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Sample records for acanthamoeba keratitis ak

  1. Genotypic Identification of Acanthamoeba sp. Isolates Associated with an Outbreak of Acanthamoeba keratitis (AK)

    PubMed Central

    Booton, Gregory C.; Joslin, Charlotte E.; Shoff, Megan; Tu, Elmer Y.; Kelly, Daryl J.; Fuerst, Paul A.

    2009-01-01

    Purpose Significant increases in Acanthamoeba keratitis (AK) cases have been observed in the Chicago-Gary-Kenosha area since June, 2003. It was hypothesized that increased rates of AK infections may be due to changes resulting from changes in municipal water treatment, or, alternatively, a more pathogenic strain of Acanthamoeba may be responsible. Methods Previous sequence analysis of the 18S rDNA of Acanthamoeba isolates resulted in the identification of 15 different genotypic classes. These analyses indicate AK cases are predominantly associated (~97%) with a single genotype (designated T4) of Acanthamoeba, and rarely with other genotypes (eg., T3 and T11). In this study we test the hypothesis that a new or more pathogenic genotype of Acanthamoeba is the cause of the recent surge in AK. Results We determined the genotype of 15 Acanthamoeba sp. isolates from AK cases from this outbreak using sequence analysis of a region of the 18S rDNA. Our results indicate that these isolates are predominantly genotype T4 (87%), with the remaining isolates being genotype T3 (13%). Both genotypes have previously been observed in AK cases. Conclusion There is no support for the hypothesis that the current AK outbreak is associated with infection by a new, more pathogenic, Acanthamoeba genotype. In addition, these results offer support for the hypothesis that the increased AK incidence may be due to changes in water treatment protocols leading to increased bacterial colonization of the water supply, and subsequent increases of already present Acanthamoeba sp, ultimately culminating in an increase of AK cases. PMID:19512903

  2. Acanthamoeba keratitis.

    PubMed

    Pogson, C

    1993-01-01

    1. Acanthamoeba keratitis is an uncommon but increasingly prevalent infection with the potential to cause severe ocular damage. Acanthamoeba is a nonflagellated free-living amoeba that is ubiquitous in the environment. The most common type is A castellani, but A polyphagia, A rhysodes, A culbertson, and A hatchetti have been isolated from infected eyes. 2. Clinical features include foreign body sensation, blurred vision, tearing, and photophobia. There is minimal pain in the early stages of infection, but severe pain is a manifestation of the advanced stages of the disease. 3. The incidence of Acanthamoeba keratitis appears to be decreasing because of increased awareness, but education emphasizing proper lens sterilization is essential and should be carried out when contact lenses are first dispensed. PMID:8158668

  3. [Painless acanthamoeba keratitis].

    PubMed

    Stemberger, K; Dick, B; Kramann, C; Thieme, H; Weber, A; Petry, F; Pfeiffer, N

    2007-05-01

    A 37-year-old contact lens wearer was treated for herpes simplex keratitis. After an initial improvement the keratitis became much worse. An annular infiltrate gave rise to the suspicion of acanthamoeba keratitis even though the patient was not in pain. This diagnosis was confirmed by histological and microbiological examination of the corneal disc after keratoplasty. Acanthamoeba keratitis should be considered even in the absence of pain, especially when the patients are contact lens wearers.

  4. Pathogenesis of Acanthamoeba Keratitis

    PubMed Central

    Panjwani, Noorjahan

    2010-01-01

    Acanthamoeba keratitis (AK) is a serious infection of the cornea. At present, diagnosis of the disease is not straightforward and treatment is very demanding. While contact lens wear is the leading risk factor for AK, Acanthamoeba parasites are increasingly recognized as an important cause of keratitis in non-contact lens wearers. The first critical step in the pathogenesis of infection is the adhesion of the microbe to the surface of the host tissues. Acanthamoebae express a major virulence protein, the mannose-binding protein (MBP), which mediates the adhesion of amoebae to the surface of the cornea. The MBP is a transmembrane protein with characteristics of a typical cell surface receptor. Subsequent to the MBP-mediated adhesion to host cells, the amoebae produce a contact-dependent metalloproteinase and several contact-independent serine proteinases. These proteinases work in concert to produce a potent cytopathic effect (CPE) involving killing of the host cells, degradation of epithelial basement membrane and underlying stromal matrix, and penetration into the deeper layers of the cornea. In the hamster animal model, oral immunization with the recombinant MBP protects against AK, and this protection is associated with an increased level of anti-MBP IgA in tears of protected animals. Normal human tear fluid contains IgA antibodies against Acanthamoeba MBP that is likely to provide protection by inhibiting the adhesion of parasites to host cells. Indeed, in in vitro CPE assays, even a low concentration of tears (10 [MU]μL of undiluted tears per milliliter of media) almost completely inhibits Acanthamoeba-induced CPE. In addition to adherence-inhibiting, IgA-mediated protection, human tears also contain IgA-independent factors that provide protection against Acanthamoeba-induced CPE by inhibiting the activity of cytotoxic proteinases. Characterization of the CPE-inhibitory factors of human tears should lead to a better understanding of the mechanism by which the

