Papillomaviruses: a systematic review

Abstract In the last decades, a group of viruses has received great attention due to its relationship with cancer development and its wide distribution throughout the vertebrates: the papillomaviruses. In this article, we aim to review some of the most relevant reports concerning the use of bovines as an experimental model for studies related to papillomaviruses. Moreover, the obtained data contributes to the development of strategies against the clinical consequences of bovine papillomaviruses (BPV) that have led to drastic hazards to the herds. To overcome the problem, the vaccines that we have been developing involve recombinant DNA technology, aiming at prophylactic and therapeutic procedures. It is important to point out that these strategies can be used as models for innovative procedures against HPV, as this virus is the main causal agent of cervical cancer, the second most fatal cancer in women.

Papillomaviruses: a systematic review.

In the last decades, a group of viruses has received great attention due to its relationship with cancer development and its wide distribution throughout the vertebrates: the papillomaviruses. In this article, we aim to review some of the most relevant reports concerning the use of bovines as an experimental model for studies related to papillomaviruses. Moreover, the obtained data contributes to the development of strategies against the clinical consequences of bovine papillomaviruses (BPV) that have led to drastic hazards to the herds. To overcome the problem, the vaccines that we have been developing involve recombinant DNA technology, aiming at prophylactic and therapeutic procedures. It is important to point out that these strategies can be used as models for innovative procedures against HPV, as this virus is the main causal agent of cervical cancer, the second most fatal cancer in women.

Integrated analysis of recombinant BPV-1 L1 protein for the production of a bovine papillomavirus VLP vaccine.

Bovine papillomatosis is an infectious disease that is caused by bovine papillomavirus (BPV), which results in important economic losses. However, no BPV vaccines or effective treatment methods are commercially available to date. Moreover, the absence of papillomavirus replication in vitro makes the use of recombinant protein a promising candidate for vaccine formulations. Hence, we developed an integrated study on the L1 capsid protein of BPV-1, obtained from a bacterial expression system, regarding its purification, biosafety, thermostability and immunogenicity. The results indicated an absence of genotoxicity of the purified recombinant L1 protein, ß-sheet prevalence of secondary structure folding, protein stability under high temperatures as well as the presence of capsomeres and VLPs. In addition, preliminary experimental vaccination of calves showed the production of specific antibodies against BPV-1 L1.

Genetics and metabolic deregulation following cancer initiation: A world to explore.

Cancer is a group of highly complex and heterogeneous diseases with several causes. According to the stochastic model, cancer initiates from mutation in somatic cells, leading to genomic instability and cell transformation. This canonical pathway of carcinogenesis is related to the discovery of important mechanisms that regulate cancer initiation. However, there are few studies describing genetic and metabolic alterations that deregulate transformed cells, resulting in epithelial-mesenchymal transition (EMT) and its most dramatic consequence, the metastasis. This review summarizes the main genetics and metabolic changes induced by reactive oxygen species (ROS) that lead to EMT.

Mutagenic Potential ofBos taurus Papillomavirus Type 1 E6 Recombinant Protein: First Description.

Bovine papillomavirus (BPV) is considered a useful model to study HPV oncogenic process. BPV interacts with the host chromatin, resulting in DNA damage, which is attributed to E5, E6, and E7 viral oncoproteins activity. However, the oncogenic mechanisms of BPV E6 oncoprotein per se remain unknown. This study aimed to evaluate the mutagenic potential of Bos taurus papillomavirus type 1 (BPV-1) E6 recombinant oncoprotein by the cytokinesis-block micronucleus assay (CBMNA) and comet assay (CA). Peripheral blood samples of five calves were collected. Samples were subjected to molecular diagnosis, which did not reveal presence of BPV sequences. Samples were treated with 1 µg/mL of BPV-1 E6 oncoprotein and 50 µg/mL of cyclophosphamide (positive control). Negative controls were not submitted to any treatment. The samples were submitted to the CBMNA and CA. The results showed that BPV E6 oncoprotein induces clastogenesis per se, which is indicative of genomic instability. These results allowed better understanding the mechanism of cancer promotion associated with the BPV E6 oncoprotein and revealed that this oncoprotein can induce carcinogenesis per se. E6 recombinant oncoprotein has been suggested as a possible vaccine candidate. Results pointed out that BPV E6 recombinant oncoprotein modifications are required to use it as vaccine.

