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1.
Rev Bras Parasitol Vet ; 18 Suppl 1: 1-6, 2009 Dec.
Artículo en Portugués | MEDLINE | ID: mdl-20040183

RESUMEN

The aim of this study was to determine whether Southern Blot/Hybridization (SB) associated to Polymerase Chain Reaction (PCR) improves the sensitivity in the detection of hemoplasma DNA in domestic cats (Felis catus). Whole blood was collected in tubes containing the anticoagulant ethylenediamine tetra-acetic acid and DNA extracted from 149 animals. PCR was performed using species specific primers to amplify the 16S ribosomal RNA subunit of Mycoplasma haemofelis and 'Candidatus M. haemominutum' from these samples. Hybridization was performed using a 16S rDNA probes chemically labeled and the results were visualized using a chemiluminescent substrate addition followed by autoradiography. Eighteen (12.1%) of the 149 tested samples had a positive PCR result for hemoplasma species DNA. SB/hybridization technique showed that 24/149 (16.1%) samples were positive for hemoplasmas, confirming the 18 PCR-positive results and reveling six additional positive animals (p<0.001). SB/hybridization method with specific probes was more sensitive than PCR performed alone, being complimentary to this technique to diagnose infections caused by feline hemoplasmas.


Asunto(s)
Animales Domésticos , Southern Blotting , Enfermedades de los Gatos/diagnóstico , Infecciones por Mycoplasma/veterinaria , Reacción en Cadena de la Polimerasa , Animales , Gatos , Infecciones por Mycoplasma/diagnóstico , Sensibilidad y Especificidad
2.
Rev. bras. parasitol. vet ; 18(supl.1): 1-6, out.-dez. 2009. ilus
Artículo en Portugués | LILACS | ID: lil-624821

RESUMEN

O objetivo deste trabalho foi verificar se a técnica de Southern Blot/Hibridização (SB) em associação à reação de polimerização em cadeia (PCR) aumenta a sensibilidade na detecção de DNA de hemoplasmas em gatos domésticos (Felis catus). O sangue total foi coletado em tubos contendo o anticoagulante ácido etilenodiamino tetra-acético, o DNA extraído a partir de 149 animais e a PCR realizada com o uso de sequências iniciadoras espécie-específicas, para amplificar subunidade 16S do RNA ribossomal de Mycoplasma haemofelis e 'Candidatus M. haemominutum' dessas amostras. Para a hibridização, foram utilizadas sondas específicas quimicamente marcadas, e os resultados visualizados por meio da adição de substrato quimiluminescente seguida de autoradiografia. Dezoito (12,1%) das 149 amostras testadas apresentaram resultado PCR-positivo para o DNA de hemoplasmas. A técnica de SB mostrou que 24/149 (16,1%) amostras apresentaram resultado positivo para hemoplasmas, confirmando os 18 resultados PCR-positivos, além de revelar seis outros adicionais (p < 0,001). O método de SB com sondas específicas mostrou-se mais sensível do que a PCR realizada isoladamente, sendo complementar para o diagnóstico das infecções causadas pelos hemoplasmas felinos.


The aim of this study was to determine whether Southern Blot/Hybridization (SB) associated to Polymerase Chain Reaction (PCR) improves the sensitivity in the detection of hemoplasma DNA in domestic cats (Felis catus). Whole blood was collected in tubes containing the anticoagulant ethylenediamine tetra-acetic acid and DNA extracted from 149 animals. PCR was performed using species specific primers to amplify the 16S ribosomal RNA subunit of Mycoplasma haemofelis and 'Candidatus M. haemominutum' from these samples. Hybridization was performed using a 16S rDNA probes chemically labeled and the results were visualized using a chemiluminescent substrate addition followed by autoradiography. Eighteen (12.1%) of the 149 tested samples had a positive PCR result for hemoplasma species DNA. SB/hybridization technique showed that 24/149 (16.1%) samples were positive for hemoplasmas, confirming the 18 PCR-positive results and reveling six additional positive animals (p < 0.001). SB/hybridization method with specific probes was more sensitive than PCR performed alone, being complimentary to this technique to diagnose infections caused by feline hemoplasmas.


