Phylogenetic studies in Ravenelia esculenta and related rust fungi.

Ravenelia esculenta Naras. and Thium. is a rust fungus, which infects mostly thorns, inflorescences, flowers and fruits of Acacia eburnea Willd. Aecial stages of the rust produce hypertrophy in infected parts. DNA of the rust fungus was isolated from aeciospores by 'freeze thaw' method. 18S rDNA was amplified and sequenced by automated DNA sequencer. BLAST of the sequence at NCBI retrieved 96 sequences producing significant alignments. Multiple sequence alignment of these sequences was done by ClustalW. Phylogenetic analysis was done by using MEGA 3.1. UPGMA Minimum Evolution tree with bootstrap value of 1000 replicates was constructed using these sequences. From phylogenetic tree it is observed that Ravenelia esculenta and the genus Gymnosporangium share a common ancestry, though Ravenelia esculenta is autoecious on angiosperm and the genus Gymnosporangium is heteroecious with pycnia, aecia on angiosperm and uredia, telia on gymnosperm. Two major clades are recognized which are based on the nature of aecial host (gymnosperm or angiosperm). These clades were also showing shift from pteridophytes to angiosperms as telial hosts. The tree can be interpreted in the other way also where there is separation of 14 families of Uredinales depending upon nature of teliospores, nature of aeciospores and structure of pycnia. These studies determine the phylogenetic position of Ravenelia esculenta among other rust fungi besides broad separation of Uredinales into two clades. These studies also show that there is phylogenetic correlation between molecular and morphological data. This is first report of DNA sequencing and phylogenetic positioning in genus Ravenelia from India.

Autophagocytosis of carotenoid-rich lipid droplets into vacuoles during aeciospore ageing in Puccinia distincta.

• Aeciospores of rust fungi contain numerous lipid droplets (LDs) which are stained yellow due to the presence of ß-and γ-carotene, thereby facilitating observations of their fate during developmental processes. • The ageing process of aeciospores of Puccinia distincta was examined by light and transmission electron microscopy. • The cytoplasm of freshly harvested spores was filled with LDs but devoid of vacuoles. Within 12 h, numerous small vacuoles developed, and the LDs became associated with their cytoplasmic tonoplast surface. The vacuoles enlarged and fused together; they were also the destination of endocytotic uptake of the vital dye, Neutral Red. Within 36 h, LDs entered these vacuoles by autophagocytosis and were degraded in the vacuolar lumen. After 96 h, most lipid reserves had been used up and the highly vacuolated spores lost viability as indicated by their failure to germinate or take up Neutral Red. • Spore maturation by vacuolation thus seems to occur at the direct expense of metabolic energy stored as LDs and is an extremely rapid process in aeciospores of P. distincta.