Retraction Note to: Comparative analysis of the replication of bovine herpesvirus 1 (BHV1) and BHV5 in bovine-derived neuron-like cells.

The Editor-in-Chief has retracted this article.

Retraction Note to: Mitochondrial bioenergy alterations in avian HD11 macrophages infected with infectious bronchitis virus.

The Editor-in-Chief has retracted this article [1]. Figures 1A, 1D and 2B (bottom right) are identical with Figures 1A, 1H and 1B respectively in another article [2] which reports a study in a different species. In addition, Table 1 contains data presented in a third article [3], which also reports a study in a different species. The Editor-in-Chief therefore no longer has confidence in the validity of the data and the conclusions drawn. Tereza C. Cardoso disagrees with this retraction. Helena L. Ferreira agrees with this retraction. Sergio E. L. da Silva, Andrea F. Garcia, Felipe E. S. Silva, Roberto Gameiro, Carolina U. F. Fabri and Dielson S. Vieira have not responded to any correspondence about this retraction.

Mitochondrial bioenergy alterations in avian HD11 macrophages infected with infectious bronchitis virus.

To establish an association between mitochondrial dysfunction and apoptosis following infectious bronchitis virus (IBV) infection, HD11 avian macrophage cells were infected with the Massachusetts 41 (M41) strain. Our results show that the M41 strain of IBV induced cytopathic effects followed by the release of new viral particles. Elevated numbers of apoptotic cells were observed at 24, 48 and 72 h post-infection (p.i.). Viral infection was associated with mitochondrial membrane depolarization and reactive oxygen species (ROS) production at all of the examined timepoints p.i. In summary, IBV M41 replication in infected HD11 macrophages seems to induce mitochondrial bioenergy failure, acting as a respiratory chain uncoupler, without compromising viral replication.

Avian coronavirus isolated from a pigeon sample induced clinical disease, tracheal ciliostasis, and a high humoral response in day-old chicks.

The detection of avian coronaviruses (AvCoV) in wild birds and the emergence of new AvCoV have increased in the past few years. In the present study, the pathogenicity of three AvCoV isolates was investigated in day-old chicks. One AvCoV isolated from a pigeon, which clustered with the Massachusetts vaccine serotype, and two AvCoV isolated from chickens, which grouped with a Brazilian genotype lineage, were used. Clinical signs, gross lesions, histopathological changes, ciliary activity, viral RNA detection, and serology were evaluated during 42 days post infection. All AvCoV isolates induced clinical signs, gross lesions in the trachea, moderate histopathological changes in the respiratory tract, and mild changes in other tissues. AvCoV isolated from the pigeon sample caused complete tracheal ciliostasis over a longer time span. Specific viral RNA was detected in all tissues, but the highest RNA loads were detected in the digestive tract (cloacal swabs and ileum). The highest antibody levels were also detected in the group infected with an isolate from the pigeon. These results confirm the pathogenicity of Brazilian variants, which can cause disease and induce gross lesions and histopathological changes in chickens. Our results suggest that non-Galliformes birds can also play a role in the ecology of AvCoV.

Isolation, characterization and immunomodulatory-associated gene transcription of Wharton's jelly-derived multipotent mesenchymal stromal cells at different trimesters of cow pregnancy.

The possibility of isolating bovine mesenchymal multipotent stromal cells (MSCs) from fetal adnexa is an interesting prospect due to the potential use of these cells in biotechnological applications. However, little is known about the properties of these progenitor cells in bovine species. Wharton's jelly (WJ) MSC cells were obtained from the umbilical cord of bovine fetuses at three different stages of pregnancy and divided into groups 1, 2 and 3 according to gestational trimester. Cell morphology, from the three stages of pregnancy, typically appeared fibroblast-like spindle-shaped, presenting the same viability and number. Moreover, the proliferative ability of T-cells in response to a mitogenic stimulus was suppressed when WJMSC cells were added to the culture. Multilineage properties were confirmed by their ability to undergo adipogenic, osteogenic/chondrogenic and neurogenic differentiation. Mesenchymal phenotyping, CD105+, CD29+, CD73+ and CD90+ cell markers were detected in all three cell groups, yet these markers were considered more expressed in MSCs of group 2 (p < 0.005). Expression of cytokines IL2, IL6RR, INFAC, INFB1, IFNG, TNF and LTBR were downregulated, whereas IL1F10 expression was upregulated in all tested WJMSCs. The present study demonstrated that WJMSCs harvested from the bovine umbilical cord at different gestational stages showed proliferative capacity, immune privilege and stemness potential.