Morphometric evaluation and nonclassical criteria for the diagnosis of HPV infection and cytological atypia in cervical samples.

Herein, we evaluated cervical samples from normal tissue or HPV-infected tissue, to determine if the relative nuclear/cytoplasmic ratio (NA/CA) and the presence of nonclassical cytological criteria are a novel cytological criterion for the diagnosis of HPV. Significantly, larger NA/CA ratios were found for the HPV-ATYPIA+ and HPV+ATYPIA+ groups compared with HPV-ATYPIA- group, regardless of collection method. For the samples collected with a spatula, only three samples from the HPV-ATIPIA- group showed four or more nonclassical parameters (i.e., were positive), while a larger number of the samples in the HPV-ATYPIA+, HPV+ATYPIA-, and HPV+ATYPIA+ groups were positive (13, 4, and 13 samples, respectively). Among those collected with a brush, no sample showed four or more nonclassical criteria in the HPV-ATYPIA- group, while a number of samples were positive in the HPV-ATYPIA+, HPV+ATYPIA-, and HPV+ATYPIA+ groups (4, 3, and 4 samples, respectively). HPV infection was associated with significant morphometrical changes; no increase in the NA/CA ratio was found in the HPV+ATYPIA- samples, compared with the HPV-ATIPIA- samples collected with either a spatula or a brush. In conclusion, by including nonclassical cytological criteria into the patient diagnosis, we were able to reduce the number of false negative and false positive HPV diagnoses made using conventional cytology alone.

Bovine papillomavirus type 4 L1 gene transfection in a Drosophila S2 cell expression system: absence of L1 protein expression

The development of a bovine papillomavirus (BPV) vaccine is an outstanding challenge. BPV protein L1 gene transfection in the Drosophila melanogaster S2 cell expression system failed to produce L1 protein notwithstanding correct L1 gene insertion. Severe genetic inbalance in the host cell line, including cytogenetic alterations, may account for the lack of protein expression.

O desenvolvimento de uma vacina para papilomavirus bovino (BPV) consiste em grande desafio. A transfecção do gene codificante da proteína L1 de BPV em sistema de células S2 de Drosophila melanogaster não logrou sucesso, apesar da correta inserção da seqüência gênica em vetor apropriado.Graves alterações genéticas na linhagem celular S2, que incluem aberrações cromossômicas, provavelmente estão relacionadas à ausência da expressão da proteína desejada.

Bovine papillomavirus type 4 L1 gene transfection in a Drosophila S2 cell expression system: absence of L1 protein expression.

The development of a bovine papillomavirus (BPV) vaccine is an outstanding challenge. BPV protein L1 gene transfection in the Drosophila melanogaster S2 cell expression system failed to produce L1 protein notwithstanding correct L1 gene insertion. Severe genetic inbalance in the host cell line, including cytogenetic alterations, may account for the lack of protein expression.

PapillomatosIs in cattle: in situ detection of bovine papillomavirus DNA sequences in reproductive tissues

Bovine papillomatosis is a common viral infection in Brazil that is caused by a bovine papillomavirus(BPV). Dissemination is by direct contact between infected animals, although the investigation of othermodes of transmission is a very important aspect in the management of this condition. BPV DNA sequenceshave been detected in many tissues by using the polymerase chain reaction. In this work, we used in situhybridization to detect BPV DNA sequences in bovine reproductive tissues and cells. The detection ofBPV in these tissues strongly suggests that these sequences could be an important alternative of viraltransmission that could contribute to the widespread incidence of bovine papillomatosis and its complexpathology. Alternatively, the viral sequences could result from cell apoptosis and may therefore not bedirectly involved in the infection.

Detection of micronuclei formation and nuclear anomalies in regenerative nodules of human cirrhotic livers and relationship to hepatocellular carcinoma.

