(Macro)autophagy is a bulk degradation process that mediates the clearance of long-lived proteins and organelles. Autophagy is initiated by double-membraned structures, which engulf portions of cytoplasm. The resulting autophagosomes ultimately fuse with lysosomes, where their contents are degraded. Although the term autophagy was first used in 1963, the field has witnessed dramatic growth in the last 5 years, partly as a consequence of the discovery of key components of its cellular machinery. In this review we focus on mammalian autophagy, and we give an overview of the understanding of its machinery and the signaling cascades that regulate it. As recent studies have also shown that autophagy is critical in a range of normal human physiological processes, and defective autophagy is associated with diverse diseases, including neurodegeneration, lysosomal storage diseases, cancers, and Crohn's disease, we discuss the roles of autophagy in health and disease, while trying to critically evaluate if the coincidence between autophagy and these conditions is causal or an epiphenomenon. Finally, we consider the possibility of autophagy upregulation as a therapeutic approach for various conditions.

https://www.physiology.org/doi/pdf/10.1152/physrev.00030.2009

Regulation of Mammalian Autophagy in Physiology and Pathophysiology

Malaria presents a diagnostic challenge to laboratories in most countries. Endemic malaria, population movements, and travelers all contribute to presenting the laboratory with diagnostic problems for which it may have little expertise available. Drug resistance and genetic variation has altered many accepted morphological appearances of malaria species, and new technology has given an opportunity to review available procedures. Concurrently the World Health Organization has opened a dialogue with scientists, clinicians, and manufacturers on the realistic possibilities for developing accurate, sensitive, and cost-effective rapid diagnostic tests for malaria, capable of detecting 100 parasites/microl from all species and with a semiquantitative measurement for monitoring successful drug treatment. New technology has to be compared with an accepted "gold standard" that makes comparisons of sensitivity and specificity between different methods. The majority of malaria is found in countries where cost-effectiveness is an important factor and ease of performance and training is a major consideration. Most new technology for malaria diagnosis incorporates immunochromatographic capture procedures, with conjugated monoclonal antibodies providing the indicator of infection. Preferred targeted antigens are those which are abundant in all asexual and sexual stages of the parasite and are currently centered on detection of HRP-2 from Plasmodium falciparum and parasite-specific lactate dehydrogenase or Plasmodium aldolase from the parasite glycolytic pathway found in all species. Clinical studies allow effective comparisons between different formats, and the reality of nonmicroscopic diagnoses of malaria is considered.

Rapid Diagnostic Tests for Malaria Parasites

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The Uses Of Argument

Acanthamoeba spp. are free-living amebae that inhabit a variety of air, soil, and water environments. However, these amebae can also act as opportunistic as well as nonopportunistic pathogens. They are the causative agents of granulomatous amebic encephalitis and amebic keratitis and have been associated with cutaneous lesions and sinusitis. Immuno compromised individuals, including AIDS patients, are particularly susceptible to infections with Acanthamoeba. The immune defense mechanisms that operate against Acanthamoeba have not been well characterized, but it has been proposed that both innate and acquired immunity play a role. The ameba's life cycle includes an active feeding trophozoite stage and a dormant cyst stage. Trophozoites feed on bacteria, yeast, and algae. However, both trophozoites and cysts can retain viable bacteria and may serve as reservoirs for bacteria with human pathogenic potential. Diagnosis of infection includes direct microscopy of wet mounts of cerebrospinal fluid or stained smears of cerebrospinal fluid sediment, light or electron microscopy of tissues, in vitro cultivation of Acanthamoeba, and histological assessment of frozen or paraffin-embedded sections of brain or cutaneous lesion biopsy material. Immunocytochemistry, chemifluorescent dye staining, PCR, and analysis of DNA sequence variation also have been employed for laboratory diagnosis. Treatment of Acanthamoeba infections has met with mixed results. However, chlorhexidine gluconate, alone or in combination with propamidene isethionate, is effective in some patients. Furthermore, effective treatment is complicated since patients may present with underlying disease and Acanthamoeba infection may not be recognized. Since an increase in the number of cases of Acanthamoeba infections has occurred worldwide, these protozoa have become increasingly important as agents of human disease.

Acanthamoeba spp. as agents of disease in humans.

Acanthamoeba is an opportunistic protozoan that is widely distributed in the environment and is well recognized to produce serious human infections, including a blinding keratitis and a fatal encephalitis. This review presents our current understanding of the burden of Acanthamoeba infections on human health, their pathogenesis and pathophysiology, and molecular mechanisms associated with the disease, as well as virulence traits of Acanthamoeba that may be targets for therapeutic interventions and/or the development of preventative measures.

Acanthamoeba: biology and increasing importance in human health

The authors present the first human case of Clinostomum pharyngitis in Taegu, Korea. The patient was a 56-year old male who visited an otolaryngology clinic due to foreign body sensation and pain of the pharyngeal region for 3-4 days. He used to eat raw fresh-water fish. Otolaryngological examinations revealed a living worm adhered to the right posterior pharyngeal wall. The worm removed was identified as C. complanatum after morphological observations. It is likely that more attention should be paid to eating raw fresh-water fish in Korea with regards to Clinostomum pharyngitis.

