Meeting of the Trichomonad Community

December 2-4, 2003, Philadelphia, USA

 

A meeting of members of the Trichomonad commmunity took place over three days in December 2003 at the 52nd annual meeting of the American Society of Tropical Medicine and Hygiene (ASTMH) in Philadelphia. The meeting was divided into three stages: a reception and dinner welcoming all attendees on December 2; a one-day workshop "The Genome and Biology of the Trichomonads" for discussion of the T. vaginalis genome sequencing project and other research on December 3; and a two-hour symposium ÒMind the Gap: Bridging the Divide Between Clinical and Molecular Genomic Studies of the TrichomonadsÓ within the ASTMH meeting on December 4. Funding for the workshop was provided by The Ellison Medical Foundation, Burroughs Wellcome Fund, National Institute of Allergy and Infectious Diseases (NIAID), and The Institute for Genomic Research (TIGR). A brief description of the workshop follows.

 

The workshop commenced with an introductory lecture given by the grandfather of trichomonas research, Miklos Mueller (Rockefeller University). Miklos presented an overview of the biology of T. vaginalis, the "enigmatic eukaryote", with reference to the derivation of the hydrogenosome organelle. Three subsequent sessions focused on the genome, transcriptome and proteome of T. vaginalis and associated research. First, an update of the T. vaginalis genome sequencing project was given (Jane Carlton, TIGR), in conjunction with  analysis of some of the many repeat sequences that are being identified in the genome (Joana Silva, TIGR). In brief, the genome is much larger in size than previously thought, currently estimated at between 80-120 Mb, and contains many small repeat sequences of 1-2 kb which make predicting the size of the genome difficult and assembly of the genome sequence data complex. Presentations were given by Petrus Tang (Chang Gung University, Taiwan) and  Robert Hirt (Natural History Museum, London) concerning their expressed sequence tag (EST) projects and research. A list of all ongoing EST projects was given to all attendees (see Tvag_EST_Projects attachment). Some 70,000-80,000 ESTs are to be generated and made available shortly for the community through public databases. Petrus Tang curently has ~7,000 ESTs from various strains of T. vaginalis, and a brief description was given concerning these. Robert Hirt has analyzed 4,000 ESTs and identified a large number of diverse GTPases among them. Stepanka Vanacova (University of California Los Angeles, USA), described the first intron-containing genes identified by her research and from the genome sequence data. Finally, two proteomics projects that are starting to utilize the T. vaginalis genome sequence data to identify proteins of the hydrogenosome were presented by Katrin Henze (Duesseldorf University, Germany) and Rachel Schneider (University of California Los Angeles, USA). Katrin Henze described the identification of rubrerythrin in T. vaginalis, which is involved in an oxidative stress protection system in Desulfovibrio vulgaris. Rachel Schneider described the identification of the Hmp 35, 37, 31 and related proteins.

 

Subsequently, major topics were divided into Diagnosis and Pathogenesis, Biochemistry and Metabolism, and Clinical Studies and Drug Resistance. Martia Hobbs (University of North Carolina, USA) gave an overview of the methods used for T. vaginalis detection, and described a study of the male sexual partners of infected women, who were determined to have a high prevalence of infection. John Alderete (University of Texas, USA) entertained the audience with a video clip of different clumped and motile forms of parasite field isolates not visible in cultured lab isolates, and gave a description of a new commercially available diagnostic test. Patricia Johnson (University of California Los Angeles, USA) gave an update concerning analyses of core promoter elements, and described how the T. vaginalis genome sequence could be searched for additional promoters and transcription factors. Miklos Muller presented for a second time, on the core energy metabolism of the trichomonads which remains a puzzle and has yet to provide the answers to several key questions such as the redox balance of the organism in anaerobiosis. Finally, Jan Tachezy (Charles University, Czech Republic) presented an overview of his research on iron metabolism, and presented data on the FeS cluster assembly machinery newly identified in the hydrogenosome. In the Clinical Studies and Drug Resistance section, Peter and Jacqui Upcroft (Queensland Institute of Medical Research, Australia) presented work that suggests clinical isolates of T. vaginalis from various regions of the world (Australia, Papua New Guinea, South Africa and Los Angeles) have a varying number of chromosomes and karyotype. It remains to be seen whether the standard laboratory strains, such as the G3 strain currently being sequenced, exhibit such a phenomenon. Evan Secor (Centers for Disease Control, USA) presented data of a large number of drug resistant isolates that have been collected at CDC and the drug susceptibility tests that are routinely run at CDC, and reminded the audience that both of these are valuable resources that are available to the community.

 

A penultimate presentation by Lynette Corbeil (University of California San Diego, USA), gave an overview of immunity to bovine and human trichomoniasis, and provided evidence that women with trichomoniasis have vaginal IgG and IgA antibodies to the surface of T. vaginalis parasites. Finally, John Logsdon (University of Iowa, USA) gave an overview of genes known to be involved in the mechanics of meiosis, and their identification in the T. vaginalis genome data and from an independent genome survey sequence (GSS) project.