Human cirrhosis is considered an important factor in hepatocarcinogenesis. The lack of substantial genetics and cytogenetics data in human cirrhosis led us to investigate spontaneous micronuclei formation, as an indicator of chromosomal damage. The analysis was performed in hepatocytes of regenerative, macroregenerative, and tumoral nodules from 30 cases of cirrhosis (paraffin-embedded archival material), retrospectively selected: cryptogenic, hepatitis C virus, and hepatitis C virus associated with hepatocellular carcinoma (HCC). Thirteen control liver samples of healthy organ donors were included. Micronucleated hepatocytes were analyzed with Feulgen-fast-green dyeing techniques. The spontaneous frequency of micronucleated hepatocytes in both regenerative and macroregenerative nodules of all cirrhotic patients was significantly higher than for the normal control group. There was no significant difference in frequency of micronucleated hepatocytes in regenerative nodules compared with macroregenerative nodules for all cases analyzed, whereas a significantly higher frequency of micronucleated hepatocytes was detected in tumoral nodules, compared with cirrhotic regenerative nodules and normal parenchyma. A higher frequency of the nuclear anomalies termed broken-eggs was observed in hepatitis C virus-related samples. Chromatinic losses and genotoxicity already existed in the cirrhotic regenerative nodules, which might predispose to development of HCC.

Viral DNA sequences in peripheral blood and vertical transmission of the virus: a discussion about BPV-1

Neste estudo detectou-se DNA de BPV-1 em: verruga, sangue e plasma de animais afetados por papilomatose. Estes resultados trazem evidências sobre possível localização intracelular do BPV-1 no sangue. Avaliamos um animal afetado por papilomatose e positivo para BPV-1 em amostras de verruga, sangue, placenta e líquido amniótico e que teve sua cria recém-nascida também positiva para BPV-1 no sangue. Estes resultados indicam possível transmissão vertical do BPV-1.

Bovine papillomavirus type 2 in reproductive tract and gametes of slaughtered bovine females

Os vírus do papiloma bovino, descritos como agentes infectantes específicos do epitélio, têm sido associados a diversas formas de câncer em diferentes espécies animais. Dada a intensa disseminação da papilomatose nos rebanhos, a investigação de diferentes formas de transmissão e seus respectivos mecanismos tem exigido especial atenção. No presente estudo, é relatada a detecção de seqüências genômicas do papilomavirus bovino (BPV) em ovócitos e tecidos do trato reprodutivo oriundos de fêmeas abatidas comercialmente, não apresentando papilomatose cutânea. A presença de DNA de BPV-2 em tecidos do trato reprodutivo, lavado uterino, ovócitos e células do cumulus traz evidências de que a infecção viral pode se desenvolver fora do tecido epitelial. Esses achados alertam para a possibilidade de transmissão do BPV através dos procedimentos de transferência de embriões e de fertilização in vitro.

Somatic pairing, endomitosis and chromosome aberrations in snakes (Viperidae and Colubridae)

The positioning of macrochromosomes of Bothrops jararaca and Bothrops insularis (Viperidae) was studied in undistorted radial metaphases of uncultured cells (spermatogonia and oogonia) not subjected to spindle inhibitors. Colchicinized metaphases from uncultured (spleen and intestine) and cultured tissues (blood) were also analyzed. We report two antagonic non-random chromosome arrangements in untreated premeiotic cells: the parallel configuration with homologue chromosomes associated side by side in the metaphase plate and the antiparallel configuration having homologue chromosomes with antipolar distribution in the metaphase ring. The antiparallel aspect also appeared in colchicinized cells. The spatial chromosome arrangement in both configurations is groupal size-dependent and maintained through meiosis. We also describe, in untreated gonia cells, endomitosis followed by reductional mitosis which restores the diploid number. In B. jararaca males we observed that some gonad regions present changes in the meiotic mechanism. In this case, endoreduplicated cells segregate the diplochromosomes to opposite poles forming directly endoreduplicated second metaphases of meiosis with the suppression of first meiosis. By a successive division, these cells form nuclei with one set of chromosomes. Chromosome doubling in oogonia is known in hybrid species and in parthenogenetic salamanders and lizards. This species also presented chromosome rearrangements leading to aneuploidies in mitosis and meiosis. It is suggested that somatic pairing, endomitosis, meiotic alterations, and chromosomal aberrations can be correlated processes. Similar aspects of nuclei configurations, endomitosis and reductional mitosis were found in other Viperidae and Colubridae species

