Bruno S.A.F. Brasil

DNA barcoding unveils a high rate of mislabeling in a commercial freshwater catfish from Brazil

Background and Aims. Molecular markers have contributed to species authentication by flagging mislabeling and the misidentification of commercial landings. Such tools are of great value since the market substitution of fish of lower value for highly commercialized species is expected to become more pronounced due to a shortage of natural stocks. Materials and Methods. Here we report on the molecular identification 4results from processed fish products (i.e. fillets) and whole fishes sold in Brazilian markets under the common name surubim (Pseudoplatystoma spp.). Results. DNA barcoding revealed the incorrect labeling of around 80% of all samples analyzed, with mislabeling being more pronounced within fillets rather than whole fish. Conclusion. To our knowledge, this is the first report correlating the rate of fraud with processed fish products. The establishment of an official list of acceptable common names for freshwater fish and seafood is urgently needed in Brazil for further trade regulations to take place.

Survey of pyrethroid and organophosphate resistance in Brazilian field populations of Rhipicephalus (Boophilus) microplus: Detection of C190A mutation in domain II of the para-type sodium channel gene

The cattle tick Rhipicephalus (Boophilus) microplus causes expressive damage to livestock in Brazil and other countries. Its control is becoming more difficult due to the development of resistance in populations. Early detection of resistance can help in developing effective control strategies. This study evaluated the susceptibility of R. microplus to cypermethrin and chlorpyriphos and was the first attempt to identify the mechanism of resistance (target site insensitivity) in cattle tick populations from Minas Gerais state (Southeastern Brazil). Engorged female ticks were collected from 10 ranches within the state of Minas Gerais, and susceptibility was evaluated with the larval packet test (LPT) using technical grade cypermethrin and chlorpyriphos. It was possible to analyze LPT results of seven populations. Target site insensitivity was investigated in all 10 isolates by using molecular approaches for detection of the T2134A substitution within the domain III S6 segment and the C190A in the domain II S4-5 linker from the para-type sodium channel gene. LPT showed that all seven populations were resistant to cypermethrin with resistance ratio (RR) ranging from 16.0 to 25.0 and 85.7% were resistant to chlorpyriphos (RR=2.2-15.6). Although the T2134A mutation was not detected, the C190A mutation was highly prevalent, being present in 82-100% of the alleles sampled in field populations. A significant correlation was found between the LC50 values for cypermethrin and the frequency of the C190A mutation suggesting that it might be responsible for the phenotypic resistance detected.

A VACCINIA VIRUS-DRIVEN INTERPLAY BETWEEN THE MKK4/7-JNK1/2 PATHWAY AND CYTOSKELETON REORGANIZATION

ABSTRACT Viral manipulation of transduction pathways associated with key cellular functions such as survival, response to microbial infection, and cytoskeleton reorganization can provide the supportive milieu for a productive infection. Here, we demonstrate that vaccinia virus (VACV) infection leads to activation of the stress-activated protein kinase (SAPK)/extracellular signal-regulated kinase (ERK) 4/7 (MKK4/7)–c-Jun N-terminal protein kinase 1/2 (JNK1/2) pathway; further, the stimulation of this pathway requires postpenetration, prereplicative events in the viral replication cycle. Although the formation of intracellular mature virus (IMV) was not affected in MKK4/7- or JNK1/2-knockout (KO) cells, we did note an accentuated deregulation of microtubule and actin network organization in infected JNK1/2-KO cells. This was followed by deregulated viral trafficking to the periphery and enhanced enveloped particle release. Furthermore, VACV infection induced alterations in the cell contractility and morphology, and cell migration was reduced in the JNK-KO cells. In addition, phosphorylation of proteins implicated with early cell contractility and cell migration, such as microtubule-associated protein 1B and paxillin, respectively, was not detected in the VACV-infected KO cells. In sum, our findings uncover a regulatory role played by the MKK4/7-JNK1/2 pathway in cytoskeleton reorganization during VACV infection.

Genetic characterization of influenza virus circulating in Brazilian pigs during 2009 and 2010 reveals a high prevalence of the pandemic H1N1 subtype

Influenza A viruses circulating in pigs in Brazil are still not characterized, and only limited data are available about swine influenza epidemiology in the country. Therefore, we characterized the hemagglutinin (HA) and neuraminidase (NA) genes of influenza viruses isolated from Brazilian pigs. We also evaluated one case of probable swine‐to‐human transmission.

Characterization of alpha-enolase as an interferon-alpha 2 alpha 1 regulated gene.

