One animal returned a positive result in the immunohistochemistry for the detection of T. gondii. CONCLUSION: These findings reflect a concern for the conservation of these animals, as the pathogen-host interaction is unpredictable and infections by these protozoa can lead to animal mortality, which has a substantial impact on endangered species.

Persistent SARS-CoV-2 antigen presence in multiple organs of a naturally infected cat from Brazil.

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the etiological agent of the disease coronavirus 2019 (COVID-19) in humans. SARS-CoV-2 has been identified in cats with or without clinical signs. Case presentation: We describe the pathological and molecular findings in a six-month-old asymptomatic cat with SARS-CoV-2 infection from Brazil, belonging to a human family with COVID-19 cases. The pool of nasopharynx and oropharynx swabs at day zero tested positive by RT-qPCR for SARS-CoV-2. No amplification resulted from molecular testing performed on days 7 and 14. The cat was hit by a car and died 43 days after the molecular diagnosis. Immunohistochemistry at post-mortem examination demonstrated nucleocapsid protein in samples from the lungs, kidneys, nasal conchae, trachea, intestine, brain and spleen. Conclusion: The present study has highlighted the possibility that viral antigens can be detected by immunohistochemistry in multiple organs six weeks after infection, although the same tissues tested negative by RT-PCR.

Clinical and genomic data of sars-cov-2 detected in maternal-fetal interface during the first wave of infection in Brazil.

Brazil has the highest SARS-CoV-2 case-fatality rate in pregnant women in the Americas. In this study, clinical and virological findings of five mildly symptomatic pregnant women and their infected fetuses/newborns treated at a referral hospital for COVID19-pregnant women in Midwestern Brazil are reported. Mother and fetal samples were tested by RT-qPCR, ECLIA and Illumina MiSeq sequencing. From the five cases, one resulted in spontaneous abortion, one was stillborn, two were preterm births and one full-term birth. Maternal and fetal placenta, newborn and stillborn secretions were SARS-CoV-2+; one neonate developed ground-glass opacities in his lungs. One neonate's umbilical cord was IgG+ and all were IgM negative upon hospital discharge. Genomes recovered from two placentas belong to the B.1.1.28 and B.1.1.33 lineages and present nonsynonymous mutations associated with virus fitness and infectivity; other not frequently reported mutations (B.1.1.33: NSP3 V2090G, M A2S and ORF3ab S253P and Y264N; B.1.1.28: NSP3 E995D, NSP12 R240K, NSP14H1897Y and in ORF7b V21F) were found in proteins involved in viral replication, viral induction of apoptosis, viral interference on interferon and on NF-Κß pathways. Phylogeny indicates the south of Brazil as the possible origin of these lineages circulating in Mato Grosso State. These findings contribute to describe SARS-CoV-2 infection and outcomes in pregnant women and their fetuses, at any stage of gestation and even in mild symptomatic cases.

Canine gastrointestinal pythiosis: a case report in Brazilian Pantanal from a diagnostic approach / Pitiose gastrointestinal canina: relato de caso no Pantanal brasileiro a partir de uma abordagem diagnóstica

Pythiosis is caused by an aquatic fungus-like organism (Pythium insidiosum). It is considered an important public health issue as it can affect both animals and humans. This paper reports a case of gastrointestinal pythiosis in a dog. The patient was hospitalized for four days, during which the animal received supportive and symptomatic treatment. But the applied treatment was unsuccessful and the dog's clinical condition worsened, culminating in death. Complementary imaging tests such as radiography and ultrasonography, as well as hematological tests, were performed during the hospitalization period. The definitive diagnosis was reached in the postmortem as macroscopic and microscopic characteristics suggested the presence of intestinal granuloma and accentuated multifocal suppurative necrotic enteritis. Additionally, the histological evaluation revealed morphological structures compatible with P. insidiosum. Also, the results of nested PCR performed showed partial amplification (105 bp) of the ITS1 region of the ribosomal gene of P. insidiosum.(AU)

A pitiose é causada por um organismo aquático semelhante a um fungo (Pythium insidiosum) e considerada um importante problema de saúde pública, pois pode afetar animais e humanos. Este artigo relata um caso de pitiose gastrointestinal em um cão. O paciente ficou internado por quatro dias, período em que o animal recebeu tratamento de suporte e sintomático. No entanto, o tratamento aplicado não teve sucesso e o quadro clínico do cão piorou, culminando com a morte. Exames de imagem complementares, como radiografia e ultrassonografia, bem como exames hematológicos, foram realizados durante o período de internação. O diagnóstico definitivo foi feito na autópsia, pois as características macroscópicas e microscópicas sugeriam a presença de granuloma intestinal e acentuada enterite necrótica multifocal supurativa. Além disso, a avaliação histológica revelou estruturas morfológicas compatíveis com P. insidiosum. Além disso, a nested PCR foi realizada e mostrou amplificação parcial (105 pb) da região ITS1 do gene ribossomal de P. insidiosum.(AU)

A Novel Host of an Emerging Disease: SARS-CoV-2 Infection in a Giant Anteater (Myrmecophaga tridactyla) Kept Under Clinical Care in Brazil.

