35 research outputs found

    Validation of a death assay for Angiostrongylus cantonensis larvae (L3) using propidium iodide in a rat model (Rattus norvegicus)

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    Angiostrongylus cantonensis is a pathogenic nematode and the cause of neuroangiostrongyliasis, an eosinophilic meningitis more commonly known as rat lungworm disease. Transmission is thought to be primarily due to ingestion of infective third stage larvae (L3) in gastropods, on produce, or in contaminated water. The gold standard to determine the effects of physical and chemical treatments on the infectivity of A. cantonensis L3 larvae is to infect rodents with treated L3 larvae and monitor for infection, but animal studies are laborious and expensive and also raise ethical concerns. This study demonstrates propidium iodide (PI) to be a reliable marker of parasite death and loss of infective potential without adversely affecting the development and future reproduction of live A. cantonensis larvae. PI staining allows evaluation of the efficacy of test substances in vitro, an improvement upon the use of lack of motility as an indicator of death. Some potential applications of this assay include determining the effectiveness of various anthelmintics, vegetable washes, electromagnetic radiation and other treatments intended to kill larvae in the prevention and treatment of neuroangiostrongyliasis

    The Tripartite Type III Secreton of Shigella flexneri Inserts Ipab and Ipac into Host Membranes

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    Bacterial type III secretion systems serve to translocate proteins into eukaryotic cells, requiring a secreton and a translocator for proteins to pass the bacterial and host membranes. We used the contact hemolytic activity of Shigella flexneri to investigate its putative translocator. Hemolysis was caused by formation of a 25-Å pore within the red blood cell (RBC) membrane. Of the five proteins secreted by Shigella upon activation of its type III secretion system, only the hydrophobic IpaB and IpaC were tightly associated with RBC membranes isolated after hemolysis. Ipa protein secretion and hemolysis were kinetically coupled processes. However, Ipa protein secretion in the immediate vicinity of RBCs was not sufficient to cause hemolysis in the absence of centrifugation. Centrifugation reduced the distance between bacterial and RBC membranes beyond a critical threshold. Electron microscopy analysis indicated that secretons were constitutively assembled at 37°C before any host contact. They were composed of three parts: (a) an external needle, (b) a neck domain, and (c) a large proximal bulb. Secreton morphology did not change upon activation of secretion. In mutants of some genes encoding the secretion machinery the organelle was absent, whereas ipaB and ipaC mutants displayed normal secretons

    High prevalence of \u3ci\u3eAngiostrongylus cantonensis\u3c/i\u3e (rat lungworm) on eastern Hawai`i Island: A closer look at life cycle traits and patterns of infection in wild rats (\u3ci\u3eRattus\u3c/i\u3e spp.)

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    The nematode Angiostrongylus cantonensis is a zoonotic pathogen and the etiological agent of human angiostrongyliasis or rat lungworm disease. Hawai`i, particularly east Hawai`i Island, is the epicenter for angiostrongyliasis in the USA. Rats (Rattus spp.) are the definitive hosts while gastropods are intermediate hosts. The main objective of this study was to collect adult A. cantonensis from wild rats to isolate protein for the development of a blood-based diagnostic, in the process we evaluated the prevalence of infection in wild rats. A total of 545 wild rats were sampled from multiple sites in the South Hilo District of east Hawai`i Island. Adult male and female A. cantonensis (3,148) were collected from the hearts and lungs of humanely euthanized Rattus rattus, and R. exulans. Photomicrography and documentation of multiple stages of this parasitic nematode in situ were recorded. A total of 45.5% (197/433) of rats inspected had lung lobe(s) (mostly upper right) which appeared granular indicating this lobe may serve as a filter for worm passage to the rest of the lung. Across Rattus spp., 72.7% (396/545) were infected with adult worms, but 93.9% (512/545) of the rats were positive for A. cantonensis infection based on presence of live adult worms, encysted adult worms, L3 larvae and/or by PCR analysis of brain tissue. In R. rattus we observed an inverse correlation with increased body mass and infection level of adult worms, and a direct correlation between body mass and encysted adult worms in the lung tissue, indicating that larger (older) rats may have developed a means of clearing infections or regulating the worm burden upon reinfection. The exceptionally high prevalence of A. cantonensis infection in Rattus spp. in east Hawai`i Island is cause for concern and indicates the potential for human infection with this emerging zoonosis is greater than previously thought
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