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1.
Hemoglobin ; 46(1): 15-19, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-35950589

RESUMO

Acknowledging and understanding the extent of thalassemia and hemoglobinopathy issues in a country is crucial for the benefit of implementing a national preventive and control program to reduce its prevalence. In order to obtain reliable prevalence data, the gene frequencies of the thalassemias and other hemoglobinopathies should be investigated. Molecular studies on thalassemia have yet to be done for Brunei's population. It was estimated that carriers of thalassemia or hemoglobinopathies in Brunei is approximately 5.0% or less of the overall population. There are about 200 current cases of thalassemia and other hemoglobinopathies including adults and children reported across all four districts of Brunei. Blood parameter analysis, microscopy, hemoglobin (Hb) electrophoresis and high performance liquid chromatography (HPLC) are the most common methods of investigation in aiding diagnosis in the hospital laboratory. Genotyping analysis conducted in an overseas laboratory has been employed to confirm some diagnosis. Compiled data from 2009-2017 at the Hematology Laboratory of the Raja Isteri Pengiran Anak Saleha Hospital, Jalan Putera Al-Muhtadee Billah, Bandar Seri Begawan, Brunei Darussalam, showed that the most reported diagnoses are α-thalassemia (α-thal) trait, ß-thalassemia (ß-thal) trait, heterozygous Hb E (HBB: c.79G>A)/ß-thal, ß-thal major (ß-TM) and ß-thal intermedia (ß-TI). The data reported indicate the importance of establishing a thalassemia registry with relevant data on patients and patient outcomes as a tool for monitoring and improving patient care.


Assuntos
Hemoglobinopatias , Talassemia alfa , Talassemia beta , Adulto , Brunei , Criança , Hemoglobinopatias/genética , Heterozigoto , Humanos , Talassemia alfa/diagnóstico , Talassemia alfa/epidemiologia , Talassemia alfa/genética , Talassemia beta/genética
2.
J Cell Sci ; 131(18)2018 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-30154213

RESUMO

The cytokine leukaemia inhibitory factor (LIF) promotes self-renewal of mouse embryonic stem cells (ESCs) through activation of the transcription factor Stat3. However, the contribution of other ancillary pathways stimulated by LIF in ESCs, such as the MAPK and PI3K pathways, is less well understood. We show here that naive-type mouse ESCs express high levels of a novel effector of the MAPK and PI3K pathways. This effector is an isoform of the Gab1 (Grb2-associated binder protein 1) adaptor protein that lacks the N-terminal pleckstrin homology (PH) membrane-binding domain. Although not essential for rapid unrestricted growth of ESCs under optimal conditions, the novel Gab1 variant (Gab1ß) is required for LIF-mediated cell survival under conditions of limited nutrient availability. This enhanced survival is absolutely dependent upon a latent palmitoylation site that targets Gab1ß directly to ESC membranes. These results show that constitutive association of Gab1 with membranes through a novel mechanism promotes LIF-dependent survival of murine ESCs in nutrient-poor conditions.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Células-Tronco Embrionárias/metabolismo , Fator Inibidor de Leucemia/metabolismo , Animais , Células Cultivadas , Transdução de Sinais
3.
J Biol Chem ; 292(21): 8694-8704, 2017 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-28373283

RESUMO

The properties and function of large-conductance calcium- and voltage-activated potassium (BK) channels are modified by the tissue-specific expression of regulatory ß1-subunits. Although the short cytosolic N-terminal domain of the ß1-subunit is important for controlling both BK channel trafficking and function, whether the same, or different, regions of the N terminus control these distinct processes remains unknown. Here we demonstrate that the first six N-terminal residues including Lys-3, Lys-4, and Leu-5 are critical for controlling functional regulation, but not trafficking, of BK channels. This membrane-distal region has features of an amphipathic helix that is predicted to control the orientation of the first transmembrane-spanning domain (TM1) of the ß1-subunit. In contrast, a membrane-proximal leucine residue (Leu-17) controls trafficking without affecting functional coupling, an effect that is in part dependent on controlling efficient endoplasmic reticulum exit of the pore-forming α-subunit. Thus cell surface trafficking and functional coupling with BK channels are controlled by distinct domains of the ß1-subunit N terminus.