 

At the conclusion of the presentations, a round table discussion was opened to deliberate completion of the genome project and future research that might capitalize on the genome sequence. Bullet points from the discussion follow:

 

¥       The Entamoeba histolytica genome sequencing project:perspective from another community and another difficult genome

Brendan Loftus (TIGR) the PI on the project to sequence the genome of Entamoeba histolytica, gave an overview of the status of the E. histolytica genome sequencing project. The repetitive nature of the genome has led the Entamoeba community to accept that not all gaps in the sequence data will be closed, and that funds would be wasted to try to achieve such an unrealistic goal. Funds which were previously set aside for closure of the genome have subsequently been diverted into sequencing other related strains that will provide data for comparative genomics studies, as well as for the generation of an E. histolytica microarray chip consisting of all open reading frames which will be available to the community. A Steering Committee made up of investigators of the original grant application has been formed to channel comments to the project PI, and also to identify the researchers assigned to analyze particular genes or biological phenotypes and contribute their findings for a publication on the genome sequence.

 

¥        Completion of the T. vaginalis genome sequencing project

Jane Carlton discussed plans for completion of the T. vaginalis genome project. 30,000 ESTs will be generated from the C1 strain over the next 2-3 months, and made available to the community by Summer 2004. Due the the larger genome size than expected, the random phase of sequencing has taken longer than predicted. A large-insert BAC library is currently being generated and sequencing of more small-insert clones will increase the coverage of the data. As more sequence data is generated, updates to the T. vaginalis database will be made (http://www.tigr.org/tdb/e2k1/tvg/). However researchers are asked to abide by the terms of TIGR's Data Release Policy (http://www.tigr.org/tigr-scripts/license/new.pl?genre=euk) which state that scientists refrain from publishing any articles containing analyses of genes or genomic data on a whole genome or chromosome scale prior to TIGR's publication of its comprehensive genome analysis. Please contact the PI Jane Carlton with any such queries (carlton@tigr.org).

 

Due to the highly repetitive nature of the genome, it is unrealistic to aim for a completely closed genome sequence of T. vaginalis. In the light of this, the genome project will focus on producing the best possible assembly, incorporating repeats into the assembly where feasible using novel assembly algorithms. Complete telomere-to-telomere chromosome sequences are not a possibility. All unique coding regions will be sequenced. Closure will be limited to closing of sequence gaps only, time and funds permitting. Gene finding will use the data generated from the multiple EST projects as training data, and should be comprehensive enough to identify all genes reliably. Annotation will be automated with some manual curation of particular genes of interest.

 

Some discussion was given to post-annotation analysis of the genome and manuscript writing, although it was emphasized that this is likely to be premature at this early stage. It was decided that the community should identify individual labs for analysis of specific genes/pathways/biological phenotypes. As a preliminary guide, a draft list of researchers who may be interested in providing a paragraph for the genome paper, and their subject area, was generated. This included some researchers not present at the meeting but identified by the community as having made significant contributions to their field. This list is preliminary, it is not binding and requires extensive addition and discussion before implementation. It was explained that TIGR and the PI have ultimate editorial control over the finished manuscript.

 

Some discussion was given to sequencing other trichomonad genomes, in particular Tritrichomonas foetus and Trichomonas tenax. Due to the significantly larger than expected genome size and complexity of the T. vaginalis genome, it is highly unlikely that funds from the T. vaginalis project could be diverted to sequencing of another species unless there was justification that this would signifcantly improve the T. vaginalis genome sequence. If there is a need within the community to sequence further species, other funding sources could be applied to. There was continued discussion concerning the preferred species to sequence: this would have to be determined based upon genome size (it is not clear if all trichomonad species have a similar genome size to that of T. vaginalis), repetitive nature of the genome (T. foetus is also a highly repetitive genome) and evolutionary relatedness, among other considerations.

 

¥        Future research capitalizing on the genome sequence

            (i) Microarray projects

Petrus Tang informed the audience that his group plans on generating an EST-based microarray with at least 6,000 non-redundant ESTs by the end of 2004. These will be made available to the community for a small cost.

            (ii) Proteomic projects

No announcements were made concerning additional proteomics projects.

            (iii) Population genetics

Jane Carlton announced plans for her group to capitalize on the genome data by identifying genome-wide repeats (microsatellites and minisatellites) that can be developed into genetic markers for population and molecular evolution studies.

           

¥        Funding initiatives

Bob Quackenbush, NIAID, encouraged all researchers to submit through the usual mechanisms of R01, R21 etc. Although Trichomonas is not classified as a bioterrorism agent, the Sexually Transmitted Diseases branch of NIAID are lobbying for increased funding opportunities.

Victoria McGovern, The Burroughs Wellcome Fund, encouraged applications to the Fund's Investigators in Pathogenesis of Infectious Disease program which the Fund is acceptiong applications for in 2004.

 

¥       AOB

Donald Burgess (American Type Tissue Collection, USA) encouraged all researchers to deposit their trichomonad reagents at ATCC, as well as ordering reagents from the collection.

 

¥        Close 

At the close of the workshop, it was agreed that the meeting had been a great success and of value to all participants. It was proposed to hold a second meeting sometime in 2004, where an update on the progress of the T. vaginalis genome project could be given, and more concrete plans laid for the analysis and publication process. One possible location could be prior to the Woods Hole Molecular Parasitology Meeting  (MPM) in Massachusettes, USA, which is to be held from September 19-23, 2004. Participants at the meeting could then attend MPM. The MPM organizers are to be approached regarding the inclusion of a special Trichomonad session at the meeting.