The first human case of Clinostomum complanatum (Trematoda: Clinostomidae) infection in Korea

We purified and characterized a serine proteinase secreted by Acanthamoeba healyi to evaluate it as a possible virulence factor in the pathogenesis of granulomatous amoebic encephalitis (GAE). Ammonium sulfate precipitated culture supernatant of A. healyi OC-3A strain was purified by chromatography on CM-Sepharose, Sephacryl-S200, and Q-2 anion-exchange columns. The purified 33-kDa enzyme had a pH optimum of 8.0 and a temperature optimum of 40 C. Phenylmethylsulfonylfluoride and diisopropyl fluorophosphate, serine proteinase inhibitors, diminished activity of the enzyme to near zero. In addition to types I and IV collagen and fibronectin, the main components of the extracellular matrix, other proteins such as fibrinogen, IgG, IgA, albumin, and hemoglobin were also degraded by the enzyme. The broad substrate specificity of this secreted serine proteinase suggests that it may play an important role in pathogenesis of GAE by A. healyi.

Purification and characterization of a secretory serine proteinase of acanthamoeba healyi isolated from gae

Members of the genus Acanthamoeba, amphizoic protozoan parasites, are causative agents of granulomatous amoebic encephalitis and amoebic keratitis. Proteinases play a role in various biologic actions in Acanthamoeba, including host tissue destruction, pathogenesis, and digestion of phagocytosed food. Interestingly, we found that encystation of Acanthamoeba was inhibited by the serine proteinase inhibitor phenylmethanesulfonyl fluoride. In this study, we characterize a serine proteinase that is involved in mediating the encystation of Acanthamoeba. This encystation-mediating serine proteinase (EMSP) is shown to be highly expressed during encystation by real-time PCR and Western blot analysis. Chemically synthesized small interfering RNA against EMSP inhibited the expression of EMSP mRNA and significantly reduced the encystation efficiency of Acanthamoeba. An EMSP-enhanced green fluorescent protein fusion protein localized to vesicle-like structures within the amoeba. Using LysoTracker analysis, these vesicular structures were confirmed to be lysosomes. After incubation of the transfected amoeba in encystment media, small fluorescent vesicle-like structures gathered and formed ball-like structures, which were identified as colocalizing with the autophagosome. Taken together, these results indicate that EMSP plays an important role in the differentiation of Acanthamoeba by promoting autolysis.

https://ec.asm.org/content/eukcell/7/9/1513.full.pdf

Characterization of a serine proteinase mediating encystation of Acanthamoeba.

Autophagy protein 8 mediating autophagosome in encysting Acanthamoeba

Twelve isolates of Acanthamoeba spp. assigned to either A. castellanii or A. polyphaga, and type strains of A. culbertsoni, A. healyi, A. palestinensis, and A. astronyxis were examined by restriction fragment length polymorphism (RFLP) of a conserved region of small subunit ribosomal RNA gene (ssu rDNA) amplified by polymerase chain reaction (PCR). The PCR products of the isolates measured approximately 910-930 bp, except for that of A. astronyxis which was extraordinarily long, approximately 1,170 bp. Average of estimated sequence divergence of the amplified DNA among the isolates assigned to A. castellaii was 9.8% whereas that among the isolates assigned to A. polyphaga 9.6%. The maximum intraspecific sequence divergence among the isolates assigned to A. castellanii was observed between the Chang and Ma strains (17.3%) while that among the isolates assigned to A. polyphaga was observed between KA/S3 and KA/S7 strains (16.1%). The both maximum sequence divergences were much greater than the minimum interspecific sequence divergence between A. castellanii and A. polyphaga (2.6%) which appeared between the Castellani (or CCAP 1501/2 g) and KA/S3 strains. The PCR-RFLP patterns of A. culbertsoni, A. healyi, A. palestinensis, and A. astronyxis were quite diverse from one another and from those of isolates assigned to either A. castellanii or A. polyphaga. It is suggested that taxonomic validity of the isolates assigned to either A. castellanii or A. polyphaga should be reevaluated.

/pdf/pcr-and-rflp-variation-of-conserved-region-of-small-subunit-4idl5k3o6a.pdf

PCR and RFLP variation of conserved region of small subunit ribosomal DNA among Acanthamoeba isolates assigned to either A. castellanii or A. polyphaga

Hyun-Hee Kong

Papers

The first human case of Clinostomum complanatum (Trematoda: Clinostomidae) infection in Korea

Purification and characterization of a secretory serine proteinase of acanthamoeba healyi isolated from gae

Characterization of a serine proteinase mediating encystation of Acanthamoeba.

Autophagy protein 8 mediating autophagosome in encysting Acanthamoeba

PCR and RFLP variation of conserved region of small subunit ribosomal DNA among Acanthamoeba isolates assigned to either A. castellanii or A. polyphaga