Assuntos
Retículo Endoplasmático/metabolismo , Regulação da Expressão Gênica/fisiologia , Subunidades alfa do Canal de Potássio Ativado por Cálcio de Condutância Alta/biossíntese , Subunidades beta do Canal de Potássio Ativado por Cálcio de Condutância Alta/biossíntese , Retículo Endoplasmático/genética , Células HEK293 , Humanos , Subunidades alfa do Canal de Potássio Ativado por Cálcio de Condutância Alta/genética , Subunidades beta do Canal de Potássio Ativado por Cálcio de Condutância Alta/genética , Domínios Proteicos , Transporte Proteico/fisiologia
4.
Arch Virol ; 162(8): 2203-2210, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28361286

RESUMO

Porcine reproductive and respiratory syndrome virus (PRRSV) is a major infectious threat to the pig industry worldwide. Increasing evidence suggests that microevolution within a quasispecies population can give rise to high sequence heterogeneity in PRRSV; potentially impacting the pathogenicity of the virus. Here, we report on micro-evolutionary events taking place within the viral quasispecies population in lung and lymph node 3 days post infection (dpi) following experimental in vivo infection with the prototypical Lelystad PRRSV (LV). Sequence analysis revealed 16 high frequency single nucleotide variants (SNV) or differences from the reference LV genome which are assumed to be representative of the consensus inoculum genome. Additionally, 49 other low frequency SNVs were also found in the inoculum population. At 3 dpi, a total of 9 and 10 SNVs of varying frequencies could already be detected in the LV population infecting the lung and lymph nodes, respectively. Interestingly, of these, three and four novel SNVs emerged independently in the two respective tissues when compared to the inoculum. The remaining variants, though already present at lower frequencies in the inoculum, were positively selected and their frequency increased within the quasispecies population. Hence, we were able to determine directly from tissues infected with PRRSV the repertoire of genetic variants within the viral quasispecies population. Our data also suggest that microevolution of these variants is rapid and some may be tissue-specific.


Assuntos
Evolução Molecular , Síndrome Respiratória e Reprodutiva Suína/virologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/genética , Suínos/virologia , Animais , Variação Genética , Genótipo , Pulmão/virologia , Linfonodos/virologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/isolamento & purificação
5.
PLoS Genet ; 9(4): e1003453, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23637623

RESUMO

Following domestication, livestock breeds have experienced intense selection pressures for the development of desirable traits. This has resulted in a large diversity of breeds that display variation in many phenotypic traits, such as coat colour, muscle composition, early maturity, growth rate, body size, reproduction, and behaviour. To better understand the relationship between genomic composition and phenotypic diversity arising from breed development, the genomes of 13 traditional and commercial European pig breeds were scanned for signatures of diversifying selection using the Porcine60K SNP chip, applying a between-population (differentiation) approach. Signatures of diversifying selection between breeds were found in genomic regions associated with traits related to breed standard criteria, such as coat colour and ear morphology. Amino acid differences in the EDNRB gene appear to be associated with one of these signatures, and variation in the KITLG gene may be associated with another. Other selection signals were found in genomic regions including QTLs and genes associated with production traits such as reproduction, growth, and fat deposition. Some selection signatures were associated with regions showing evidence of introgression from Asian breeds. When the European breeds were compared with wild boar, genomic regions with high levels of differentiation harboured genes related to bone formation, growth, and fat deposition.


Assuntos
Cruzamento , Sus scrofa , Animais , Genoma , Genômica , Fenótipo , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , Seleção Genética , Sus scrofa/genética , Suínos
6.
Virol J ; 11: 42, 2014 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-24588855

RESUMO

BACKGROUND: Porcine Reproductive and Respiratory Syndrome (PRRS) is a disease of major economic impact worldwide. The etiologic agent of this disease is the PRRS virus (PRRSV). Increasing evidence suggest that microevolution within a coexisting quasispecies population can give rise to high sequence heterogeneity in PRRSV. FINDINGS: We developed a pipeline based on the ultra-deep next generation sequencing approach to first construct the complete genome of a European PRRSV, strain Olot/9, cultured on macrophages and then capture the rare variants representative of the mixed quasispecies population. Olot/91 differs from the reference Lelystad strain by about 5% and a total of 88 variants, with frequencies as low as 1%, were detected in the mixed population. These variants included 16 non-synonymous variants concentrated in the genes encoding structural and nonstructural proteins; including Glycoprotein 2a and 5. CONCLUSION: Using an ultra-deep sequencing methodology, the complete genome of Olot/91 was constructed without any prior knowledge of the sequence. Rare variants that constitute minor fractions of the heterogeneous PRRSV population could successfully be detected to allow further exploration of microevolutionary events.


