L-Proline uptake in Crithidia deanei is influenced by its endosymbiont bacterium

Crithidia deanei, a monoxenic trypanosomatid, presents an endosymbiotic bacterium in its cytoplasm. Both the protozoan and the bacterium maintain intensive metabolic exchange, resulting in an interesting model to study the coevolution of metabolisms. The relevance of L-proline for the growth of C. deanei and its transport into these cells was studied. Both the endosymbiont-containing (wild) and the endosymbiont-free protozoa (aposymbiont or cured) strains, when grown in medium supplemented with L-proline, reached higher cell densities than those grown in unsupplemented media. We biochemically characterized the uptake of L-proline in both the wild (K(m)=0.153 +/- 0.022 mM, V(max)=0.239 +/- 0.011 nmol min(-1) per 4 x 10(7) cells) and the aposymbiont strains (K(m)=0.177 +/- 0.049 mM, V(max)=0.132 +/- 0.012 nmol min(-1) per 4 x 10(7) cells). These data suggest a single type of proline transporter whose activity is upregulated by the presence of the symbiotic bacterium. Proline transport was further characterized and was found to be insensitive to the extracellular concentration of Na(+), but sensitive to K(+) and pH. The abolition of proline uptake by respiratory chain inhibitors and valinomycin indicates that the proline transport in C. deanei is dependent on the plasma membrane K(+) gradient.

Wolbachia in Anastrepha Fruit Flies (Diptera: Tephritidae)

Endosymbiotic bacteria of the genus Wolbachia are widespread among arthropods and cause a variety of reproductive abnormalities, such as cytoplasmic incompatibility, thelytokous parthenogenesis, male-killing, and host feminization. In this study, we used three sets of Wolbachia-specific primers (16S rDNA, ftsZ, and wsp) in conjunction with the polymerase chain reaction (PCR), cloning and sequencing to study the infection of fruit flies (Anastrepha spp. and Ceratitis capitata) by Wolbachia. The flies were collected at several localities in Brazil and at Guayaquil, Ecuador. All of the fruit flies studied were infected with Wolbachia supergroup A, in agreement with the high prevalence of this group in South America. Phylogenetic analysis showed that the wsp gene was the most sensitive gene for studying the relationships among Wolbachia strains. The Wolbachia sequences detected in these fruit flies were similar to those such as wMel reported for other fruit flies. These results show that the infection of Anastrepha fruit flies by Wolbachia is much more widespread than previously thought.

Phylogenetic Validation of the Genera Angomonas and Strigomonas of Trypanosomatids Harboring Bacterial Endosymbionts with the Description of New Species of Trypanosomatids and of Proteobacterial Symbionts

We comparatively examined the nutritional, molecular and optical and electron microscopical characteristics of reference species and new isolates of trypanosomatids harboring bacterial endosymbionts. Sequencing of the V7V8 region of the small subunit of the ribosomal RNA (SSU rRNA) gene distinguished six major genotypes among the 13 isolates examined. The entire sequences of the SSU rRNA and glycosomal glyceraldehyde phosphate dehydrogenase (gGAPDH) genes were obtained for phylogenetic analyses. In the resulting phylogenetic trees, the symbiont-harboring species clustered as a major clade comprising two subclades that corresponded to the proposed genera Angomonas and Strigomonas. The genus Angomonas comprised 10 flagellates including former Crithidia deanei and C. desouzai plus a new species. The genus Strigomonas included former Crithidia oncopelti and Blastocrithidia cuiicis plus a new species. Sequences from the internal transcribed spacer of ribosomal DNA (ITS rDNA) and size polymorphism of kinetoplast DNA (kDNA) minicircles revealed considerable genetic heterogeneity within the genera Angomonas and Strigomonas. Phylogenetic analyses based on 16S rDNA and ITS rDNA sequences demonstrated that all of the endosymbionts belonged to the Betaproteobacteria and revealed three new species. The congruence of the phylogenetic trees of trypanosomatids and their symbionts support a co-divergent host-symbiont evolutionary history. (C) 2011 Elsevier GmbH. All rights reserved.