Expression of miR-155 associated with Toll-like receptors 3, 7, and 9 transcription in the olfactory bulbs of cattle naturally infected with BHV5.

Bovine herpesvirus 5 (BHV5) infection of young cattle is frequently associated with fatal neurological disease and, as such, represents an attractive model for studying the pathogenesis of viral-induced meningoencephalitis. Following replication in the nasal mucosa, BHV5 invades the central nervous system (CNS) mainly through the olfactory pathway. The innate immune response triggered by the host face to virus replication through the olfactory route is poorly understood. Recently, an upregulation of conserved pathogen-associated molecular pattern, as Toll-like receptors (TLRs), has been demonstrated in the CNS of BHV5 experimentally infected cows. A new perspective to understand host-pathogen interactions has emerged elucidating microRNAs (miRNAs) network that interact with innate immune response during neurotropic viral infections. In this study, we demonstrated a link between the expression of TLRs 3, 7, and 9 and miR-155 transcription in the olfactory bulbs (OB) of 16 cows suffering from acute BHV5-induced neurological disease. The OBs were analyzed for viral antigens and genome, miR-155 and TLR 3, 7, and 9 expression considering three major regions: olfactory receptor neurons (ORNs), glomerular layer (GL), and mitral cell layer (ML). BHV5 antigens and viral genomes, corresponding to glycol-C gene, were detected in all OBs regions by fluorescent antibody assay (FA) and PCR, respectively. TLR 3, 7, and 9 transcripts were upregulated in ORNs and ML, yet only ORN layers revealed a positive correlation between TLR3 and miR-155 transcription. In ML, miR-155 correlated positively with all TLRs studied. Herein, our results evidence miR-155 transcription in BHV5 infected OB tissue associated to TLRs expression specifically ORNs which may be a new window for further studies.

Bovine herpesviruses induce different cell death forms in neuronal and glial-derived tumor cell cultures.

Oncolytic viruses have the ability to infect tumor cells and leave healthy cells intact. In this study, bovine herpesvirus 1 (BHV1; Los Angeles, Cooper, and SV56/90 strains) and bovine herpesvirus 5 (BHV5; SV507/99 and GU9457818 strains) were used to infect two neuronal tumor cell lineages: neuro2a (mouse neuroblastoma cells) and C6 (rat glial cells). BHV1 and BHV5 strains infected both cell lines and positively correlated with viral antigen detection (p < 0.005). When neuro2a cells were infected by Los Angeles, SV507/99, and GU9457818 strains, 40 % of infected cells were under early apoptosis and necroptosis pathways. Infected C6 cells were >40 % in necroptosis phase when infected by BHV5 (GU9457818 strain). Blocking caspase activation did not interfere with cell death. However, when necroptosis was blocked, 60-80 % of both infected cells with either virus switched to early apoptosis pathway with no interference with virus replication. Moreover, reactive oxygen species production and mitochondrial membrane dysfunction were detected at high levels in both infected cell lines. In spite of apoptosis and necroptosis blockage, tumor necrosis factor alpha (TNFA) and virus transcription were positively correlated for all viral strains studied. Thus, these results contribute to the characterization of BHV1 and BHV5 as potential oncolytic viruses for non-human cells. Nonetheless, the mechanisms underlying their oncolytic activity in human cells are still to be determined.