Assuntos
Variação Genética , Genoma Viral , Macrófagos/virologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/classificação , Vírus da Síndrome Respiratória e Reprodutiva Suína/genética , RNA Viral/genética , Animais , Análise por Conglomerados , Evolução Molecular , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Dados de Sequência Molecular , Filogenia , Vírus da Síndrome Respiratória e Reprodutiva Suína/crescimento & desenvolvimento , Vírus da Síndrome Respiratória e Reprodutiva Suína/isolamento & purificação , Análise de Sequência de DNA , Suínos , Cultura de Vírus
7.
Syst Rev ; 13(1): 100, 2024 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-38576013

RESUMO

BACKGROUND: Breast cancer incidence has been on the rise significantly in the Asian population, occurring at an earlier age and a later stage. The potential predictive value of molecular subtypes, biomarkers, and genetic variations has not been deeply explored in the Asian population. This study evaluated the effect of molecular subtype classification and the presence or absence of biomarkers and genetic variations on pathological complete response (pCR) after neoadjuvant treatment in Asian breast cancer patients. METHODS: A systematic search was conducted in MEDLINE (PubMed), Science Direct, Scopus, and Cochrane Library databases. Studies were selected if they included Asian breast cancer patients treated with neoadjuvant chemotherapy and contained data for qualitative or quantitative analyses. The quality of the included studies was assessed using the Newcastle Ottawa Scale. Following the random effects model, pooled odds ratios or hazard ratios with 95% confidence intervals for pCR were analysed using Review Manager Software. Heterogeneity between studies was assessed using Cochran's Q-test and I2 test statistics. RESULTS: In total, 19,708 Asian breast cancer patients were pooled from 101 studies. In the neoadjuvant setting, taxane-anthracycline (TA) chemotherapy showed better pCR outcomes in triple-negative breast cancer (TNBC) (p<0.0001) and human epidermal growth factor receptor 2 enriched (HER2E) (p<0.0001) than luminal breast cancer patients. Similarly, taxane-platinum (TP) chemotherapy also showed better pCR outcomes in TNBC (p<0.0001) and HER2E (p<0.0001). Oestrogen receptor (ER)-negative, progesterone receptor (PR)-negative, HER2-positive and high Ki-67 were significantly associated with better pCR outcomes when treated with either TA or TP. Asian breast cancer patients harbouring wildtype PIK3CA were significantly associated with better pCR outcomes when treated with TA in the neoadjuvant setting (p=0.001). CONCLUSIONS: In the neoadjuvant setting, molecular subtypes (HER2E and TNBC), biomarkers (ER, PR, HER2, HR, Ki-67, nm23-H1, CK5/6, and Tau), and gene (PIK3CA) are associated with increased pCR rates in Asian breast cancer patients. Hence, they could be further explored for their possible role in first-line treatment response, which can be utilised to treat breast cancer more efficiently in the Asian population. However, it needs to be further validated with additional powered studies. SYSTEMATIC REVIEW REGISTRATION: PROSPERO CRD42021246295.


Assuntos
Neoplasias da Mama , Hidrocarbonetos Aromáticos com Pontes , Neoplasias de Mama Triplo Negativas , Feminino , Humanos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Classe I de Fosfatidilinositol 3-Quinases , Variação Genética , Antígeno Ki-67/metabolismo , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Receptores de Estrogênio/genética , Receptores de Estrogênio/metabolismo , Taxoides/uso terapêutico , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/genética
8.
Phenomics ; 3(1): 64-76, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36939796

