Interaction of cowpea with Phytophthora vignae: inheritance of resistance and production of phenylalanine ammonia-lyase as a resistance response

The interactions between Phytophthora vignae and two cowpea cultivars are described. The cultivars are near-isogenic but differ in their resistance to P. vignae race 2. Cultivar Caloona is resistant, cv. Poona is susceptible. Resistance of cv. Caloona to race 2 is controlled by a single dominant gene and is expressed throughout the life of the plant in all tissues at temperatures up to 35°C. Inoculation of cut cowpea epicotyls with a plug of agar containing race 2 hyphae provides a reliable method of obtaining host tissue which responds in a predictable way to the pathogen. Phenylalanine ammonia-lyase (PAL) activity increases rapidly in invaded epicotyl tissue of cv. Caloona in response to inoculation with race 2. Increased PAL activity in advance of the hyphae is not detected. Low levels of PAL are produced by cv. Poona and cv. Caloona in response to inoculation with a race virulent on both cultivars (race 3). Application of l-2-aminooxy-3-phenylpropionic acid (1 mm) or aminooxy acetic acid (10 mm) through the cut bases of hypocotyls of cv. Caloona seedlings results in this cultivar, normally resistant to P. vignae race 2, becoming susceptible

Procyclin Null Mutants of Trypanosoma brucei Express Free Glycosylphosphatidylinositols on Their Surface

Procyclins are abundant, glycosylphosphatidylinositol (GPI)-anchored proteins on the surface of procyclic (insect) form trypanosomes. To investigate whether trypanosomes are able to survive without a procyclin coat, all four procyclin genes were deleted sequentially. Bloodstream forms of the null mutant exhibited no detectable phenotype and were able to differentiate to procyclic forms. Initially, differentiated null mutant cells were barely able to grow, but after an adaptation period of 2 mo in culture they proliferated at the same rate as wild-type trypanosomes. Analysis of these culture-adapted null mutants revealed that they were covered by free GPIs. These were closely related to the mature procyclin anchor in structure and were expressed on the surface in numbers comparable with that of procyclin in wild-type cells. However, free GPIs were smaller than the procyclin anchor, indicative of a lower number of poly-N-acetyllactosamine repeats, and a proportion contained diacylphosphatidic acid. Free GPIs are also expressed by wild-type cells, although to a lesser extent. These have been overlooked in the past because they partition in a solvent fraction (chloroform/water/methanol) that is normally discarded when GPI-anchored proteins are purified

Virulence Attenuation of a UDP-galactose/ N-acetylglucosamine β1,4 Galactosyltransferase Expressing Leishmania donovani Promastigote

Protozoan parasites of the genus Leishmania are the causative agent of leishmaniasis, a disease whose manifestations in humans range from mild cutaneous lesions to fatal visceral infections. Human visceral leishmaniasis is caused by Leishmania donovani. Long-term culture in vitro leads to the attenuation of the parasite. This loss of parasite virulence is associated with the expression of a developmentally regulated UDP-Galactose/N-acetylglucosamine β 1–4 galactosyltransferase and galactose terminal glycoconjugates as determined by their agglutination with the pea nut agglutinin (PNA). Thus, all promastigotes passaged for more than 11 times were 100% agglutinated with PNA, and represent a homogeneous population of avirulent parasites. Identical concentrations of PNA failed to agglutinate promastigotes passaged for ≤5 times. These PNA− promastigotes were virulent. Promastigotes passaged from 5 to 10 times showed a mixed population. The identity of populations defined by virulence and PNA agglutination was confirmed by isolating PNA+ avirulent and PNA− virulent clones from the 7th passage promastigotes. Only the PNA+ clones triggered macrophage microbicidal activity. The PNA+ clones lacked lipophosphoglycan. Intravenous administration of [14C] galactose-labeled parasite in BALB/c mice resulted in rapid clearance of the parasite from blood with a concomitant accumulation in the liver. By enzymatic assay and RT-PCR we have shown the association of a UDP-Galactose/Nacetylglucosamine β1,4 galactosyltransferase with only the attenuated clones. By immunofluorescence we demonstrated that the enzyme is located in the Golgi apparatus. By western blot analysis and SDS-PAGE of the affinitypurified protein, we have been able to identify a 29 KDa galactose terminal protein from the avirulent clones