2009 Annual Report
1a.Objectives (from AD-416)
Objective 1: Employing single-gene and genomic approaches, improve diagnosis of protists and nematodes that parasitize major food animals and that facilitate establishment, internalization, and survival of bacterial pathogens in produce.
Subobjective A: better characterize the molecular epidemiology of parasitic coccidia and trichinella.
Subobjective B: better characterize those bacterophagous eukaryotic microbes that may convey and help establish in produce pathogenic bacteria.
Objective 2: Develop a molecular phylogeny of coccidia in fish in order to better define their potential risk to food safety and security, and in order to better understand the relationship between Eimeriidae (including the agents of avian coccidiosis) and the Sarcocystidae (including the agent of human toxoplasmosis).
Objective 3: Better define the historical and ongoing interactions among wildlife and livestock reservoirs of Toxoplasma gondii through comparative population genetic analysis.
1b.Approach (from AD-416)
Several genes will be sequenced from parasites obtained from a wide array of animals, both domesticated and wild. These will be compared to each other, and to sequences obtained from human beings, in order to define the diversity and epidemiology of these parasite species. Homologues will be characterized for genes whose global variation has already begun to be studied in T. gondii, including the Intergenic Spacer sequence between rRNA genes, beta tubulin introns, and others. Characterization of microsatellite alleles will be considered as a second approach which, although requiring a greater investment of time and resources, should provide greater population genetic resolution than is possible with current loci.
To advance interests in molecular diagnostics, in collaboration with Chinese colleagues, we reported non-invasive means to diagnose species of Sarcocystis in its various life-history stages, including oocysts excreted by definitive hosts. This was important as it represented the first such application for clinical cases of Sarcocystis hominis, a parasite of people acquired by consuming infected beef. In addition, an invited book chapter accepted for publication this year describes the application of molecular diagnostic methods to this and other species of Sarcocystis. Progress in genome sequencing was achieved, first on two isolates of Listeria monocytogenes of interest to food safety and then, having optimized protocols for the Illumina Genome Analyzer, on isolates of Trichinella spiralis.
Progress was achieved in elucidating the function and evolutionary history of a key determinant to virulence in the zoonotic parasite Toxoplasma gondii. This original report, published in the prestigious journal PLoS Genetics, illustrates the increase in virulence that resulted from increased expression of a kinase, secreted by the parasite, that regulates host cell physiology. Progress of a conceptual nature included reviews on the genomics and phylogenetics of Trichinella, on the evolution of Toxoplasma gondii, and on how human activities have influenced our exposure to zoonotic pathogens. Each of these reviews provides needed perspective on the forces that have governed, and continue to govern, how such parasites have diversified and spread.
Natural selection at a single locus has elevated the expression of a secreted kinase resulting in the increased virulence of the zoonotic parasite Toxoplasma gondii to mice. In a groundbreaking study published in the prestigious PLoS Genetics, ARS scientists at the Beltsville Agricultural Research Center, working with researchers at the Washington University Medical School of Saint Louis and Cambridge University, uncovered the evolutionary history of a virulence factor and demonstrated that certain parasites have a version which has grown in its capacity to induce disease in experimental animal infections, in part through an increase in this gene’s expression. A series of experiments demonstrated that replacing alternative versions of this gene is sufficient to induce a virulent phenotype. This project will enable future progress in understanding how such intracellular parasites communicate with, and alter their hosts.
Domesticating and transporting livestock has altered how zoonotic parasites have evolved. ARS researchers at the Beltsville Agricultural Research Center have identified a ‘genetic signature’ evident in several parasites of agricultural livestock, including parasites that may induce human disease. Such parasites, over an especially broad geographic expanse, are particularly lacking in neutral genetic diversity. This pattern, which contrasts with that evident among related parasites of wildlife hosts, seems to reflect human influence in the transmission and dissemination of such hosts. A distillation of this process, summarizing recent progress and outlining future research needs, was published by an ARS researcher at the Beltsville Agricultural Research Center in the prestigious journal Trends in Parasitology and a highly original research report describing this phenomenon in the agent of trichinellosis was published by an ARS research team, working with Italian and Chinese colleagues, in the journal Infection, Genetics, and Evolution.
Cysts of zoonotic sarcocystsis may be diagnosed using genetic tools. Morphologically indistinguishable ‘oocysts’ of many distinct coccidian parasite species are excreted by their definitive hosts. Better understanding of the epidemiology and clinical consequences of such infections require better means to diagnose such infections and differentiate among causative agents. Working with Chinese collaborators, ARS scientists at the Beltsville Agricultural Research Service demonstrated the efficacy of methods to accurately diagnose such cysts excreted by dogs, cats, and people who had become infected by eating meats contaminated by each of three distinct parasite species. This project provides the means for future, non-invasive surveys of human infection with such parasites in order to better assess the magnitude of public health risk they may represent.
5.Significant Activities that Support Special Target Populations
The work on toxoplasmosis is especially important to the health of pregnant women, fetal and newborn health, and the health of persons afflicted by HIV-AIDS.
Pritt, B., Trainer, T., Simmons-Arnold, L., Evans, M., Dunams, D.B., Rosenthal, B.M. 2008. Detection of Sarcocystis parasites in retail beef: A regional survey combining histological and genetic detection methods. Journal of Food Protection. 71(10):2144-2147.
Xiang, Z., Chen, X., Yang, L., Zhou, B., Jiang, R., Rosenthal, B.M., Lei, L., Luan, P., Liu, J., Attwood, S., Zuo, Y., Cui, L., Zhang, Y., Yang, Z. 2009. Non-invasive methods for identifyig oocysts of Sarcocystis spp. from definitive hosts. Parasitology International. 58(3):293-296.
Khan, A., Taylor, S., Ajioka, J., Rosenthal, B.M., Sibley, D. 2009. Selection at a single Locus leads to widespread expansion of Toxoplasma gondii lineages that are virulent in mice. PLoS Genetics. 5(3): e1000404.
Rosenthal, B.M. 2008. How human history has influenced geography and genetics of parasite populations. Trends in Parasitology. 25(2):67-70.
Rosenthal, B.M. 2009. Sarcocystis. In: Liu.,D., editor. Molecular Detection of Sarcocystis. Boca Raton, Florida; CRC Press. p. 731-739.