RESUMO

Headache is one of the commonest complaints that doctors need to address in clinical settings. The genetic mechanisms of different types of headache are not well understood while it has been suggested that self-reported headache and self-reported migraine were genetically correlated. In this study, we performed a meta-analysis of genome-wide association studies (GWAS) on the self-reported headache phenotype from the UK Biobank and the self-reported migraine phenotype from the 23andMe using the Unified Score-based Association Test (metaUSAT) software for genetically correlated phenotypes (N = 397,385). We identified 38 loci for headaches, of which 34 loci have been reported before and four loci were newly suggested. The LDL receptor related protein 1 (LRP1)-Signal Transducer and Activator of Transcription 6 (STAT6)-S hort chain D ehydrogenase/R eductase family 9C member 7 (SDR9C7) region in chromosome 12 was the most significantly associated locus with a leading p value of 1.24 × 10-62 of rs11172113. The One Cut homeobox 2 (ONECUT2) gene locus in chromosome 18 was the strongest signal among the four new loci with a p value of 1.29 × 10-9 of rs673939. Our study demonstrated that the genetically correlated phenotypes of self-reported headache and self-reported migraine can be meta-analysed together in theory and in practice to boost study power to identify more variants for headaches. This study has paved way for a large GWAS meta-analysis involving cohorts of different while genetically correlated headache phenotypes. Supplementary Information: The online version contains supplementary material available at 10.1007/s43657-022-00078-7.

9.
Front Biosci (Landmark Ed) ; 27(7): 213, 2022 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-35866409

RESUMO

INTRODUCTION: Colorectal cancer (CRC) is one of the most common cancer types, with rising incidence due to imbalanced lifestyle and dietary habit. Association between CRC cases and KRAS mutation has been established recently. Brunei Darussalam, located within the Borneo island, is of diverse ethnicity which could represent the genome of Southeast Asia population. Our study, for the first time, determined the survival outcome of metastatic colorectal cancer (mCRC) and established the link with KRAS mutation by modelling the population in Brunei Darussalam. METHODS: We collected data of 76 metastatic CRC (mCRC) patients undergoing treatment at The Brunei Cancer Centre, the national centre for cancer treatment in Brunei. These patients were diagnosed with Stage 4 CRC between 1 January 2013 and 31 December 2017. Age, gender, ethnicity, date of diagnosis, site of primary tumour, metastatic sites and molecular analysis of KRAS mutation status (either KRAS mutated or KRAS wild-type) of tumour were recorded. The survival outcomes of these mCRC patients were analysed. RESULTS: The end of this study period recorded 73.1% deceased mutant KRAS mCRC patients and 46.0% deceased wild-type KRAS mCRC patients, contributing to death rates of 45.2% and 54.8%, correspondingly. Chi-squared analysis showed a significant difference between the survival outcomes of wild-type KRAS and mutant KRAS mCRC patients (p-value = 0.024). CONCLUSIONS: There is a significant difference between the survival outcomes of wild-type KRAS and mutant KRAS mCRC patients in the Brunei population. In addition, we found that mutations in codon 12 of KRAS gene on mutant KRAS mCRC patients have shorter survival median periods than those with mutations within codon 13 of KRAS gene. This is the first study in Brunei Darussalam to analyse both the survival outcomes of mCRC patients and those of mutant KRAS mCRC patients.


Assuntos
Neoplasias Colorretais , Proteínas Proto-Oncogênicas p21(ras) , Códon , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Humanos , Mutação , Metástase Neoplásica , Proteínas Proto-Oncogênicas p21(ras)/genética
10.
BMC Genomics ; 12: 110, 2011 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-21324158

RESUMO

BACKGROUND: C57BL/6J mice possess a single intelectin (Itln) gene on chromosome 1. The function of intelectins is not well understood, but roles have been postulated in insulin sensitivity, bacterial recognition, intestinal lactoferrin uptake and response to parasites and allergens. In contrast to C57BL/6J mice, there is evidence for expansion of the Itln locus in other strains and at least one additional mouse Itln gene product has been described. The aim of this study was to sequence and characterise the Itln locus in the 129S7 strain, to determine the nature of the chromosomal expansion and to inform possible future gene deletion strategies. RESULTS: Six 129S7 BAC clones were sequenced and assembled to generate 600 kbp of chromosomal sequence, including the entire Itln locus of approximately 500 kbp. The locus contained six distinct Itln genes, two CD244 genes and several Itln- and CD244-related pseudogenes. It was approximately 433 kbp larger than the corresponding C57BL/6J locus. The expansion of the Itln locus appears to have occurred through multiple duplications of a segment consisting of a full-length Itln gene, a CD244 (pseudo)gene and an Itln pseudogene fragment. Strong evidence for tissue-specific distribution of Itln variants was found, indicating that Itln duplication contributes more than a simple gene dosage effect. CONCLUSIONS: We have characterised the Itln locus in 129S7 mice to reveal six Itln genes with distinct sequence and expression characteristics. Since C57BL/6J mice possess only a single Itln gene, this is likely to contribute to functional differences between C57BL/6J and other mouse strains.