  5. Medical interventions for acanthamoeba keratitis

    PubMed Central

    Alkharashi, Majed; Lindsley, Kristina; Law, Hua Andrew; Sikder, Shameema

    2016-01-01

    Background Acanthamoeba are microscopic, free-living, single-celled organisms which can infect the eye and lead to Acanthamoeba keratitis (AK). AK can result in loss of vision in the infected eye or loss of eye itself; however, there are no formal guidelines or standards of care for the treatment of AK. Objectives To evaluate the relative effectiveness and safety of medical therapy for the treatment of AK. Search methods We searched CENTRAL (which contains the Cochrane Eyes and Vision Group Trials Register) (2015, Issue 1), Ovid MEDLINE, Ovid MEDLINE In-Process and Other Non-Indexed Citations, Ovid MEDLINE Daily, Ovid OLDMEDLINE (January 1946 to January 2015), EMBASE (January 1980 to January 2015), PubMed (1948 to January 2015), Latin American and Caribbean Health Sciences Literature Database (LILACS) (1982 to January 2015), the metaRegister of Controlled Trials (mRCT) (www.controlled-trials.com), ClinicalTrials.gov (www.clinicaltrials.gov) and the World Health Organization (WHO) International Clinical Trials Registry Platform (ICTRP) (www.who.int/ictrp/search/en). We did not use any date or language restrictions in the electronic search for trials. We last searched the electronic databases on 9 January 2015. Selection criteria We included randomized controlled trials (RCTs) of medical therapy for AK, regardless of the participants' age, sex, or etiology of disease. We included studies that compared either anti-amoeba therapy (drugs used alone or in combination with other medical therapies) with no anti-amoeba therapy or one anti-amoeba therapy with another anti-amoeba therapy. Data collection and analysis Two authors independently screened search results and full-text reports, assessed risk of bias, and abstracted data. We used standard methodological procedures as set forth by the Cochrane Collaboration. Main results We included one RCT (56 eyes of 55 participants) in this review. The study compared two types of topical biguanides for the treatment of AK

  6. IL-17A-mediated protection against Acanthamoeba keratitis.

    PubMed

    Suryawanshi, Amol; Cao, Zhiyi; Sampson, James F; Panjwani, Noorjahan

    2015-01-15

    Acanthamoeba keratitis (AK) is a very painful and vision-impairing infection of the cornea that is difficult to treat. Although past studies have indicated a critical role of neutrophils and macrophages in AK, the relative contribution of the proinflammatory cytokine, IL-17A, that is essential for migration, activation, and function of these cells into the cornea is poorly defined. Moreover, the role of the adaptive immune response, particularly the contribution of CD4(+) T cell subsets, Th17 and regulatory T cells , in AK is yet to be understood. In this report, using a mouse corneal intrastromal injection-induced AK model, we show that Acanthamoeba infection induces a strong CD4(+) T effector and regulatory T cell response in the cornea and local draining lymph nodes. We also demonstrate that corneal Acanthamoeba infection induces IL-17A expression and that IL-17A is critical for host protection against severe AK pathology. Accordingly, IL-17A neutralization in Acanthamoeba-infected wild-type mice or Acanthamoeba infection of mice lacking IL-17A resulted in a significantly increased corneal AK pathology, increased migration of inflammatory cells at the site of inflammation, and a significant increase in the effector CD4(+) T cell response in draining lymph nodes. Thus, in sharp contrast with other corneal infections such as herpes and Pseudomonas aeruginosa keratitis where IL-17A exacerbates corneal pathology and inflammation, the findings presented in this article suggest that IL-17A production after Acanthamoeba infection plays an important role in host protection against invading parasites.