Asunto(s)
Animales , Gatos , Animales Domésticos , Southern Blotting , Enfermedades de los Gatos/diagnóstico , Infecciones por Mycoplasma/veterinaria , Reacción en Cadena de la Polimerasa , Infecciones por Mycoplasma/diagnóstico , Sensibilidad y Especificidad
3.
Rev. Bras. Parasitol. Vet. (Online) ; 18(supl. 1): 1-6, 2009. ilus
Artículo en Portugués | LILACS, VETINDEX | ID: biblio-1487757

RESUMEN

O objetivo deste trabalho foi veriicar se a técnica de Southern Blot/Hibridização (SB) em associação à reação de polimerização em cadeia (PCR) aumenta a sensibilidade na detecção de DNA de hemoplasmas em gatos domésticos (Felis catus). O sangue total foi coletado em tubos contendo o anticoagulante ácido etilenodiamino tetra-acético, o DNA extraído a partir de 149 animais e a PCR realizada com o uso de sequências iniciadoras espécie-especíicas, para ampliicar subunidade 16S do RNA ribossomal de Mycoplasma haemofelis e ‘Candidatus M. haemominutum’ dessas amostras. Para a hibridização, foram utilizadas sondas especíicas quimicamente marcadas, e os resultados visualizados por meio da adição de substrato quimiluminescente seguida de autoradiograia. Dezoito (12,1%) das 149 amostras testadas apresentaram resultado PCR-positivo para o DNA de hemoplasmas. A técnica de SB mostrou que 24/149 (16,1%) amostras apresentaram resultado positivo para hemoplasmas, conirmando os 18 resultados PCR-positivos, além de revelar seis outros adicionais (p < 0,001). O método de SB com sondas especíicas mostrou-se mais sensível do que a PCR realizada isoladamente, sendo complementar para o diagnóstico das infecções causadas pelos hemoplasmas felinos.


The aim of this study was to determine whether Southern Blot/Hybridization (SB) associated to Polymerase Chain Reaction (PCR) improves the sensitivity in the detection of hemoplasma DNA in domestic cats (Felis catus). Whole blood was collected in tubes containing the anticoagulant ethylenediamine tetra-acetic acid and DNA extracted from 149 animals. PCR was performed using species speciic primers to amplify the 16S ribosomal RNA subunit of Mycoplasma haemofelis and ‘Candidatus M. haemominutum’ from these samples. Hybridization was performed using a 16S rDNA probes chemically labeled and the results were visualized using a chemiluminescent substrate addition followed by autoradiography. Eighteen (12.1%) of the 149 tested samples had a positive PCR result for hemoplasma species DNA. SB/hybridization technique showed that 24/149 (16.1%) samples were positive for hemoplasmas, conirming the 18 PCR-positive results and reveling six additional positive animals (p < 0.001). SB/hybridization method with speciic probes was more sensitive than PCR performed alone, being complimentary to this technique to diagnose infections caused by feline hemoplasmas.


Asunto(s)
Animales , Gatos , Reacción en Cadena de la Polimerasa , Southern Blotting/métodos
4.
J Feline Med Surg ; 10(2): 120-9, 2008 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-17905624

RESUMEN

The aim of this study was to determine the prevalence and risk factors for Mycoplasma haemofelis (Mhf) and 'Candidatus Mycoplasma haemominutum' (Mhm) infections in domestic cats tested for feline immunodeficiency virus (FIV) and feline leukemia virus (FeLV) with a commercial enzyme-linked immunosorbent assay (ELISA) kit. Based on serological testing, cats were grouped as i) FIV-positive (n=25); ii) FeLV-positive (n=39); iii) FIV/FeLV-positive (n=8); and iv) FIV/FeLV-negative (n=77). Complete blood counts were followed by DNA extraction, species-specific polymerase chain reaction (16S rRNA gene) for Mhf and Mhm and Southern blotting for all animals. Mhf DNA was found in 4.0, 2.6, 12.5 and 7.8% of the cats from groups i, ii, iii and iv, respectively, while 32, 5.1, 50 and 5.2% of these animals had an Mhm infection. Cats with FIV (OR=4.25, P=0.009) and both FIV and FeLV (OR=7.56, P=0.014) were at greater risk of being hemoplasma infected than retroviral-negative cats, mainly due to Mhm infection (OR=8.59, P=0.001 and OR=18.25, P=0.001, respectively). Among pure-breed cats, FIV-positive status was associated with hemoplasma infection (OR 45.0, P=0.001).


Asunto(s)
Enfermedades de los Gatos/epidemiología , Síndrome de Inmunodeficiencia Adquirida del Felino/epidemiología , Leucemia Felina/epidemiología , Infecciones por Mycoplasma/veterinaria , Animales , Recuento de Células Sanguíneas/veterinaria , Southern Blotting/veterinaria , Brasil/epidemiología , Gatos , Comorbilidad , ADN Bacteriano/química , ADN Bacteriano/genética , Ensayo de Inmunoadsorción Enzimática/métodos , Ensayo de Inmunoadsorción Enzimática/veterinaria , Femenino , Masculino , Mycoplasma/aislamiento & purificación , Infecciones por Mycoplasma/epidemiología , Reacción en Cadena de la Polimerasa/veterinaria , Prevalencia , Estudios Prospectivos , Factores de Riesgo
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