Assuntos
Dosagem de Genes , Loci Gênicos , Lectinas/genética , Animais , Antígenos CD/genética , Sequência de Bases , Sítios de Ligação , Cromossomos Artificiais Bacterianos , Cromossomos de Mamíferos/genética , Evolução Molecular , Biblioteca Gênica , Genômica , Proteínas de Homeodomínio/metabolismo , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Anotação de Sequência Molecular , Dados de Sequência Molecular , Filogenia , Regiões Promotoras Genéticas , Pseudogenes , Receptores Imunológicos/genética , Duplicações Segmentares Genômicas , Análise de Sequência de DNA , Família de Moléculas de Sinalização da Ativação Linfocitária , Fatores de Transcrição/metabolismo
11.
Public Health Genomics ; 22(3-4): 132-139, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31587001

RESUMO

Pharmacogenomics (PGx) is increasingly being recognized as a potential tool for improving the efficacy and safety of drug therapy. Therefore, several efforts have been undertaken globally to facilitate the implementation process of PGx into routine clinical practice. Part of these efforts include the formation of PGx working groups working on PGx research, synthesis, and dissemination of PGx data and creation of PGx implementation strategies. In Asia, the Southeast Asian Pharmacogenomics Research Network (SEAPharm) is established to enable and strengthen PGx research among the various PGx communities within but not limited to countries in SEA; with the ultimate goal to support PGx implementation in the region. From the perspective of SEAPharm member countries, there are several key elements essential for PGx implementation at the national level. They include pharmacovigilance database, PGx research, health economics research, dedicated laboratory to support PGx testing for both research and clinical use, structured PGx education, and supportive national health policy. The status of these essential elements is presented here to provide a broad picture of the readiness for PGx implementation among the SEAPharm member countries, and to strengthen the PGx research network and practice in this region.


Assuntos
Relações Interprofissionais , Farmacogenética/estatística & dados numéricos , Ásia , Sudeste Asiático , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Difusão de Inovações , Toxidermias/prevenção & controle , Humanos , Farmacogenética/economia
12.
Cell Stress Chaperones ; 13(3): 375-85, 2008 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-18465210

RESUMO

Squamous epithelium in mammals has evolved an atypical stress response involving down-regulation of the classic HSP70 protein and induction of sets of proteins including one named SEP53. This atypical stress response might be due to the unusual environmental pressures placed on squamous tissue. In fact, SEP53 plays a role as an anti-apoptotic factor in response to DNA damage induced by deoxycholic acid stresses implicated in oesophageal reflux disease. SEP53 also has a genetic signature characteristic of an adaptively and rapidly evolving gene, and this observation has been used to imply a role for SEP53 in immunity. Physiological models of squamous tissue are required to further define the regulation and function of SEP53. We examined whether porcine squamous epithelium would be a good model to study SEP53, since this animal suffers from a bile-reflux disease in squamous oesophageal tissue. We have (1) cloned and sequenced the porcine SEP53 locus from porcine bacterial artificial chromosome genomic DNA, (2) confirmed the strikingly divergent nature of the C-terminal portion of the SEP53 gene amongst mammals, (3) discovered that a function of the conserved N-terminal domain of the gene is to maintain cytoplasmic localisation, and (4) examined SEP53 expression in normal and diseased porcine pars oesophagea. SEP53 expression in porcine tissue was relatively confined to gastric squamous epithelium, consistent with its expression in normal human squamous epithelium. Immunohistochemical staining for SEP53 protein in normal and damaged pars oesophagea demonstrated significant stabilisation of SEP53 protein in the injured tissue. These results suggest that porcine squamous epithelium would be a robust physiological model to examine the evolution and function of the SEP53 stress pathway in modulating stress-induced responses in squamous tissue.