Comparative analysis of the replication of bovine herpesvirus 1 (BHV1) and BHV5 in bovine-derived neuron-like cells.

Members of the subfamily Alphaherpesvirinae use the epithelium of the upper respiratory and/or genital tract as preferential sites for primary replication. However, bovine herpesvirus 5 (BoHV5) is neurotropic and neuroinvasive and responsible for meningoencephalitis in cattle and in animal models. A related virus, BoHV1 has also been occasionally implicated in natural cases of neurological infection and disease in cattle. The aim of the present study was to assess the in vitro effects of BoHV1 and BoHV5 replication in neuron-like cells. Overall, cytopathic effects, consisting of floating rounded cells, giant cells and monolayer lysis, induced by both viruses at 48 h postinfection (p.i.) resulted in a loss of cell viability and high virus titres (r = 0.978). The BoHV1 Cooper strain produced the lowest titres in neuron-like cells, although viral DNA was detected in infected cells during all experiments. Virus replication in infected cells was demonstrated by immunocytochemistry, flow cytometry and qPCR assays. BoHV antigens were better visualized at 48 h p.i. and flow cytometry analysis showed that SV56/90 and Los Angeles antigens were present at higher levels. In spite of the fact that BoHV titres dropped at 48 h p.i, viral DNA remained detectable until 120 h p.i. Sensitive TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling) and annexin V assays were used to identify apoptosis. BoHV5 induced death in approximately 50% of cells within 24 h p.i., similar to what has been observed for BoHV1 Los Angeles. Infection with the BoHV1 Cooper strain resulted in 26.37% of cells being in the early stages of apoptosis; 63.69% of infected cells were considered viable. Modulation of mitochondrial function, as measured by mitochondrial membrane depolarization, was synchronous with the virus replication cycle, cell viability and virus titres at 48 h p.i. Our results indicate that apoptosis plays an important role in preventing neuronal death and provides a bovine-derived in vitro system to study herpesvirus-neuron interactions.

Cellular vacuolation and mitochondrial-associated factors induced by Clostridium perfringens epsilon toxin detected using acoustic flow cytometry.

Epsilon toxin (ETX) produced by Clostridium perfringens types B and D is a potent toxin that is responsible for fatal enterotoxaemia. In vitro, ETX, which is considered as a pore-forming toxin, forms a heptamer in Madin-Darby canine kidney (MDCK) cell membranes, which is considered to be a pre-pore stage. After binding of the ETX, vacuoles inside cell cytoplasm are produced. ETX causes decreased levels of essential coenzymes required for host cell energy. Here, we optimized and applied acoustic flow cytometry analysis in order to gain further insight into ETX-pathogenesis. Using acoustic flow cytometer analysis, which considered highly sensitive, ETX-exposed MDCK cells revealed mitochondrial membrane decreases followed by 25.48% and 45.45% of the exposed cells expressing the Bax and BCL-2 proteins at a pre-pore stage, respectively. These results together with high cytotoxicity and visualization of cell vacuoles, demonstrates that acoustic flow cytometry analysis potentially represents an effective tool to study ETX pathogenesis.

Isolation and characterization of Wharton's jelly-derived multipotent mesenchymal stromal cells obtained from bovine umbilical cord and maintained in a defined serum-free three-dimensional system.

