Detection of polymorphism in the Trypanosoma cruzi TcP2β gene family by single strand conformational analysis (SSCA)

Single strand conformation analysis (SSCA) is a technique that has been used to detect point mutations. We explored its usefulness in the analysis of four different members of the Trypanosoma cruzi TcP2β gene family and its suitability for detection of polymorphism in different parasite strains. The availability of primers covering a 97-bp sequence at the 5′ end of the genes allowed assessment of the effect of a single base substitution, while the analysis of a 321 bp long sequence permitted the evaluation of sequences differing in several bases. PCR products were analysed under four different electrophoretic conditions: with or without the addition of 10% glycerol in a 6% polyacrylamide gel run at room temperature or at 4°C. Shifts in mobility were radically dependent on the migration condition. Both 97-bp and 321-bp amplicons were best resolved at 4°C, without glycerol. Amplification products derived from total genomic DNA showed a pattern that resembled closely a combination of the products derived from the cloned genes. The results herein demonstrate the usefulness of SSCA to differentiate forms of a complex protozoan gene family, and to scan its polymorphic nature. Furthermore, due to the remarkable sensitivity of the technique it can generate genomic markers, such as Sequence Tagged Sites (STS), of great need in the T. cruzi genome project     

Eggplant Mottled Dwarf Virus Associated with Vein Yellowing of Honeysuckle

A rhabdovirus isolated in Tunisia by mechanical inoculation from honeysuckle (Lonicera sp.) plants with vein yellowing, was compared with a Tunisian isolate of eggplant mottled dwarf virus (EMDV). The two viruses had similar host ranges and caused the same symptoms in artificially infected hosts. The honeysuckle virus induced in eggplant a syndrome indistinguishable from that typical of EMDV. The two viruses could not be differentiated serologically, had particles of the same size and elicited identical cytopathological alterations in naturally and artificially infected hosts. Honeysuckle is the first host, besides eggplant, found to be naturally infected with EMDV.