Assuntos
Bile/metabolismo , Modelos Animais de Doenças , Epitélio/patologia , Esôfago/anatomia & histologia , Esôfago/patologia , Proteínas de Membrana/metabolismo , Proteínas de Neoplasias/metabolismo , Sequência de Aminoácidos , Animais , Linhagem Celular Tumoral , Epitélio/metabolismo , Esôfago/metabolismo , Humanos , Proteínas de Membrana/genética , Dados de Sequência Molecular , Proteínas de Neoplasias/genética , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Suínos
13.
Antiviral Res ; 144: 322-329, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28698014

RESUMO

Although vaccination has been hugely successful in protecting birds against infection by the New castle disease virus (NDV), newly-emerged highly virulent strains have been found to overcome established immune protection and threaten the poultry industry. The need to improve the immunization efficacy is, therefore, urgent. Here, we tested the potential immunostimulatory adjuvant activity of the adenoviral-expressed recombinant chicken granulocyte monocyte colony stimulating factor (rchGM-CSF) in an inactivated Newcastle Disease Virus (NDV) vaccine. 126 commercial layer chicks, divided into six groups, were first vaccinated at day 7, followed by a subsequent boost and later an intramuscular challenge at day 21 and 35 respectively. rchGM-CSF expressed by adenovirus raised NDV-specific hemagglutinin-inhibition (HI) titers from 10 to 12 (log2) and significantly upregulated the production of interferon α/ß/γ (IFN-α/ß/γ), interleukin-4 (IL-4) and major histocompatibility complex II (MHC-II) in spleens. Crucially, chicks inoculated with the inactivated NDV vaccine plus the rchGM-CSF adjuvant displayed only mild clinical signs, lower tissue viral loads, fewer tissue lesions, and decreased mortality and viral shedding than those in the group immunized with the vaccine alone. Our present work has demonstrated that chicken GM-CSF may act as an enhancer in the orchestration of host immune responses induced by the inactivated NDV vaccine. The molecule, expressed by an adenovirus, has the potential to be used as an immune adjuvant to improve protection by NDV vaccination.


Assuntos
Adjuvantes Imunológicos/administração & dosagem , Fator Estimulador de Colônias de Granulócitos e Macrófagos/administração & dosagem , Doença de Newcastle/prevenção & controle , Vírus da Doença de Newcastle/imunologia , Vacinas Virais/imunologia , Adenoviridae/genética , Adjuvantes Imunológicos/genética , Animais , Anticorpos Antivirais/sangue , Galinhas , Portadores de Fármacos , Testes de Inibição da Hemaglutinação , Antígenos de Histocompatibilidade Classe II/análise , Interferons/metabolismo , Interleucina-4/metabolismo , Leucócitos Mononucleares/imunologia , Doença de Newcastle/patologia , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/genética , Baço/imunologia , Análise de Sobrevida , Vacinas de Produtos Inativados/administração & dosagem , Vacinas de Produtos Inativados/imunologia , Vacinas Virais/administração & dosagem
14.
Genome Announc ; 3(3)2015 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-25999564

RESUMO

We report here the complete genome of the pathogenic eastern European subtype 3 porcine reproductive and respiratory syndrome virus (PRRSV) strain SU1-Bel, sequenced directly from a pig lymph node. While sharing substantial sequence similarity with other subtype 3 strains, SU1-Bel is found to harbor unique indels and contain putative novel subgenomic RNAs.

15.
J Vet Diagn Invest ; 15(6): 585-8, 2003 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-14667025

RESUMO

Polymerase chain reaction (PCR) from paraffin-embedded tissues provides a powerful tool to amplify DNA from a variety of recent and archival material. Because DNA from paraffin-embedded samples is more degraded than from fresh material, the amplification of reference genes is essential to exclude false-negative results. This study describes the use of the proliferative cell nuclear antigen (PCNA) gene as a reference gene in a range of animal species and in humans. The PCNA-PCR to amplify a fragment extending from exon 5 through exon 6 and including the intervening intron 6 gave a reproducible pattern, with a 280-base pair (bp) band from canine, equine, bovine, ovine, and caprine samples showing high sequence homology. Porcine, guinea pig, tiger, and lion samples, however, gave an additional fragment of approximately 197 bp. The whole intron 6 from these fragments is missing, possibly representing a pseudogene. In feline samples only the 197-bp fragment could be detected. This study shows that the PCNA gene is highly conserved across a broad range of animal species and is well suited as an internal control for PCR analysis in veterinary medicine.


