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Citrus tristeza virus (CTV) is widespread and causes one of the most economical important viral diseases of citrus. The large scale analysis of citrus samples for CTV detection is conducted routinely by the enzyme-linked immunosorbent assay (ELISA) or its variant named direct tissue blot immunoassay (= immunoprinting). In this study, we evaluated the use of polyclonal antibodies developed to recombinant p25 CTV coat protein for virus detection under an immunoprinting system by using healthy and field CTV infected citrus samples. The combination of goat (CB) anti- CTV/IgG anti-goat-AP conjugate provided a positive reaction with CTV infected plants, and negative with healthy citrus. In addition, the combination of goat CB anti- CTV coupled with biotin, plus avidin-AP as commercial conjugate was equally effective in discriminating CTV infected from healthy citrus. Furthermore, antibodies developed to the recombinant p25 CTV coat protein, either untagged or biotin conjugated showed comparable results with commercially available immunoprinting kits for CTV detection by Plant Print and Agdia. [ABSTRACT FROM AUTHOR]

Four commercial ELISA kits for serological detection of Citrus tristeza virus (CTV) were evaluated under uniform laboratory conditions. All four kits were efficient for discrimination of positive and negative CTV samples. Optical density (OD405) values after 120 min of hydrolisis of p-nitrophenyl phosphate (pNPP) for 16 CTV isolates were between 0.290 and 1.386 (X = 0.997) for Agdia kit, between 0.163 and 1.590 (X = 0.977) for Bioreba, between 0.250 and 2.049 (X = 1.367) for Ingenasa, and between 0.250 y 2.023 (X = 1.183) for Sanofi. For healthy controls after 120 min of hydrolysis of pNPP, Ingenasa and Sanofi kits yielded OD405 readings lower than 0.100; whereas, the kits from Agdia and Bioreba yielded values between 0.100 y 0.150, respectively. According to these results, the four kits can be reliably used for the serological detection of CTV. The selection of any of them would be based upon cost and product availability. Results of this study are also useful for any research work with CTV, but mostly for those certified laboratories devoted for the routine detection of CTV in Mexico. [ABSTRACT FROM AUTHOR]

An in situ immunoassay is described for localization of Citrus tristeza virus (CTV) inclusion bodies by light microscopy with antibodies developed to the recombinant p20 non-structural protein. Free hand bark sections of ca. 100 μm tick from healthy or CTV infected plants were fixed with 70 % ethanol and subsequently incubated with specific polyclonal antiserum anti-p20 either from rat or rabbit at dilutions 1:5,000 y 1:10,000 y 1:1,000 and 1:3,000 respectively and IgG anti species at 1:30,000 dilution; at the end, the chromogenic solution NBT/BCIP was added. Viral inclusion bodies were visualized by light microscopy at 10 and 40X magnifications as dark blue irregular areas in phloem associated tissues in CTV infected plants; no dark blue color was observed in the phloem tissues of healthy plants. [ABSTRACT FROM AUTHOR]

El objetivo de este trabajo the identificar marcadores DNA Polimórfico Amplificado al Azar (RAPDs) y Secuencias Simples Repetidas (SSRs) también conocidos como Microsatélites asociados con la resistencia al daño causado por fitoplasmas en papa. Se evaluó la severidad de la punta morada de la papa (PMP) y la aparición de color pardo en el tubérculo, así como también se realizaron análisis moleculares en variedades y líneas avanzadas de papa. El estudio se realizó en el laboratorio de biotecnología de la Facultad de Ciencias Biológicas de la Universidad Autónoma de Nuevo León - Instituto Nacional de Investigaciones Forestales Agrícolas y Pecuarias (INIFAP), México en el 2003 y 2004. Las técnicas utilizadas fueron RAPDs y SSRs también conocido microsatélites. Tres genotipos mostraron resistencia a la aparición del color pardo del tubérculo, mientras que del análisis molecular seis materiales formaron un grupo que incluyó los genotipos con valores de daño mínimos (entre uno y tres de la escala). Los genotipos restantes formaron dos grupos con presencia de color pardo en el tubérculo mayor (entre cinco y siete). Se identificaron marcadores moleculares posiblemente asociados con tolerancia a la coloración parda del tubérculo. En el análisis de componentes principales y el dendograma generado los materiales que presentaron menor severidad de daño, quedaron incluidos en un mismo grupo, mientras que materiales con mayor severidad de daño quedaron en un grupo diferente. [ABSTRACT FROM AUTHOR]

Polyclonal antibodies specific for the recombinant coat protein (rCP) p25 gene of Citrus tristeza virus (CTV), were developed for isolates MX08 and MX14 from Mexico and B227 from India. The reactivity of rCP antibodies was evaluated using healthy and CTV infected tissue. The combination of rCP antibodies developed in goats for isolate MX14 used as primary (trapping) antibodies, and rCP antibodies developed in rabbits for isolates MX08, MX14 y B227 as for antigen detection, reacted efficiently, with optical density (OD405) values between 0.400 and 1.400 for CTV infected tissue and lower than 0.100 for healthy plant tissue. The combination of rCP antibodies TI (goat anti-CTV MX14) as primary antibody and C3 (rabbit anti-CTV MX08) as detection antibody was as equally effective as the commercial kit from Agdia for positive and negative sample discrimination from a collection of 32 CTV isolates from Nuevo Leon, Mexico. In addition, the rCP from isolates MX 14 and B227 mixed with healthy tissue, were equally effective as positive control in ELISA tests, with OD405 values in the range of 0.300 and 0.400. [ABSTRACT FROM AUTHOR]