  7. Influence of Acanthamoeba Genotype on Clinical Course and Outcomes for Patients with Acanthamoeba Keratitis in Spain

    PubMed Central

    Lumbreras-Fernández, Blanca; Martín-Navarro, Carmen M.; Valladares, Basilio; Lopez-Velez, Rogelio; Morcillo-Laiz, Rafael; Lorenzo-Morales, Jacob

    2014-01-01

    Genotype T4 is by far the most frequent genotype of Acanthamoeba keratitis (AK) and therefore has been considered the most virulent. This study included 14 cases of AK of genotype T4 and three cases of non-T4 genotype. We found that cases of non-T4 genotype had a worse response to medical therapy, greater need for surgical intervention, greater risk of extracorneal involvement, and remarkably poorer final visual outcome than those of T4 genotype, suggesting an association between Acanthamoeba virulence and genotype that requires additional case investigation. PMID:24430449

  8. Acanthamoeba keratitis challenges a case report.

    PubMed

    Cristina, Stan; Cristina, Vlăduţiu; Mihaela, Popovici

    2016-01-01

    Acanthamoeba keratitis is a rare, chronic, mainly contact lens-related infection caused by a free-living amoeba found ubiquitously in water and soil. A case of a 9-year-old child, who presented to our clinic with painful, red left eye, associated with photophobia, and decreased visual acuity, wais reported. The clinical examination revealed a discoid opacity inferiorly bounded by a dense, gray infiltrate. The progressive nature of the corneal infiltrate, the epithelial defect, and the lack of response to treatment was highly suggestive for Acanthamoeba keratitis. The distinctiveness of this case was the presence of Acanthamoeba keratitis in a child without a history of trauma or contact lens usage, the lack of an appropriate diagnosis and management of this vision-threatening infection. PMID:27220232

  9. Genotypic characterization of amoeba isolated from Acanthamoeba keratitis in Poland.

    PubMed

    Derda, Monika; Solarczyk, Piotr; Cholewiński, Marcin; Hadaś, Edward

    2015-03-01

    Free-living amoebae belonging to the genus Acanthamoeba are the causative factor of many diseases. Among others, they cause Acanthamoeba keratitis (AK), a condition that usually occurs in contact lens wearers, though it is also observed in non-wearers. The number of diagnosed cases of AK increased more than eightfold during 8 years in the USA, and a proportional increase in frequency also occurred in Poland and Europe. Cases of AK are usually diagnosed late, and their therapy is difficult and rarely successful. AK is an uncommon diagnosis in Poland. The increased number of positive cases observed in our laboratory may reflect the growing at-risk population of contact lens wearers. Acanthamoeba as a genus of facultative human parasites is currently classified into 17 genotypes. Isolates belonging to seven genotypes were found to be associated with AK. One genotype in particular, T4, was found to be overrepresented in human disease. The main finding of our study is that in Poland, AK is almost always associated with the T4 genotype.

  10. Development of an immunochromatographic assay kit using fluorescent silica nanoparticles for rapid diagnosis of Acanthamoeba keratitis.

    PubMed

    Toriyama, Koji; Suzuki, Takashi; Inoue, Tomoyuki; Eguchi, Hiroshi; Hoshi, Saichi; Inoue, Yoshitsugu; Aizawa, Hideki; Miyoshi, Kazutomi; Ohkubo, Michio; Hiwatashi, Eiji; Tachibana, Hiroshi; Ohashi, Yuichi

    2015-01-01

    We developed an immunochromatographic assay kit that uses fluorescent silica nanoparticles bound to anti-Acanthamoeba antibodies (fluorescent immunochromatographic assay [FICGA]) and evaluated its efficacy for the detection of Acanthamoeba and diagnosis of Acanthamoeba keratitis (AK). The sensitivity of the FICGA kit was evaluated using samples of Acanthamoeba trophozoites and cysts diluted to various concentrations. A conventional immunochromatographic assay kit with latex labels (LICGA) was also evaluated to determine its sensitivity in detecting Acanthamoeba trophozoites. To check for cross-reactivity, the FICGA was performed by using samples of other common causative pathogens of infectious keratitis, such as Pseudomonas aeruginosa, Staphylococcus aureus, Staphylococcus epidermidis, and Candida albicans. Corneal scrapings from patients with suspected AK were tested with the FICGA kit to detect the presence of Acanthamoeba, and the results were compared with those of real-time PCR. The FICGA kit detected organisms at concentrations as low as 5 trophozoites or 40 cysts per sample. There were no cross-reactivities with other pathogens. The FICGA was approximately 20 times more sensitive than the LICGA for the detection of Acanthamoeba trophozoites. The FICGA kit yielded positive results for all 10 patients, which corresponded well with the real-time PCR results. The FICGA kit demonstrated high sensitivity for the detection of Acanthamoeba and may be useful for the diagnosis of AK.

