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Diagnóstico Laboratorial De Dirofilariosis

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Diagnóstico Laboratorial De Dirofilariosis DIAGNÓSTICO LABORATORIAL DE DIROFILARIOSIS Elena Carretón, José Alberto Montoya-Alonso Medicina interna, Facultad de Veterinaria, Universidad de las Palmas de Gran Canaria DIROFILARIOSIS Dirofilaria immitis Perros, gatos y hurones DIROFILARIOSIS CANINA Arterias pulmonares Ventrículo derecho Wolbachia pipientis DIROFILARIOSIS CANINA Enfermedad cosmopolita Distribución mundial ↑Temperatura y humedad DIROFILARIOSIS CANINA Transmisión anual Transmisión estacional DIROFILARIOSIS CANINA España: condiciones óptimas Prevalencia que presente cada zona condiciones medioambientales Montoya et al, 2007 hábitos de la población canina DIROFILARIOSIS CANINA Prevalencias más elevadas: Islas Canarias Gran Canaria: 67% - 19% Prevalencia 80 60 40 20 0 1994 1995 1996 2000 2002 2007 2010 1. TESTS DE ANTÍGENOS Técnicas de ELISA o inmunocromatografía Proteínas parásitos adultos hembras 1. TESTS DE ANTÍGENOS Elevada ESPECIFICIDAD Falsos Positivos Elevada SENSIBILIDAD Falsos Negativos Snap 3Dx®, Snap Filaria® (Idexx) idexx Sensibilidad 98.7 % Especificidad 100 % Speed Diro® (Virbac) Sensibilidad 95.2 % Especificidad 99 % Anigen HW Ag 2.0 ® (Bionote) anigen Sensibilidad 94.4 % Especificidad 100 % CHW Ag ® (Quicking Biotech) quicking Sensibilidad 91.3 % Especificidad 100 % Uranotest Dirofilaria® (Uranovet) Sensibilidad 94.4 % Especificidad 100 % Witness ® (Synbiotics - Pfizer) Sensibilidad 97.5 % Especificidad 100 % FasTest HW Antigen® (Megacor) Sensibilidad 98.6 % Especificidad 99.1 % DFV test dirofilaria® (Divasa) Sensibilidad 94.4% Especificidad 100 % SensPERT CHW ® (SensPert) Sensibilidad ??% Especificidad ?? % Tests de Antígenos de pocillos Petchek Filaria PF (Idexx) Filarchek (Ingenasa) Dirochek (Pfizer) 1. TESTS DE ANTÍGENOS Una sola hembra adulta Factores pueden afectar la sensibilidad edad del parásito tamaño del perro 1. TESTS DE ANTÍGENOS No carga antigénica suficiente < 6 meses 1. TESTS DE ANTÍGENOS Cantidad de antígenos: nº hembras adultas?? ↑muerte reciente ↓hembras jóvenes ↓filarias machos 1. TESTS DE ANTÍGENOS Falsos positivos: errores técnicos retrasar la lectura del resultado 1. TESTS DE ANTÍGENOS Falsos negativos: Fallos técnicos Baja sensibilidad Carga parasitaria muy ligera Sólo por filarias macho Dirofilarias <6 meses 1. TESTS DE ANTÍGENOS Si se duda (positivo muy débil) repetir la prueba repetir en otra marca 2. DETECCIÓN DE MICROFILARIAS OBSERVACIÓN DIRECTA objetivo de x10 aumentos 2. DETECCIÓN DE MICROFILARIAS El nº no guarda relación con el nº adultos ni gravedad No identifica tipo de microfilarias Dirofilaria immitis ● Arterias pulmonares ● Ventrículo derecho Dirofilaria repens ● Tejido subcutáneo Acanthocheilonema (Dipetalonema) ● Peritoneo y cavidad reconditum abdominal Acanthocheilonema (Dipetalonema) ● Cavidad peritoneal dracunculoides 2. DETECCIÓN DE MICROFILARIAS Dirofilaria immitis Dipetalonema más móviles reconditum movimiento ondulante y no movimiento rectilíneo y progresivo progresivo 2. DETECCIÓN