Project description:The protozoan Ichthyophthirius multifiliis (Ich) is a eukaryotic ciliate parasite of freshwater fish. Ich causes ichthyophthiriosis or ‘white spot disease’ characterized by white cysts covering the host skin and gills. The parasite is responsible for high mortalities and severe economic losses to farmed species as well as to ornamental species of fish. Despite the global importance of Ich, little is known about the genetic processes underlying its development and infectivity. Ich has three main life-stages, an infective theront, a parasitic trophont, and a reproductive tomont. To compare gene expression among Ich life-stages, oligonucleotide microarrays were constructed and utilized. All publicly-available Ich ESTs (~35K) were clustered to generate 9,129 unique consensus sequences represented as probes on custom microarrays produced in coordination with Roche NimbleGen. To facilitate comparative genomic analysis and to potentially increase gene content through cross-hybridization, gene coding sequences of related protozoans Tetrahymena thermophila and Plasmodium falciparum were also added to the microarrays. Gene expression was analyzed in samples taken from each of the three Ich life-stages. The results of this study will add in the understanding of protozoan global gene regulation and biology and should aid in the development of strategies aimed at the control of this important fish parasite. Submitted is a nine chip oligo array design using 385 K Nimblegen arrays. A total of nine microarrays were used for the experiment: three replicates from each of the three Ich life-stages (tomont, trophont, and theront life-stages). Probes were designed using 9,129 unique Ich ESTs (clustered contigs and singletons) as well as 26,273 Tetrahymena thermophila and 5,184 Plasmodium falciparum coding sequences. The probe design strategy was to create 12 60-mer oligonucleotide probes per I. multifiliis sequence, and 10 60-mer oligonucleotide probes for both T. thermophila and P. falciparum sequences. Total RNA was isolated in triplicate from the three life-stages of I. multifiliis and submitted to Nimblegen for labeling, hybridization, and scaning.

Project description:The protozoan Ichthyophthirius multifiliis (Ich) is a eukaryotic ciliate parasite of freshwater fish. Ich causes ichthyophthiriosis or ‘white spot disease’ characterized by white cysts covering the host skin and gills. The parasite is responsible for high mortalities and severe economic losses to farmed species as well as to ornamental species of fish. Despite the global importance of Ich, little is known about the genetic processes underlying its infectivity. Ich has three main life-stages, an infective theront, a parasitic trophont, and a reproductive tomont. Further, Ich has been demonstrated previously to display a loss of infectivity as the number of lab-passages on a fish increase, presumably relating to senescence of the organism. To compare gene expression among two of the three Ich life-stages (the tomont and trophont life-stages) at different passages, oligonucleotide microarrays were utilized. Gene expression was analyzed in samples taken from two of the three Ich life-stages (the tomont and trophont life-stages) at the first serial passage on channel catfish in the lab (P1), and at serial passage 100 (P100). The results of this study will add in the understanding of protozoan global gene regulation and biology and should aid in the development of strategies aimed at the control of this important fish parasite. Submitted is a 12 chip oligo array design using 385 K Nimblegen arrays. A total of 12 microarrays were used for the experiment: three replicates from two of the three Ich life-stages (tomont and trophont life-stages) at serial passage #1 (P1) and serial passage #100 (P100). Probes were designed using 9,129 unique Ich ESTs (clustered contigs and singletons) as well as 26,273 Tetrahymena thermophila and 5,184 Plasmodium falciparum coding sequences. The probe design strategy was to create 12 60-mer oligonucleotide probes per I. multifiliis sequence, and 10 60-mer oligonucleotide probes for both T. thermophila and P. falciparum sequences. Total RNA was isolated in triplicate from the three life-stages of I. multifiliis and submitted to Nimblegen for labeling, hybridization, and scaning. This microarray study is based on the GPL9449 platform built for the developmental stages of the parasite.

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Rickettsia endosymbiont of Ichthyophthirius multifiliis

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