Viral DNA sequences in peripheral blood and vertical transmission of the virus: a discussion about BPV-1

This study has detected BPV-1 DNA sequences in wart, blood and plasma samples collected from animals affected by papillomatosis, suggesting viral presence inside the cell. We sellected an animal in which we could detect BPV-1 DNA sequences in wart, blood, placenta and amniotic liquid samples and her offspring which presented BPV-1 DNA sequences in blood sample collected immediately after birth. These results show a possible vertical transmission of BPV-1.

Neste estudo detectou-se DNA de BPV-1 em: verruga, sangue e plasma de animais afetados por papilomatose. Estes resultados trazem evidências sobre possível localização intracelular do BPV-1 no sangue. Avaliamos um animal afetado por papilomatose e positivo para BPV-1 em amostras de verruga, sangue, placenta e líquido amniótico e que teve sua cria recém-nascida também positiva para BPV-1 no sangue. Estes resultados indicam possível transmissão vertical do BPV-1.

Bovine papillomavirus type 2 in reproductive tract and gametes of slaughtered bovine females

Papillomaviruses are described selectively infecting epithelial tissues and are associated with many forms of cancer in different species. Considering the widespread dissemination of papillomatosis in livestock, interest is being centred on possible forms of viral transmission and respective mechanisms. In the present study, we report the detection of bovine papillomavirus (BPV) DNA sequences in female reproductive tract tissues, fluids and oocytes from slaughtered bovines not afflicted by cutaneous papillomatosis. BPV-2 DNA sequences were found in ovarian and uterine tissues as well as in oocytes, cumulus cells and uterine flushings. The presence of papillomavirus sequences in reproductive organ tissues and fluids shows that viral infection in organisms can be verified in others tissues, not only in epithelial ones. The present findings alert to the possibility of BPV transmission in embryo transfer programs and assisted fertilization procedures.

Os vírus do papiloma bovino, descritos como agentes infectantes específicos do epitélio, têm sido associados a diversas formas de câncer em diferentes espécies animais. Dada a intensa disseminação da papilomatose nos rebanhos, a investigação de diferentes formas de transmissão e seus respectivos mecanismos tem exigido especial atenção. No presente estudo, é relatada a detecção de seqüências genômicas do papilomavirus bovino (BPV) em ovócitos e tecidos do trato reprodutivo oriundos de fêmeas abatidas comercialmente, não apresentando papilomatose cutânea. A presença de DNA de BPV-2 em tecidos do trato reprodutivo, lavado uterino, ovócitos e células do cumulus traz evidências de que a infecção viral pode se desenvolver fora do tecido epitelial. Esses achados alertam para a possibilidade de transmissão do BPV através dos procedimentos de transferência de embriões e de fertilização in vitro.

Viral DNA sequences in peripheral blood and vertical transmission of the virus: a discussion about BPV-1

This study has detected BPV-1 DNA sequences in wart, blood and plasma samples collected from animals affected by papillomatosis, suggesting viral presence inside the cell. We sellected an animal in which we could detect BPV-1 DNA sequences in wart, blood, placenta and amniotic liquid samples and her offspring which presented BPV-1 DNA sequences in blood sample collected immediately after birth. These results show a possible vertical transmission of BPV-1.

Neste estudo detectou-se DNA de BPV-1 em: verruga, sangue e plasma de animais afetados por papilomatose. Estes resultados trazem evidências sobre possível localização intracelular do BPV-1 no sangue. Avaliamos um animal afetado por papilomatose e positivo para BPV-1 em amostras de verruga, sangue, placenta e líquido amniótico e que teve sua cria recém-nascida também positiva para BPV-1 no sangue. Estes resultados indicam possível transmissão vertical do BPV-1.

