A functionally divergent hydrogenosomal peptidase with protomitochondrial ancestry

Matrix proteins of mitochondria, hydrogenosomes and mitosomes are typically targeted and translocated into their respective organelles using N-terminal presequences that are subsequently cleaved by a peptidase. Here we characterize a approximately 47 kDa metallopeptidase, from the hydrogenosome-bearing, unicellular eukaryote Trichomonas vaginalis, that contains the active site motif (HXXEHX(76)E) characteristic of the beta subunit of the mitochondrial processing peptidase (MPP) and localizes to hydrogenosomes. The purified recombinant protein, named hydrogenosomal processing peptidase (HPP), is capable of cleaving a hydrogenosomal presequence in vitro, in contrast to MPP which requires both an alpha and beta subunit for activity. T. vaginalis HPP forms an approximately 100 kDa homodimer in vitro and also exists in an approximately 100 kDa complex in vivo. Our phylogenetic analyses support a common origin for HPP and betaMPP and demonstrate that gene duplication gave rise to alphaMPP and betaMPP before the divergence of T. vaginalis and mitochondria-bearing lineages. These data, together with published analyses of MPPs and putative mitosomal processing peptidases, lead us to propose that the length of targeting presequences and the subunit composition of organellar processing peptidases evolved in concert. Specifically, longer mitochondrial presequences may have evolved to require an alpha/beta heterodimer for accurate cleavage, while shorter hydrogenosomal and mitosomal presequences did not.     

A proposed role for Leishmania major carboxypeptidase in peptide catabolism

Leishmaniasis is a tropical disease caused by Leishmania, eukaryotic parasites transmitted to humans by sand flies. Towards the development of new chemotherapeutic targets for this disease, biochemical and in vivo expression studies were performed on one of two M32 carboxypeptidases present within the Leishmania major (LmaCP1) genome. Enzymatic studies reveal that like previously studied M32 carboxypeptidases, LmaCP1 cleaves substrates with a variety of C-terminal amino acids—the primary exception being those having C-terminal acidic residues. Cleavage assays with a series of FRET-based peptides suggest that LmaCP1 exhibits a substrate length restriction, preferring peptides shorter than 9-12 amino acids. The in vivo expression of LmaCP1 was analyzed for each major stage of the L. major life cycle. These studies reveal that LmaCP1 expression occurs only in procyclic promastigotes—the stage of life where the organism resides in the abdominal midgut of the insect. The implications of these results are discussed.     

Development of ChillPeach genomic tools and identification of cold-responsive genes in peach fruit

The ChillPeach database was developed to facilitate identification of genes controlling chilling injury (CI), a global-scale post-harvest physiological disorder in peach. It contained 7,862 high-quality ESTs (comprising 4,468 unigenes) obtained from mesocarp tissues of two full-sib progeny contrasting for CI, about 48 and 13% of which are unique to Prunus and Arabidopsis, respectively. All ESTs are in the Gateway vector to facilitate functional assessment of the genes. The data set contained several putative SNPs and 184 unigenes with high quality SSRs, of which 42% were novel to Prunus. Microarray slides containing 4,261 ChillPeach unigenes were printed and used in a pilot experiment to identify differentially expressed genes in cold-treated compared to control mesocarp tissues, and in vegetative compared to mesocarp tissues. Quantitative RT-PCR (qRT-PCR) confirmed microarray results for all 13 genes tested. The microarray and qRT-PCR analyses indicated that ChillPeach is rich in putative fruit-specific and novel cold-induced genes. A website ( http://bioinfo.ibmcp.upv.es/genomics/ChillPeachDB ) was created holding detailed information on the ChillPeach database.     

Two similar enhanced root-colonizing Pseudomonas strains differ largely in their colonization strategies of avocado roots and Rosellinia necatrix hyphae