BACKGROUND: The possibility for isolating bovine mesenchymal multipotent cells (MSCs) from fetal adnexa is an interesting prospect because of the potential for these cells to be used for biotechnological applications. Bone marrow and adipose tissue are the most common sources of MSCs derived from adult animals. However, little knowledge exists about the characteristics of these progenitors cells in the bovine species. Traditionally most cell cultures are developed in two dimensional (2D) environments. In mammalian tissue, cells connect not only to each other, but also support structures called the extracellular matrix (ECM). The three-dimensional (3D) cultures may play a potential role in cell biotechnology, especially in tissue therapy. In this study, bovine-derived umbilical cord Wharton's jelly (UC-WJ) cells were isolated, characterized and maintained under 3D-free serum condition as an alternative of stem cell source for future cell banking. RESULTS: Bovine-derived UC-WJ cells, collected individually from 5 different umbilical cords sources, were successfully cultured under serum-free conditions and were capable to support 60 consecutive passages using commercial Stemline(®) mesenchymal stem cells expansion medium. Moreover, the UC-WJ cells were differentiated into osteocytes, chondrocytes, adipocytes and neural-like cells and cultured separately. Additionally, the genes that are considered important embryonic, POU5F1 and ITSN1, and mesenchymal cell markers, CD105(+), CD29(+), CD73(+) and CD90(+) in MSCs were also expressed in five bovine-derived UC-WJ cultures. Morphology of proliferating cells typically appeared fibroblast-like spindle shape presenting the same viability and number. These characteristics were not affected during passages. There were 60 chromosomes at the metaphase, with acrocentric morphology and intense telomerase activity. Moreover, the proliferative capacity of T cells in response to a mitogen stimulus was suppressed when bovine-derived UC-WJ cells was included in the culture which demonstrated the immunossupression profile typically observed among isolated mesenchymal cells from other species. After classified the UC-WJ cells as mesenchymal stromal phenotype the in vitro 3D cultures was performed using the AlgiMatrix(®) protocol. Based on the size of spheroids (283,07 µm ± 43,10 µm) we found that three weeks of culture was the best period to growth the UC-WJ cells on 3D dimension. The initial cell density was measured and the best value was 1.5 × 10(6) cells/well. CONCLUSIONS: We described for the first time the isolation and characterization of UC-WJ cells in a serum-free condition and maintenance of primitive mesenchymal phenotype. The culture was stable under 60 consecutive passages with no genetic abnormalities and proliferating ratios. Taken together all results, it was possible to demonstrate an easy way to isolate and culture of bovine-derived UC-WJ cells under 2D and 3D serum-free condition, from fetal adnexa with a great potential in cell therapy and biotechnology.

Evaluation of developmental changes in bovine in vitro produced embryos following exposure to bovine Herpesvirus type 5.

BACKGROUND: Bovine Herpesvirus type-5 (BoHV-5) is a neurovirulent α-Herpesvirus which is potentially pathogenic for cows and suspected to be associated with reproductive disorders. Interestingly, natural transmission of BoHV-5 by contaminated semen was recently described in Australia. Additionally, BoHV-5 was also isolated from the semen of a healthy bull in the same country and incriminated in a natural outbreak of reproductive disease after artificial insemination. In contrast with BoHV-1, experimental exposure of in vitro produced bovine embryos to BoHV-5 does not affect embryo viability and seems to inhibit some pathways of apoptosis. However, the mechanisms responsible for these phenomena are poorly understood. In this study, we examined mitochondrial activity, antioxidant protection, stress response and developmental rates of in vitro produced bovine embryos that were exposed and unexposed to BoHV-5. METHODS: For this purpose, bovine embryos produced in vitro were assayed for cell markers after experimental infection of oocytes (n = 30; five repetitions), in vitro fertilization and development. The indirect immunofluorescence was employed to measure the expression of superoxide dismutase 1 (SOD1), anti-oxidant like protein 1 (AOP-1), heat shock protein 70.1 (Hsp 70.1) and also viral antigens in embryos derived from BoHV-5 exposed and unexposed oocytes. The determination of gene transcripts of mitochondrial activity (SOD1), antioxidant protection (AOP-1) and stress response (Hsp70.1) were evaluated using the reverse transcriptase polymerase chain reaction (RT-PCR). MitoTracker Green FM, JC-1 and Hoechst 33342-staining were used to evaluate mitochondrial distribution, segregation patterns and embryos morphology. The intensity of labeling was graded semi-quantitatively and embryos considered intensively marked were used for statistical analysis. RESULTS: The quality of the produced embryos was not affected by exposure to BoHV-5. Of the 357 collected oocytes, 313 (+/- 6.5; 87.7%) were cleaved and 195 (+/- 3.2; 54.6%) blastocysts were produced without virus exposure. After exposure, 388 oocytes were cleaved into 328 (+/- 8.9, 84.5%), and these embryos produced 193 (+/- 3.2, 49.7%) blastocysts. Viral DNA corresponding to the US9 gene was only detected in embryos at day 7 after in vitro culture, and confirmed by indirect immunofluorescence assay (IFA). These results revealed significant differences (p < 0.05) between exposed and unexposed oocytes fertilized, as MitoTracker Green FM staining Fluorescence intensity of Jc-1 staining was significantly higher (p < 0.005) among exposed embryos (143 +/- 8.2). There was no significant difference between the ratios of Hoechst 33342-stained nuclei and total cells in good-quality blastocysts (in both the exposed and unexposed groups). Using IFA and reverse transcriptase polymerase chain reaction (RT-PCR) for the set of target transcripts (SOD1, AOP-1 and Hsp 70.1), there were differences in the mRNA and respective proteins between the control and exposed embryos. Only the exposed embryos produced anti-oxidant protein-like 1 (AOP-1). However, neither the control nor the exposed embryos produced the heat shock protein Hsp 70.1. Interestingly, both the control and the exposed embryos produced superoxide dismutase (SOD1), revealing intense mitochondrial activity. CONCLUSION: This is the first demonstration of SOD1 and AOP-1 production in bovine embryos exposed to BoHV-5. Intense mitochondrial activity was also observed during infection, and this occurred without interfering with the quality or number of produced embryos. These findings further our understanding on the ability of α-Herpesviruses to prevent apoptosis by modulating mitochondrial pathways.