Bovine papillomavirus type 2 in reproductive tract and gametes of slaughtered bovine females

Papillomaviruses are described selectively infecting epithelial tissues and are associated with many forms of cancer in different species. Considering the widespread dissemination of papillomatosis in livestock, interest is being centred on possible forms of viral transmission and respective mechanisms. In the present study, we report the detection of bovine papillomavirus (BPV) DNA sequences in female reproductive tract tissues, fluids and oocytes from slaughtered bovines not afflicted by cutaneous papillomatosis. BPV-2 DNA sequences were found in ovarian and uterine tissues as well as in oocytes, cumulus cells and uterine flushings. The presence of papillomavirus sequences in reproductive organ tissues and fluids shows that viral infection in organisms can be verified in others tissues, not only in epithelial ones. The present findings alert to the possibility of BPV transmission in embryo transfer programs and assisted fertilization procedures.

Os vírus do papiloma bovino, descritos como agentes infectantes específicos do epitélio, têm sido associados a diversas formas de câncer em diferentes espécies animais. Dada a intensa disseminação da papilomatose nos rebanhos, a investigação de diferentes formas de transmissão e seus respectivos mecanismos tem exigido especial atenção. No presente estudo, é relatada a detecção de seqüências genômicas do papilomavirus bovino (BPV) em ovócitos e tecidos do trato reprodutivo oriundos de fêmeas abatidas comercialmente, não apresentando papilomatose cutânea. A presença de DNA de BPV-2 em tecidos do trato reprodutivo, lavado uterino, ovócitos e células do cumulus traz evidências de que a infecção viral pode se desenvolver fora do tecido epitelial. Esses achados alertam para a possibilidade de transmissão do BPV através dos procedimentos de transferência de embriões e de fertilização in vitro.

Comportamento sexual e etiologia da sindrome de Down / Sexual behavior and the etiology of Down syndrome

Os habitos sexuais de 123 maes, na epoca da concepcao, foram revelados atraves da Tecnica de Entrevista de Ajuda e analisados estatisticamente. As maes de criancas com "Sindrome de Down" (SD) relataram com mais frequencia longos intervalos de abstinencia, na epoca da concepcao. Independente da idade materna, ha uma associacao entre a incidencia de SD e intervalos longos de obstinencia sexual

Financiamento de pesquisa / Research financing

Mesa redonda onde se discute a situaçäo atual, as preocupaçöes, os problemas e as sugestöes visando resolver a questäo do financiamento da pesquisa. Participaram representantes da FINEP, FAPESP, DCTEC e OPAS (AMSB)

Genetic characterization of lactose fermentation and antibiotic resistance in plasmid-harboring enteric bacteria isolated from amazon amphibians

Bacterias aeróbicas gram-negativas foram isoladas de fezes de 20 anfíbios. Os animais foram capturados na regiäo que seria inundada para a formaçäo da represa da Usina Hidrelétrica de Tucuruí-PA. Entre as 420 colônias bacterianas analisadas, 123 (29%) eram portadoras de plasmídios, sendo: 89% capazes de fermentar a lactose, 41% resistentes à ampicilina, 6,5% à estreptomicina e 6,5% à tetraciclina, em níveis * 10 ug/ml. O gênero predominante entre as 123 amostras foi o Citrobacter (67%), seguido de Enterobacter (16%), Klebsiella (6%), Providencia (3%), Proteus (2%), Aeromonas (2%) e Salmonella (1%). Como grande parte dos plasmídios detectados tinha pesos moleculares suficientes para conter genes capazes de promover sua auto-transferência, a correlaçäo entre a presença dos mesmos e algumas características observadas foi investigada por conjugaçäo. A presença destes genes foi também analisada por segregaçäo natural e induzida, e por hibridizaçäo DNA-DNA, através de sondas radioativas. Os resultados näo demonstraram a presença de plasmídios R, Hly, ou Lac nestas amostras, mas foram encontradas evidências de que os caracteres de fermentaçäo de lactose e produçäo de hemolisinas säo medidos por genes presentes nos cromossomos destas bactérias