DE MICROFILARIAS TEST DE WOO Tubo de hematocrito: capa flogística 2. DETECCIÓN DE MICROFILARIAS MÉTODO DE FILTRACIÓN: 1 ml de sangre + 9 ml de carbonato sódico 0.1% Filtro (3 µm diámetro) Azul de metileno 0.1% 2. DETECCIÓN DE MICROFILARIAS TEST DE KNOTT : 1 ml sangre + 9 ml formalina 2% 5’ a 1100 rpm Sedimento + azul de metileno 2. DETECCIÓN DE MICROFILARIAS MÉTODO HISTOQUÍMICO : Extensiones de sangre Se cubren con la fosfatasa ácida 2 horas a TA Contrateñir verde metilo al 1% 2. DETECCIÓN DE MICROFILARIAS D.immitis D.repens D.reconditum D.dracunculoides : ● Poro anal ● Poro anal ● Todo el ● Poro anal parásito ● Poro excretor ● Poro excretor ● Cuerpo interno Canine and feline heartworm disease; diagnosis and therapy. ● Espacio cefálico Schrey & Trautvetter. 1998 2. DETECCIÓN DE MICROFILARIAS 20-30% amicrofilarémicos Localización ectópica de adultos Parásitos de un solo sexo Periodo de prepatencia (7 meses) Respuesta inmune frente MF 2. DETECCIÓN DE MICROFILARIAS Porcentaje se eleva: tratamiento preventivo intermitente MF POSITIVO y TEST AG NEGATIVO Falso Negativo Fallo técnico Bajo nº hembras Anticuerpos que enmascare antígenos MF POSITIVO y TEST AG NEGATIVO Microfilarias no son D.immitis : D.reconditum D.repens D.dracunculoides Estudio histoquímico Estudio morfológico MF POSITIVO y TEST AG NEGATIVO D.immitis y D.repens mayor anchura y longitud D.reconditum y D.dracunculoides más estrechas y cortas D.immitis D.repens D.reconditum D.dracunculoides Longitud µm 260 -340 274 -362 241 -287 190 -211 Anchura µm 4-6.5 5.8-7.3 3.8-5 4.8-5.8 Dirofilaria immitis ● Extremo cefálico afilado ● Cola recta y afilada Dirofilaria repens ● Extremo cefálico romo ● Cola afilada y filiforme (mango de paragüas) Acanthocheilonema (Dipetalonema) ● Extremo cefálico romo con gancho prominente reconditum ● Cola chata y en gancho Acanthocheilonema (Dipetalonema) ● Extremo cefálico romo dracunculoides ● Cola larga y afilada MF POSITIVO y TEST AG NEGATIVO Muerte de adultos: circunstancias naturales tratamiento adulticida MF sobreviven 2 años MF POSITIVO y TEST AG NEGATIVO Transfusión: de un perro microfilarémico MF POSITIVO y TEST AG NEGATIVO Vía transplacentaria: madre con microfilaremia Microfilarias no se desarrollarán en adultos HEMATOLOGÍA Anemia: 10-60% Normocítica normocrómica Hemólisis (síndrome vena cava) Leucocitosis: 20% Neutrofilia, monocitosis Eosinofilia : 85-95% Basofilia: 35-40% HEMATOLOGÍA Perfil de coagulación Trombocitopenia Hipofibrinogenemia Tiempo de protrombinoplastina parcial y tiempo de coagulación activada prolongados Altas concentraciones de PDFs ●Tromboembolismo ●CID BIOQUíMICA Alteración hepática: FA, ALT, AST: 5-10% Alteración renal: Azotemia: <5% Hipoalbúminemia URIANÁLISIS Glomerulonefropatía Depósito de complejos autoinmunes MF circulantes Ag de Wolbachia URIANÁLISIS Proteinuria (U P/C >0.5): 10-45% Group A (WSP POSITIVE) Group B (WSP POSITIVE) 0.72+0.81 0.27+0.33 Group A (WSP POSITIVE) Group B (WSP NEGATIVE) 81.5+78.8 mg/dl 28.1+24.4 mg/dl MARCADORES CARDIOPULMONARES Veterinaria: Enf CP tempranas o asintomáticas Casos dudosos Pronóstico MARCADORES CARDIOPULMONARES Hipertensión