Susceptibility of neuron-like cells derived from bovine Wharton's jelly to bovine herpesvirus type 5 infections.

BACKGROUND: Bovine herpesvirus type 5 (BoHV-5), frequently lethal in cattle, is associated with significant agricultural economic losses due to neurological disease. Cattle and rabbits are frequently used as models to study the biology and pathogenesis of BoHV-5 infection. In particular, neural invasion and proliferation are two of the factors important in BoHV-5 infection. The present study investigated the potential of bovine Wharton's jelly mesenchymal stromal cells (bWJ-MSCs) to differentiate into a neuronal phenotype and support robust BoHV-5 replication. RESULTS: Upon inducing differentiation within a defined neuronal specific medium, most bWJ-MSCs acquired the distinctive neuronal morphological features and stained positively for the neuronal/glial markers MAP2 (neuronal microtubule associated protein 2), N200 (neurofilament 200), NT3 (neutrophin 3), tau and GFAP (glial fibrillary acidic protein). Expression of nestin, N200, ß-tubulin III (TuJI) and GFAP was further demonstrated by reverse transcriptase polymerase chain reaction (RT-PCR). Following BoHV-5 inoculation, there were low rates of cell detachment, good cell viability at 96 h post-infection (p.i.), and small vesicles developed along neuronal branches. Levels of BoHV-5 antigens and DNA were associated with the peak in viral titres at 72 h p.i. BoHV-5 glycoprotein C mRNA expression was significantly correlated with production of progeny virus at 72 h p.i. (p < 0.05). CONCLUSION: The results demonstrated the ability of bWJ-MSCs to differentiate into a neuronal phenotype in vitro and support productive BoHV-5 replication. These findings constitute a remarkable contribution to the in vitro study of neurotropic viruses. This work may pave the way for bWJ-MSCs to be used as an alternative to animal models in the study of BoHV-5 biology.

Visual detection of turkey coronavirus RNA in tissues and feces by reverse-transcription loop-mediated isothermal amplification (RT-LAMP) with hydroxynaphthol blue dye.