Pseudomonas alcaligenes AVO73 and Pseudomonas pseudoalcaligenes AVO110 were selected previously as efficient avocado root tip colonizers, displaying in vitro antagonism towards Rosellinia necatrix, causal agent of avocado white root rot. Despite the higher number of antagonistic properties shown in vitro by AVO73, only AVO110 demonstrated significant protection against avocado white root rot. As both strains are enhanced root colonizers, and as colonization is crucial for the most likely biocontrol mechanisms used by these strains, namely production of non-antibiotic antifungal compounds and competition for nutrients and niches, we decided to compare the interactions of the bacterial strains with avocado roots as well as with R. necatrix hyphae. The results indicate that strain AVO110 is superior in biocontrol trait swimming motility and establishes on the root tip of avocado plants faster than AVO73. Visualization studies, using Gfp-labelled derivatives of these strains, showed that AVO110, in contrast to AVO73, colonizes intercellular crevices between neighbouring plant root epidermal cells, a microhabitat of enhanced exudation. Moreover, AVO110, but not AVO73, also colonizes root wounds, described to be preferential penetration sites for R. necatrix infection. This result strongly suggests that AVO110 meets, and can attack, the pathogen on the root. Finally, when co-inoculated with the pathogen, AVO110 utilizes hyphal exudates more efficiently for proliferation than AVO73 does, and colonizes the hyphae more abundantly than AVO73. We conclude that the differences between the strains in colonization levels and strategies are likely to contribute to, and even can explain, the difference in disease-controlling abilities between the strains. This is the first report that shows that two similar bacterial strains, selected by their ability to colonize avocado root, use strongly different root colonization strategies and suggests that in addition to the total bacterial root colonization level, the sites occupied on the root are important for biocontrol.     

2‐(Undecyloxy)‐ethanol is a major component of the male‐produced aggregation pheromone of Monochamus sutor

The small white‐marmorated longicorn beetle, Monochamus sutor (L.) (Coleoptera: Cerambycidae), is widely distributed throughout Europe and Asia. It is a potential vector of the pine wood nematode, Bursaphelenchus xylophilus (Steiner et Buhrer) Nickle, the causal agent of the devastating pine wilt disease. Volatiles were collected from both male and female beetles after maturation feeding. In analyses of these collections using gas chromatography (GC) coupled to mass spectrometry, a single male‐specific compound was detected and identified as 2‐(undecyloxy)‐ethanol. In analyses by GC coupled to electroantennography the only consistent responses from both female and male antennae were to this compound. Trapping tests were carried out in Spain, Sweden, and China. 2‐(Undecyloxy)‐ethanol was attractive to both male and female M. sutor beetles. A blend of the bark beetle pheromones ipsenol, ipsdienol, and 2‐methyl‐3‐buten‐2‐ol was also attractive to both sexes in Spain and Sweden, and further increased the attractiveness of the 2‐(undecyloxy)‐ethanol. The host plant volatiles α‐pinene, 3‐carene, and ethanol were weakly attractive, if at all, in all three countries and did not significantly increase the attractiveness of the blend of 2‐(undecyloxy)‐ethanol and bark beetle pheromones. 2‐(Undecyloxy)‐ethanol is thus proposed to be the major, if not only, component of the male‐produced aggregation pheromone of M. sutor, and its role is discussed. This compound has been reported as a pheromone of several other Monochamus species and is another example of the parsimony that seems to exist among the pheromones of many of the Cerambycidae. Traps baited with 2‐(undecyloxy)‐ethanol and bark beetle pheromones should be useful for monitoring and control of pine wilt disease, should M. sutor be proven to be a vector of the nematode.     

A melissopalynological study of artisanal honey produced in Catamarca (Argentina)

Spores belonging to 43 taxa of pteridophytes were observed from the ambient air of 11 localities in India. It is suggested that pteridophyte spores could be listed among viable particulate air pollutants, especially in the light of the earlier reports regarding their allergenicity.     

A single locus determines sensitivity to bacterial flagellin in Arabidopsis thaliana

Peptides corresponding to the most conserved domain of eubacterial flagellin act as potent elicitors in cells of different plant species. In intact Arabidposis thaliana seedlings these peptides (flg22 and flg15) caused callose deposition, induction of genes coding for pathogenesis-related proteins and a strong inhibition of growth. Half-maximal growth inhibition occurred at peptide concentrations of approximately 100 nM. In contrast, peptides representing the corresponding flagellin domains of the plant-associated bacteria A. tumefaciens and R. meliloti were inactive even at concentrations of 10 microM. With the exception of Ws-0, all ecotypes of A. thaliana tested were sensitive to flg22. Crosses of Ws-0 with the sensitive ecotypes Col-0 and La-er, respectively, resulted in sensitive F1 seedlings. In the F2 generation of both crosses, sensitivity segregated as a single trait with markers of chromosome 5 and a ratio of 3:1. Dominance of the locus sensing flagellin, termed FLS-1, suggests that it encodes an element which is important for the perception of the flagellin signal.     

Development of a SYBR green I real-time PCR assay for specific identification of the fish pathogen Aeromonas salmonicida subspecies salmonicida

Applied Microbiology and Biotechnology - A SYBR Green I real-time polymerase chain reaction protocol for specific detection of the fish pathogen Aeromonas salmonicida subsp. salmonicida was...