Characterization and localization of antigens for serodiagnosis of human paragonimiasis

Paragonimiasis is a foodborne trematode infection that affects 23 million people, mainly in Asia. Lung fluke infections lead frequently to chronic cough with fever and hemoptysis, and are often confused with lung cancer or tuberculosis. Paragonimiasis can be efficiently treated with praziquantel, but diagnosis is often delayed, and patients are frequently treated for other conditions. To improve diagnosis, we selected five Paragonimus kellicotti proteins based on transcriptional abundance, recognition by patient sera, and conservation among trematodes and expressed them as His-fusion proteins in Escherichia coli. Sequences for these proteins have 76–99% identity with amino acid sequences for orthologs in the genomes of Paragonimus westermani, Paragonimus heterotremus, and Paragonimus miyazakii. Immunohistology studies showed that antibodies raised to four recombinant proteins bound to the tegument of adult P. kellicotti worms, at the parasite host interface. Only a known egg antigen was absent from the tegument but present in developing and mature eggs. We evaluated the diagnostic potential of these antigens by Western blot with sera from patients with paragonimiasis (from MO and the Philippines), fascioliasis, and schistosomiasis, and with sera from healthy North American controls. Two recombinant proteins (a cysteine protease and a myoglobin) showed the highest sensitivity and specificity as diagnostic antigens, and they detected antibodies in sera from paragonimiasis patients with early or mature infections. In contrast, antibodies to egg yolk ferritin appeared to be specific marker for patients with adult fluke infections that produce eggs. Our study has identified and localized antigens that are promising for serodiagnosis of human paragonimiasis.

Comparative genomics and transcriptomics of 4 Paragonimus species provide insights into lung fluke parasitism and pathogenesis

Background Paragonimus spp. (lung flukes) are among the most injurious foodborne helminths, infecting ∼23 million people and subjecting ∼292 million to infection risk. Paragonimiasis is acquired from infected undercooked crustaceans and primarily affects the lungs but often causes lesions elsewhere including the brain. The disease is easily mistaken for tuberculosis owing to similar pulmonary symptoms, and accordingly, diagnostics are in demand. Results We assembled, annotated, and compared draft genomes of 4 prevalent and distinct Paragonimus species: Paragonimus miyazakii, Paragonimus westermani, Paragonimus kellicotti, and Paragonimus heterotremus. Genomes ranged from 697 to 923 Mb, included 12,072–12,853 genes, and were 71.6–90.1% complete according to BUSCO. Orthologous group analysis spanning 21 species (lung, liver, and blood flukes, additional platyhelminths, and hosts) provided insights into lung fluke biology. We identified 256 lung fluke–specific and conserved orthologous groups with consistent transcriptional adult-stage Paragonimus expression profiles and enriched for iron acquisition, immune modulation, and other parasite functions. Previously identified Paragonimus diagnostic antigens were matched to genes, providing an opportunity to optimize and ensure pan-Paragonimus reactivity for diagnostic assays. Conclusions This report provides advances in molecular understanding of Paragonimus and underpins future studies into the biology, evolution, and pathogenesis of Paragonimus and related foodborne flukes. We anticipate that these novel genomic and transcriptomic resources will be invaluable for future lung fluke research.