A sensitive reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for the rapid visual detection of turkey coronavirus (TCoV) infection. The reaction is performed in one step in a single tube at 65 °C for 45 min, with hydroxynaphthol blue (HNB) dye added prior to amplification. The detection limit of the RT-LAMP assay was approximately 10(2) EID(50/50 µl) TCoV genome, and no cross-reaction with other avian viruses was observed. The assay was evaluated further in tissue suspensions prepared from the ileum and ileum-caecal junctions of infected turkey embryos; 100% of these samples were positive in the RT-LAMP assay. All individual feces samples collected in the field were considered positive by both conventional RT-PCR and RT-LAMP. In conclusion, RT-LAMP with HNB dye was shown to be a sensitive, simple assay for the rapid diagnosis of TCoV infection, either directly from feces or in association with virus isolation methods.

RETRACTED: Bovine Herpesvirus 5 promotes mitochondrial dysfunction in cultured bovine monocyte-derived macrophages and not affect virus replication.

This article has been retracted: please see Elsevier Policy on Article Withdrawal (https://www.elsevier.com/about/our-business/policies/article-withdrawal). This article has been retracted at the request of the Editors-in-Chief and Authors. Fig 1A is a duplicate of a figure that has already been published in da Silva SEL et al. Archives of Virology 2018;163:1043-1049; 10.1007/s00705-018-3704-2. These two papers report studies performed with cells from two different animal species (bovine cells for the Veterinary Microbiology paper and chicken cells for the Archives of Virology paper). The reuse of the same figure in the Veterinary Microbiology paper to describe cells that were supposed to be from a different species is thus inappropriate and also puts into question the reliability of the other results presented in this paper. In addition, the Editors-in-Chief have remaining concerns about the strong similarities of other data presented in the two papers. Even if these concerns were addressed, the re-use of any data has to be clearly indicated and appropriately cited. As such this article represents a misuse of the scientific publishing system. The scientific community takes a very strong view on this matter and apologies are offered to readers of the journal that this was not detected during the submission process.

Direct detection of infectious bursal disease virus from clinical samples by in situ reverse transcriptase-linked polymerase chain reaction.

The presence of the very virulent (vv) Brazilian strain of infectious bursal disease virus (IBDV) was determined in the bursa of Fabricius, thymus and liver of 2-week-old broilers from a flock with a higher than expected mortality. For this purpose, a direct in situ reverse transcriptase (RT)-linked polymerase chain reaction (PCR) method was developed using specific primers for vvIBDV. Unlabelled forward and reverse biotinylated oligonucleotides were used for RT-PCR in a one-step method and the respective products were revealed by a direct enzymatic reaction. The results were compared with those obtained by standard RT-PCR using general primers for IBDV and virus isolation. The virus isolation, RT-PCR and in situ RT-PCR revealed positive results on the bursa of Fabricius in 86%, 80% and 100%, respectively. The in situ RT-PCR detected vvIBDV in all tested thymus and liver samples, whereas the standard RT-PCR detected virus in 80% and 90% of the samples, respectively. After three consecutive passages on chicken embryonated eggs, IBDV was isolated from 64% of the thymus samples and 30% of the liver samples. In the present study, no classical or antigenic variants of IBDV were detected. The developed in situ RT-PCR assay was able to detect the very virulent strain of IBDV with a higher sensitivity than the conventional RT-PCR and virus isolation.

Detection of Turkey astrovirus in young poults affected with poult enteritis complex in Brazil.

Reverse transcription polymerase chain reaction (RT-PCR) of turkey astrovirus (TAstV) capsid and polymerase genes was applied to the bursa of Fabricius (BF), thymus (TH), spleen (SP) and cloacal swabs (CS) of young poults with "Poult enteritis complex" (PEC). The histological lesions included atrophy, lymphoid depletion, cellular infiltration and necrosis of the BF, TH and SP, respectively. The RT-PCR reactions were positive for the polymerase gene of TAstV-2 in all 100 CSs, 7 out of 10 of BFs and 10 out of 20 THs and SPs, respectively. Five out of 10 THs and SPs samples, considered to be negative by RT-PCR, were positive when specific primers designed for the TAstV-2 capsid gene were applied. This is the first description of turkey astrovirus infection presenting PEC in Latin America.

