A synthetic workflow for coordinated direct observation and genetic tagging applied to a complex host-parasite interaction.

An important aspect influencing host specificity is a parasite's compatibility, or ability, to infect a potential host. Here, we examine the compatibility between different trematode genotypes of the same species and several host species. To execute this study, we developed a synthetic workflow which combines the use of a fluorescent dye and standard molecular techniques to study host-parasite interactions and host specificity. The utility of the fluorescent dye, BIODIPY FL C12, was evaluated to label and track larval trematodes during experimental infections using the Cerithidea californica-trematode host-parasite system. Our results showed that low dye concentrations (200 nM) did not significantly affect survival or infectivity of Acanthoparyphium spinulosum and proved to be useful for labeling cercariae. Parasites were genotyped based on sequences from cytochrome oxidase subunit 1 (COI) and the nuclear internal transcribed spacer 1 (ITS1) prior to labeling and experimental infections. Samples with low COI PCR product yield were reamplified using the M13 tails to obtain enough material for sequencing. Three parasite genotypes were recovered and results from experimental infections demonstrated varying levels of host specificity. Of the three host species used (C. californica, Polydora nuchalis, Tagelus californianus), genotype B was unable to infect P. nuchalis. Genotype A individuals were less likely to infect P. nuchalis than the other host species. Additionally, genotype C was unable to infect any host offered in this study. These findings reflect possible suboptimal pairings between parasite genotype and host species. Furthermore, the present study provides procedures that are useful for exploring parasite ecology at the molecular level.

Monitoring of methyl jasmonate-responsive genes in Arabidopsis by cDNA macroarray: self-activation of jasmonic acid biosynthesis and crosstalk with other phytohormone signaling pathways.

Jasmonates mediate various physiological events in plant cells such as defense responses, flowering, and senescence through intracellular and intercellular signaling pathways, and the expression of a large number of genes appears to be regulated by jasmonates. In order to obtain information on the regulatory network of jasmonate-responsive genes (JRGs) in Arabidopsis thaliana (Arabidopsis), we screened 2880 cDNA clones for jasmonate responsiveness by a cDNA macroarray procedure. Since many of the JRGs reported so far have been identified in leaf tissues, the cDNA clones used were chosen from a non-redundant EST library that was prepared from above-ground organs. Hybridization to the filters was achieved using alpha-33P-labeled single-strand DNAs synthesized from mRNAs obtained from methyl jasmonate (MeJA)-treated and untreated Arabidopsis seedlings. Data analysis identified 41 JRGs whose mRNA levels were changed by more than three fold in response to MeJA. This was confirmed by Northern blot analysis by using eight representatives. Among the 41 JRGs identified, 5 genes were JA biosynthesis genes and 3 genes were involved in other signaling pathways (ethylene, auxin, and salicylic acid). These results suggest the existence of a positive feedback regulatory system for JA biosynthesis and the possibility of crosstalk between JA signaling and other signaling pathways.