Phytomonas iron superoxide dismutase: a possible molecular marker

Abstract

We have isolated and biochemically characterized two iron superoxide dismutases activities (SODI and SODII) from a plant trypanosomatid isolated from Euphorbia characias. The isoenzyme FeSODII has immunogenic capacity, and the positivity of the anti-SODII serum persists to a dilution of 1/40,000, by Western blot. In addition, Western blot has been used to test the positivity of the anti-SODII serum against antigen fractions (SOD) from 17 isolates belonging to the family Trypanosomatidae and for which we had previously determined the isoenzymatic profile. The reaction proved positive only with those plant isolates considered to belong to the genus Phytomonas, whereas there was no reaction of the anti-SODII serum, against the antigen fractions from the species Trypanosoma cruzi, Leishmania donovani, Herpetomonas samuelpessoai, Herpetomonas davidi, Crithidia luciliae and Leptomonas collosoma. FeSODII is located mainly over the entire surface of the parasite, as well as in the nucleus, glycosomes and membranes. The above makes FeSODII promising as a molecular tool for diagnosis and identification, and as a potential chemotherapeutic target for designing drugs aimed at controlling not only of the diseases caused by Phytomonas species, but also for the great metabolic similarity to other trypanosomatids of animals and humans, it may be possible for these results to be extrapolated. Moreover, the sequencing of the amino-terminal end of the FeSODII enables the design of primers that in the near future will make it possible to sequence the gene of this isoenzyme.

Identification of excreted iron superoxide dismutase for the diagnosis of Phtytomonas

“…These fractions were separated by electrophoresis of IEF 3-9 in polyacrylamide gels as described elsewhere (Marín et al 2004a). In this way, we obtained the anti-SODeCHA and anti-SODeTOM sera.…”

“…After being washed as above, the membrane was further incubated for 2 h at room temperature with the second antibody, anti-mouse IgG (Fc specific) peroxidase conjugate (Sigma®) (dilution 1/1000). After washing, the substrate diaminobenzidine was added and the reaction stopped with several washes in distilled water (Marín et al 2004a). For the cross reaction analyses, we followed the same protocol described above but with a dilution of the anti-SODeCHA and anti-SODeTOM sera of 1/5000.…”

“…Attempts to overcome this difficulty prompted the testing of various serological, biochemical, and molecular methods, but none to date have been very successful for genus distinction (Camargo 1999). In recent years, new polymerase chain reaction method for the amplification of the SL sequence have been successfully developed, and more recently new molecular markers have become available to help in the identification (Serrano et al 1999, Dollet et al 2001, Marín et al 2004a.…”

“…SOD activity has been detected in the main species belonging to the family Trypanosomatidae, in Trypanosoma cruzi , in T. brucei brucei (Kabiri & Steverding 2001), in several species of the genus Leishmania (Ismail et al 1994), in some lower trypanosomatids, and in plant trypanosomes (Quesada et al 2001, Marín et al 2004a. SODs from protozoan parasites are considered virulence factors that protect the parasites from the attack of the host cells by the action both of oxidant and anti-inflammatory agents (Paramchuck et al 1997), and can even confer immunological capacities (Pérez-Fuentes et al 2003).…”

Diversidade, relações filogenéticas e taxonomia de Phytomonas spp.

“…Divergências de gGAPDH superiores a 1-3% tem sido consideradas suficientes para separar espécies de tripanossomatideos dependendo do gênero (Teixeira et al, 2011;Borghesan et al, 2013 Sturn et al, 2007), tentativas de cultivo de Phytomonas de coqueiros não foram bem sucedidas em nosso laboratório. Os isolados de látex são um pouco menos exigentes quanto ao cultivo, mas ainda assim, difíceis de isolar e de crescimento irregular (Camargo, 1999;Marín et al, 2004;Miranda et al, 2004). Por outro lado, isolados de frutos são, em geral, fáceis de cultivar (Camargo, 1999;Catarino et al, 2001).…”