Genotyping of Clostridium perfringens isolated from calves with neonatal diarrhea.

Neonatal diarrhea is one of the main causes of losses in cattle herds. Clostridium perfringens is a widespread enteropathogen, and is responsible for many animal diseases such as bovine neonatal diarrhea. Fecal samples from 141 diarrheic calves and 129 healthy calves, aged up to 28 days and belonging to three herds were examined. Rates of culture positivity were 36.2% and 30.2% for diarrheic and nondiarrheic calves, respectively. Multiple isolates from primary isolation plates were subjected to simultaneous genotyping by multiplex PCR, with primers amplifying fragments of alpha (cpa), beta (cpb), epsilon (etx), iota (itxA), enterotoxin (cpe) and beta2 (cpb2) toxin-encoding genes. Only 17/51 (33.3%) and 17/39 (43.6%) of these mixtures from diarrheic and nondiarrheic calves, respectively, yielded genotype information, suggesting that this may not be a viable approach to genotyping of isolates. Fourteen isolate mixtures from animals with diarrhea had only cpa (type A), one had cpa and cpb2 (type A beta2 positive), one with cpa, itxA, and cpb2 (type E, beta2 positive), and one with cpa, etx, itxA, and cpb2 toxin producing strains. Among 17 isolate mixtures from healthy calves, 10 were exclusively type A, one was type A cpb2 positive, two were type E, three were type E cpb2 positive, and one was types D and E cpb2 positive. There was no correlation between isolation of a given toxin type and the presence of diarrhea.

Circulation of canine parvovirus among dogs living in human-wildlife interface in the Atlantic forest biome, Brazil.

Despite of the role of domestic dogs as reservoirs for threatening viral diseases for wild carnivores, few studies have focused to identify circulation of viruses among dogs living in human/wildlife interfaces. To identify canine parvovirus (CPV) types circulating in dogs living in an Atlantic forest biome, faecal samples (n = 100) were collected at the same period (one week) corresponding to each of four areas, during 2014 to 2016 and corresponded to 100 different individuals. CPV was isolated in cell culture from 67 out 100 (67%) samples from healthy dogs. Cytopathic effects were characterized by total or partial cell culture lysis. Genome sequences of CPV-2a (10%), CPV-2b (7%) and CPV-2c (50%) were concomitantly detected by PCR and nucleotide sequencing. The current study addresses the importance of monitoring CPV circulation among dogs presenting potential contact with wildlife species.

Complete Genome Sequence of an Avian Metapneumovirus Subtype A Strain Isolated from Chicken (Gallus gallus) in Brazil.

We report here the complete genome sequence of an avian metapneumovirus (aMPV) isolated from a tracheal tissue sample of a commercial layer flock. The complete genome sequence of aMPV-A/chicken/Brazil-SP/669/2003 was obtained using MiSeq (Illumina, Inc.) sequencing. Phylogenetic analysis of the complete genome classified the isolate as avian metapneumovirus subtype A.

Osteogenic potential of mesenchymal cells derived from canine umbilical cord matrix co-cultured with platelet-rich plasma and demineralized bone matrix.

Canine mesenchymal cells (MSCs) derived from Wharton's jelly were co-cultured, then supplemented or not supplemented with platelet rich plasma (PRP) and demineralized bone matrix (DBM) to verify osteogenic differentiation. Osteoblastic differentiation followed by mineralized bone matrix production was found to be significantly higher (p < 0.05) when MSCs were associated with PRP/DBM in culture after 14-21-days of induction. Osteopontin and osteocalcin gene expression were significantly superior (p < 0.05) under the same culture conditions after 21 days of observation. In conclusion, addition of PRP to DBM co-cultured with MSCs successfully induced osteogenesis in vitro.