A young male free-ranging giant anteater (Myrmecophaga tridactyla) was found with paralysis of pelvic limbs on a highway and kept under human care. Radiographs confirmed multiple incomplete fractures in the thoracolumbar vertebrae. Due to the poor prognosis, euthanasia was chosen. The infection was established by viral SARS-CoV-2 RNA detection in the rectal swab, spleen and kidney samples. Immunohistochemistry detected the viral nucleocapsid protein in sections of the lungs, liver, spleen, lymph nodes, and large intestine sections, and spike protein antigen in the lung tissue. Pilosa order species should be included as potential hosts of natural infection of SARS-CoV-2.

Successful potassium iodide treatment for rhinofacial pythiosis in sheep.

BACKGROUND: Pythiosis in sheep is an important disease in Brazil, which could cause rhinitis, dermatitis and alimentary tract inflammation. It is caused by the aquatic oomycete, Pythium insidiosum. The rhinofacial pythiosis causes facial deformity and upper respiratory tract clinical signs associated with necroproliferative masses occupying the rostral nasal cavity and hard palate. Little is known regarding the therapy, prophylaxis and pathogenesis of this disease. METHODOLOGY: During the 6-year study, we examined 13 sheep presenting rhinofacial pythiosis. The diagnosis was performed through biopsy of the rhinofacial lesions followed by histopathology and immunohistochemistry using specific antibodies against P insidiosum, polymerase chain reaction and an indirect enzyme-linked immunosorbent assay. RESULTS: This study presents the clinical findings of a potassium iodide treatment of rhinofacial pythiosis in sheep. All sheep were treated with 10 ml of 10% potassium iodide solution, administered orally every day during 63-120 (mean 85) days. Among treated sheep, 84.6% demonstrated complete recovery. CONCLUSION: Potassium iodide therapy may treat rhinofacial pythiosis in sheep.

Domestic Birds as Source of Cryptococcus deuterogattii (AFLP6/VGII): Potential Risk for Cryptococcosis.

Cryptococcosis is an infection caused by encapsulated basidiomycetous yeasts belonging to the Cryptococcus neoformans/Cryptococcus gattii species complexes. It is acquired through inhalation of infectious propagules, often resulting in meningitis and meningoencephalitis. The ecological niche of these agents is a wide variety of trees species, as well as pigeon, parrot and passerine excreta. The objective of this study was to isolate Cryptococcus yeasts from excreta of commercially traded parrots and passerines. The 237 samples were collected between October 2018 and April 2019 and processed using conventional methodologies. Nineteen colonies with a dark brown phenotype, caused by phenol oxidase activity, were isolated, suggesting the presence of pathogenic Cryptococcus yeasts. All isolates tested positive for urease activity. URA5-RFLP fingerprinting identified 14 isolates (68.4%) as C. neoformans (genotype AFLP1/VNI) and 5 (26.3%) as C. deuterogattii (genotype AFLP6/VGII). Multi-locus sequence typing was applied to investigate the relatedness of the C. deuterogattii isolates with those collected globally, showing that those originating from bird-excreta were genetically indistinguishable from some clinical isolates collected during the past two decades.

Persistent SARS-CoV-2 antigen presence in multiple organs of a naturally infected cat from Brazil

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the etiological agent of the disease coronavirus 2019 (COVID-19) in humans. SARS-CoV-2 has been identified in cats with or without clinical signs. Case presentation: We describe the pathological and molecular findings in a six-month-old asymptomatic cat with SARS-CoV-2 infection from Brazil, belonging to a human family with COVID-19 cases. The pool of nasopharynx and oropharynx swabs at day zero tested positive by RT-qPCR for SARS-CoV-2. No amplification resulted from molecular testing performed on days 7 and 14. The cat was hit by a car and died 43 days after the molecular diagnosis. Immunohistochemistry at post-mortem examination demonstrated nucleocapsid protein in samples from the lungs, kidneys, nasal conchae, trachea, intestine, brain and spleen. Conclusion: The present study has highlighted the possibility that viral antigens can be detected by immunohistochemistry in multiple organs six weeks after infection, although the same tissues tested negative by RT-PCR.(AU)