Interferons (IFNs) are multifunctional cytokines that after binding to the cell surface receptor induce the expression of a large number of genes, which in turn, mediate many biological processes including host defense, cell growth control, signaling, and metabolism. Here we show that IFN-alpha activates the mitogen-activated protein kinases (MAPK) ERK1/2 and the transcription factor CREB/ATF-1, which lead to the alpha-enolase (alpha-ENO) gene expression in fibroblasts. Alpha-ENO mRNA accumulation was apparent 6 h post-IFN stimulation and required both de novo protein synthesis and active gene transcription, which is typical of a secondary response gene. Alpha-ENO expression does not appear to be restricted to fibroblasts, since it was equally verified in peripheral blood mononuclear cells (PBMC). Furthermore, IFN-alpha stimulates the expression of the primary response genes c-fos and egr-1, which was followed by an increase in DNA binding activity of c-FOS and EGR-1 proteins, as verified by shift assays using the cis-acting elements AP-1 and EGR-1 localized at the alpha-ENO promoter. Finally, we also demonstrated that IFN treatment of PBMC cause an increase in both, alpha-ENO expression on the cell surface and plasmin generation followed addition of exogenous plasminogen.

Surto de laringotraqueíte infecciosa em granjas de galinhas poedeiras de múltiplas idades em Minas Gerais, Brasil

A recent (November 2010) outbreak of infectious laryngotracheitis (ILT) in a multi-age laying hen facility in Minas Gerais state, Brazil, is described. Previous ILT outbreak in laying hens was only notified in Sao Paulo state, Brazil, in 2002. In the outbreak described here, the affected population was approximately eight million hens, with flock sizes ranging from 100,000 to 2,900,000 chickens. The average mortality ranged from 1 to 6%, and morbidity was around 90% (most of the twenty seven farms of the area were positive for ILT virus). Three multi-age laying farms from one company were selected for this report. Clinical signs included prostration, dyspnea, conjunctivitis, occasional swelling of the paranasal sinuses and bloody mucous nasal discharge. Severely affected chickens presented with dyspnea, gasping and became cyanotic before death. At necropsy, these chickens had fibrinous exudate blocking the larynx and the lumen of cranial part of the trachea. In addition, conjunctivitis with intense hyperemia, edema and sinuses with caseous exudate were present. On histopathology, there were marked necrosis and desquamation of respiratory ephitelium and conjunctiva with numerous syncytial cells formation and fibrinous exudate. Moderate to marked non suppurative (especially lymphocytes and plasma cells) infiltration in the lamina propria also was observed. Sixteen out of 20 examined chickens, eosinophilic intranuclear inclusion bodies were observed in the syncytial cells. The DNA extracted from larynx and trachea produced positive PCR results for ILT virus (ILTV) DNA using formalin-fixed, paraffin embedded (FFPE) samples. Amplicons from a small region of ICP4 gene were submitted to sequencing and showed 100% identity with ILTV EU104910.1 (USA strain), 99% with ILTV JN596963.1 (Australian strain) and 91% with ILTV JN580316.1 (Gallid herpesvirus 1 CEO vaccine strain) and JN580315.1 (Gallid herpesvirus 1 TCO vaccine strain).

Vaccinia virus regulates expression of p21WAF1/Cip1 in A431 cells

In this paper, we provide evidence that both the mRNA and protein levels of the cyclin-dependent kinase (CDK) inhibitor p21WAF1/CDK-interacting protein 1 (Cip1) increase upon infection of A431 cells with Vaccinia virus (VACV). In addition, the VACV growth factor (VGF) seems to be required for the gene expression because infection carried out with the mutant virus VACV-VGF- revealed that this strain was unable to stimulate its transcription. Our findings are also consistent with the notion that the VGF-mediated change in p21WAF1/Cip1 expression is dependent on tyrosine kinase pathway(s) and is partially dependent on mitogen-activated protein kinase/extracellular-signal regulated kinase 1/2. We believe that these pathways are biologically significant because VACV replication and dissemination was drastically affected when the infection was carried out in the presence of the relevant pharmacological inhibitors.

The Host Factor Early Growth Response Gene (EGR-1) Regulates Vaccinia virus Infectivity during Infection of Starved Mouse Cells

Evolution has equipped poxvirus genomes with the coding capacity for several virus-host interaction products which interfere with host cell gene expression and protein function, creating an adequate intracellular environment for a productive infection. We show here that Vaccinia virus (VACV) induces the expression of the cellular transcription factor EGR-1 (early growth response-1) in Mouse Embryonic Fibroblasts (MEFs) through the MEK (mitogen-activated protein kinase (MAPK)/ERK)/ERK (extracellular signal-regulated kinases) pathway, from 3 to 12 h post infection (h.p.i.). By using starved egr-1 knockout (egr-1−/−) MEFs, we demonstrate that VACV replication is reduced by ~1 log in this cell line. Although western blotting and electron microscopy analyses revealed no difference in VACV gene expression or morphogenesis, the specific infectivity of VACV propagated in egr-1−/− MEFs was lower than virus propagated in wild type (WT) cells. This lower infectivity was due to decreased VACV DNA replication during the next cycle of infection. Taken together, these results revealed that EGR-1 appears to facilitate VACV replication in starved fibroblasts by affecting viral particles infectivity.