Parasites and Worms

A parasite is an organism that lives on or in a host and gets its food from or at the expense of its host. Worms or helminths either live as parasites or free of a host in aquatic and terrestrial environments. Parasites and worms are found worldwide but mainly in the tropics. It is estimated that 20% of immigrants from endemic countries may have helminthic infections at their arrival to the UK. These people could be asymptomatic, but tend to present with unexplained symptoms, especially gastrointestinal in nature or eosinophilia. Travellers to endemic countries tend to be newly infected and have greater immune response and pronounced eosinophilia in some but not all parasitic infections. Parasites that can cause disease in humans fall under three classes: protozoa, helminths, and Ectoparasites Protozoa are microscopic, one- celled organisms that can be free living or parasitic in nature. Transmission of protozoa that live in a human’s intestine to another human typically occurs through a faeco-oral route (for example, contaminated food or water, or person- to-person contact). Protozoa that live in the blood or tissue of humans are transmitted to other humans by an arthropod vector (for example, through the bite of a mosquito or sand fly). Helminths are large, multicellular organisms that are generally visible to the naked eye in their adult stages. Like protozoa, helminths can be either free living or parasitic. There are three main groups of helminths that parasitize humans: cestodes, trematodes, and nematodes. These are flat worms that comprise Echinococcus species: intestinal tapeworms and neurocysticercosis (Taenia solium) These are leaf- shaped, and they vary in length from a few millimetres to 8 cm. They include: ■ Liver fluke: Clonorchis sinensis, Fasciola hepatica ■ Intestinal fluke: Fasciola buski, Heterophyes heterophyes, ■ Lung fluke: Paragonimus westernmani ■ Blood flukes: Schistosoma species These are cylindrical in structure. Blood- sucking arthropods such as mosquitoes are considered as ectoparasites because they depend on blood meal for their survival. Narrowly speaking, ectoparasites include organisms like ticks, fleas, lice, and mites (scabies) that attach or burrow into the skin and remain there for relatively long periods of time (e.g. weeks to months).

The complete mitochondrial genome of Paragonimus ohirai (Paragonimidae: Trematoda: Platyhelminthes) and its comparison with P. westermani congeners and other trematodes

We present the complete mitochondrial genome of Paragonimus ohirai Miyazaki, 1939 and compare its features with those of previously reported mitochondrial genomes of the pathogenic lung-fluke, Paragonimus westermani, and other members of the genus. The circular mitochondrial DNA molecule of the single fully sequenced individual of P. ohirai was 14,818 bp in length, containing 12 protein-coding, two ribosomal RNA and 22 transfer RNA genes. As is common among trematodes, an atp8 gene was absent from the mitogenome of P. ohirai and the 5′ end of nad4 overlapped with the 3′ end of nad4L by 40 bp. Paragonimusohirai and four forms/strains of P. westermani from South Korea and India, exhibited remarkably different base compositions and hence codon usage in protein-coding genes. In the fully sequenced P. ohirai individual, the non-coding region started with two long identical repeats (292 bp each), separated by tRNAGlu. These were followed by an array of six short tandem repeats (STR), 117 bp each. Numbers of the short tandem repeats varied among P. ohirai individuals. A phylogenetic tree inferred from concatenated mitochondrial protein sequences of 50 strains encompassing 42 species of trematodes belonging to 14 families identified a monophyletic Paragonimidae in the class Trematoda. Characterization of additional mitogenomes in the genus Paragonimus will be useful for biomedical studies and development of molecular tools and mitochondrial markers for diagnostic, identification, hybridization and phylogenetic/epidemiological/evolutionary studies.

Paragonimusand paragonimiasis in West and Central Africa: unresolved questions

Paragonimiasis, human lung fluke disease, is a foodborne anthropozoonosis caused by the trematodes assigned toParagonimusand is regarded by the World Health Organization as a Neglected Tropical Disease (NTD). The life cycle of this medically important parasite centres on a complex freshwater biological community that includes two intermediate hosts: a mollusc and a decapod, usually a brachyuran. Although there is a perception that the biology, symptoms, diagnosis and treatment ofParagonimusis well understood, in reality, this is not the case, especially in Africa. Much remains unknown concerning the life-cycle of the parasite, its transmission, the current epidemiology of the disease, diagnosis and the effectiveness of treatment. Furthermore, cases of paragonimiasis may be misdiagnosed as resistant tuberculosis (TB) because of the similar pulmonary symptoms and no remission after anti TB therapy. The endemic foci of human paragonimiasis in Africa have been reported mainly in the forest zones of Upper Guinea (Liberia, Guinea and Ivory Coast) and Lower Guinea (Nigeria, Cameroon, Equatorial Guinea and Gabon). Despite the perceived medical importance of paragonimiasis, relatively little attention has been paid to this NTD since its discovery in Africa in the 1960s. This review focuses on the current understanding of the life cycle and transmission ofParagonimusin Africa, discusses its diagnosis and public health importance and highlights many outstanding gaps in the knowledge that still exist for this NTD.