pulmonar: Adultos Pérdida de elasticidad ICC derecha MARCADORES CARDIOPULMONARES Tromboembolización: Endoarteritis: oclusión de arteriolas Desprendimiento íntima lesionada Muerte de parásitos MARCADORES CARDIOPULMONARES Valorar las lesiones producidas Grado de severidad Pronóstico Integridad Miocárdica Troponinas T e I: Proteínas musculares estructurales Isoformas cardiacas específicas Necrosis: pérdida de integridad de la membrana Troponinas T e I en dirofilariosis Troponina T: valores normales Troponina I: valores ligeramente aumentados Troponina I es un marcador más sensible que la troponina T Integridad Miocárdica Mioglobina: Proteína de bajo peso molecular Función: fijar el oxígeno en las células musculares Músculo cardíaco y esquelético Mioglobina en dirofilariosis Traumatismo torácico Dilatación vólvulo-torsión Dirofilariosis canina: 18.8% de perros amicrofilarémicos 23.9% de perros microfilarémicos Marcadores de tromboembolismo pulmonar Dímero-D: Evalúa presencia de procesos tromboembólicos Producto final de la degradación de un trombo Dímero-D FIBRINóGENO Elevadas concentraciones: TROMBINA Mayor formación y degradación de fibrina FACTOR XIII FIBRINA PLASMINA DÍMERO-D Dímero-D en dirofilariosis 47% presentan valores patológicos Especialmente perros microfilarémicos incrementa la tendencia al desarrollo de trombos y CID * DIROFILARIOSIS FELINA Los gatos también pueden verse infectados gatos de exterior y de interior Prevalencia: 5-10% de perros ?? SUB -DIAGNOSTICADA!! Gran Canaria: 34 % DIAGNÓSTICO Más complicado Fácil pasarla por alto Bajo número de parásitos Combinar varias técnicas Serología Radiología torácica Ecocardiografía Tests comerciales Antígenos de hembras adultas Resultado negativo: Parásitos machos Parásitos inmaduros o juveniles Antigenemia: > 6-8 meses Detección de anticuerpos Ac filarias desde los 2 meses post-infección Falsos positivos: contacto con el parásito Falsos negativos: 14-20% en gatos infectados Detección de anticuerpos Ray Dillon and Byron Blagburn, 2007 Detección de anticuerpos Anticuerpos anti-WSP Proteína de superficie de Wolbachia (WSP) Microfilaremia Muy poco probable < 20% Radiología torácica Principales hallazgos radiológicos: Arterias lobares y pulmonares: aumento y mayor tortuosidad Proyección DV y LL Radiología torácica Silueta cardíaca no suele modificarse Patrón pulmonar broncointersticial focal o multifocal Suele desaparecer espontáneamente en unos meses La mitad de los gatos presentan alguna alteración radiográfica Ecocardiografía Test antígenos negativo, pero sospechamos DÓNDE BUSCAR: Arterias pulmonares principal y lobar derecha Observar todas las localizaciones posibles Detecta parásitos en un 40- 78% Consultations in Feline Internal Medicine (Fifth Edition) Cuadro diagnóstico NEGATIVO: Bajo índice sospecha ANTICUERPOS POSITIVO: Alto índice sospecha NEGATIVO: Bajo índice sospecha ANTÍGENOS POSITIVO: DIAGNÓSTICO NORMAL: Bajo índice sospecha RADIOGRAFÍA TÓRAX ALTERACIONES: Alto índice sospecha NO PARÁSITOS: No influye índice sospecha ECOCARDIOGRAFÍA SÍ PARÁSITOS: DIAGNÓSTICO [email protected].
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