Assuntos
Mamíferos/genética , Reação em Cadeia da Polimerase/veterinária , Antígeno Nuclear de Célula em Proliferação/genética , Animais , Sequência de Bases , Reações Falso-Negativas , Dados de Sequência Molecular , Técnicas de Amplificação de Ácido Nucleico , Parafina , Valores de Referência , Reprodutibilidade dos Testes , Manejo de Espécimes , Fixação de Tecidos
16.
Am J Vet Res ; 64(11): 1421-8, 2003 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-14620780

RESUMO

OBJECTIVE: To detect and characterize the full range of chlamydial infections in cats with ocular disease by use of polymerase chain reaction (PCR) assays, cytologic examination, immunohistochemical analysis, and evaluation of clinical information including status for feline herpesvirus-1 (FeHV-1). SAMPLE POPULATION: DNA extracted from 226 conjunctival samples obtained from cats with clinically diagnosed keratitis or conjunctivitis and 30 conjunctival samples from healthy cats. PROCEDURE: PCR assays for the 16S rRNA gene specific for the order Chlamydiales and a new Chlamydophila felis (formerly Chlamydia psittaci) species-specific 23S rRNA gene were performed. Seventy-four conjunctival samples were prepared with Romanowsky-type stain, grouped on the basis of inflammatory pattern, and screened for chlamydial inclusions by use of immunohistochemical analysis. Clinical information and FeHV-1 status were recorded. RESULTS: 26 (12%) specimens had positive results for the only known feline chlamydial pathogen, C felis. Surprisingly, an additional 88 (39%) were positive for non-C felis chlamydial DNA. Identification of non-C felis chlamydial DNA by direct sequencing revealed 16S rRNA gene sequences that were 99% homologous to the sequence for Neochlamydia hartmannellae, an amebic endosymbiont. Chlamydial prevalence was significantly higher in cats with ocular disease. CONCLUSIONS AND CLINICAL RELEVANCE: Application of a broad-range detection method resulted in identification of a new agent associated with ocular disease in cats. Finding chlamydia-like agents such as N hartmannellae in coinfections with their obligate amebic host, Hartmannella vermiformis, raises questions about the potential role of these microorganisms in causation or exacerbation of ocular disease in cats.


Assuntos
Doenças do Gato/diagnóstico , Infecções por Chlamydia/veterinária , Chlamydia/isolamento & purificação , Conjuntivite de Inclusão/veterinária , Animais , Sequência de Bases , Doenças do Gato/microbiologia , Doenças do Gato/patologia , Gatos , Chlamydia/classificação , Chlamydia/genética , Infecções por Chlamydia/diagnóstico , Infecções por Chlamydia/patologia , Conjuntivite de Inclusão/diagnóstico , Conjuntivite de Inclusão/patologia , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , RNA Bacteriano/genética , RNA Bacteriano/isolamento & purificação , RNA Ribossômico 16S/genética , RNA Ribossômico 16S/isolamento & purificação , RNA Ribossômico 23S/genética , RNA Ribossômico 23S/isolamento & purificação , Alinhamento de Sequência , Homologia de Sequência do Ácido Nucleico
17.
Virus Res ; 194: 167-74, 2014 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-25312450

RESUMO

The highly heterogeneous porcine reproductive and respiratory syndrome virus (PRRSV) is the causative agent responsible for an economically important pig disease with the characteristic symptoms of reproductive losses in breeding sows and respiratory illnesses in young piglets. The virus can be broadly divided into the European and North American-like genotype 1 and 2 respectively. In addition to this intra-strains variability, the impact of coexisting viral quasispecies on disease development has recently gained much attention; owing very much to the advent of the next-generation sequencing (NGS) technologies. Genomic data produced from the massive sequencing capacities of NGS have enabled the study of PRRSV at an unprecedented rate and details. Unlike conventional sequencing methods which require knowledge of conserved regions, NGS allows de novo assembly of the full viral genomes. Evolutionary variations gained from different genotypic strains provide valuable insights into functionally important regions of the virus. Together with the advancement of sophisticated bioinformatics tools, ultra-deep NGS technologies make the detection of low frequency co-evolving quasispecies possible. This short review gives an overview, including a proposed workflow, on the use of NGS to explore the genetic diversity of PRRSV at both macro- and micro-evolutionary levels.


Assuntos
Variação Genética , Genoma Viral , Sequenciamento de Nucleotídeos em Larga Escala , Filogenia , Vírus da Síndrome Respiratória e Reprodutiva Suína/classificação , Vírus da Síndrome Respiratória e Reprodutiva Suína/genética , Animais , Evolução Molecular , Síndrome Respiratória e Reprodutiva Suína/virologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/isolamento & purificação , Suínos
18.
PLoS One ; 8(8): e71363, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23977029

RESUMO

Large animal models are an important resource for the understanding of human disease and for evaluating the applicability of new therapies to human patients. For many diseases, such as cone dystrophy, research effort is hampered by the lack of such models. Lentiviral transgenesis is a methodology broadly applicable to animals from many different species. When conjugated to the expression of a dominant mutant protein, this technology offers an attractive approach to generate new large animal models in a heterogeneous background. We adopted this strategy to mimic the phenotype diversity encounter in humans and generate a cohort of pigs for cone dystrophy by expressing a dominant mutant allele of the guanylate cyclase 2D (GUCY2D) gene. Sixty percent of the piglets were transgenic, with mutant GUCY2D mRNA detected in the retina of all animals tested. Functional impairment of vision was observed among the transgenic pigs at 3 months of age, with a follow-up at 1 year indicating a subsequent slower progression of phenotype. Abnormal retina morphology, notably among the cone photoreceptor cell population, was observed exclusively amongst the transgenic animals. Of particular note, these transgenic animals were characterized by a range in the severity of the phenotype, reflecting the human clinical situation. We demonstrate that a transgenic approach using lentiviral vectors offers a powerful tool for large animal model development. Not only is the efficiency of transgenesis higher than conventional transgenic methodology but this technique also produces a heterogeneous cohort of transgenic animals that mimics the genetic variation encountered in human patients.


Assuntos
Animais Geneticamente Modificados , Heterogeneidade Genética , Guanilato Ciclase/genética , Células Fotorreceptoras Retinianas Cones/patologia , Distrofias Retinianas/genética , Transgenes , Sequência de Aminoácidos , Animais , Modelos Animais de Doenças , Eletrorretinografia , Genes Dominantes , Vetores Genéticos , Guanilato Ciclase/metabolismo , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Lentivirus/genética , Dados de Sequência Molecular , Mutação , Fenótipo , Células Fotorreceptoras Retinianas Cones/enzimologia , Distrofias Retinianas/patologia , Homologia de Sequência de Aminoácidos , Índice de Gravidade de Doença , Suínos/genética , Acuidade Visual
20.
FEMS Microbiol Lett ; 302(1): 69-75, 2010 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19903200

RESUMO

Intracellular copper routing in Enterococcus hirae is accomplished by the CopZ copper chaperone. Under copper stress, CopZ donates Cu(+) to the CopY repressor, thereby releasing its bound zinc and abolishing repressor-DNA interaction. This in turn induces the expression of the cop operon, which encodes CopY and CopZ, in addition to two copper ATPases, CopA and CopB. To gain further insight into the function of CopZ, the yeast two-hybrid system was used to screen for proteins interacting with the copper chaperone. This led to the identification of Gls24, a member of a family of stress response proteins. Gls24 is part of an operon containing eight genes. The operon was induced by a range of stress conditions, but most notably by copper. Gls24 was overexpressed and purified, and was shown by surface plasmon resonance analysis to also interact with CopZ in vitro. Circular dichroism measurements revealed that Gls24 is partially unstructured. The current findings establish a novel link between Gls24 and copper homeostasis.


Assuntos
Proteínas de Bactérias/metabolismo , Cobre/metabolismo , Enterococcus/metabolismo , Proteínas de Choque Térmico/biossíntese , Chaperonas Moleculares/metabolismo , Transativadores/metabolismo , Ativação Transcricional , Adaptação Fisiológica , Enterococcus/genética , Regulação Bacteriana da Expressão Gênica , Proteínas de Choque Térmico/genética , Homeostase , Dados de Sequência Molecular , Óperon , Estresse Fisiológico , Técnicas do Sistema de Duplo-Híbrido
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