  11. Comparison of Fluorescence Microscopy and Different Growth Media Culture Methods for Acanthamoeba Keratitis Diagnosis

    PubMed Central

    Peretz, Avi; Geffen, Yuval; Socea, Soergiu D.; Pastukh, Nina; Graffi, Shmuel

    2015-01-01

    Acanthamoeba keratitis (AK), a potentially blinding infection of the cornea, is caused by a free-living protozoan. Culture and microscopic examination of corneal scraping tissue material is the conventional method for identifying Acanthamoeba. In this article, we compared several methods for AK diagnosis of 32 patients: microscopic examination using fluorescent dye, specific culture on growth media—non-nutrient agar (NNA), culture on liquid growth media—peptone yeast glucose (PYG), and TYI-S-33. AK was found in 14 patients. Thirteen of the specimens were found AK positive by fluorescence microscopic examination, 11 specimens were found AK positive on PYG growth media, and 9 specimens were found AK positive on TYI-S-33 growth media. Only five specimens were found AK positive on NNA growth media. Therefore, we recommend using fluorescence microscopy technique and culture method, especially PYG liquid media. PMID:25962772

  12. Comparison of Fluorescence Microscopy and Different Growth Media Culture Methods for Acanthamoeba Keratitis Diagnosis.

    PubMed

    Peretz, Avi; Geffen, Yuval; Socea, Soergiu D; Pastukh, Nina; Graffi, Shmuel

    2015-08-01

    Acanthamoeba keratitis (AK), a potentially blinding infection of the cornea, is caused by a free-living protozoan. Culture and microscopic examination of corneal scraping tissue material is the conventional method for identifying Acanthamoeba. In this article, we compared several methods for AK diagnosis of 32 patients: microscopic examination using fluorescent dye, specific culture on growth media-non-nutrient agar (NNA), culture on liquid growth media-peptone yeast glucose (PYG), and TYI-S-33. AK was found in 14 patients. Thirteen of the specimens were found AK positive by fluorescence microscopic examination, 11 specimens were found AK positive on PYG growth media, and 9 specimens were found AK positive on TYI-S-33 growth media. Only five specimens were found AK positive on NNA growth media. Therefore, we recommend using fluorescence microscopy technique and culture method, especially PYG liquid media.

  13. The Value of Cytology Smears for Acanthamoeba Keratitis

    PubMed Central

    Schaefer, Jamie L.; Paterson, Joyce; Liu, Weiguo; Gonzalez-Fernandez, Federico

    2016-01-01

    Purpose. Acanthamoeba keratitis remains a difficult diagnosis despite advances in genetic and imaging technologies. The purpose of this paper is to highlight the utility of cytology smears for diagnosis of Acanthamoeba keratitis. Methods. This is a case study of the diagnostic course for a patient with suspected Acanthamoeba keratitis. Results. A 40-year-old male with poor contact lens hygiene presented with severe left eye pain. Slit lamp examination showed two peripheral ring infiltrates without an epithelial defect. The epithelium over both infiltrates was removed with a Kimura spatula. Half of the sample was smeared on a dry microscope slide and the other half was submitted for Acanthamoeba culture and PCR. Both culture and PCR were negative for Acanthamoeba, but hematoxylin and eosin stain of the smear revealed double-walled cysts. Conclusion. H&E staining of corneal cytology specimens is an efficient and readily available test for diagnosis of Acanthamoeba keratitis. PMID:27403362

  14. First report of an Acanthamoeba genotype T13 isolate as etiological agent of a keratitis in humans.

    PubMed

    Grün, Anna-Lena; Stemplewitz, Birthe; Scheid, Patrick

    2014-06-01

    Several strains of free-living amoebae (FLA) belonging to the genus Acanthamoeba are able to cause a painful sight-threatening disease of the cornea designated as Acanthamoeba keratitis (AK). In this case report, a 22-year-old woman, wearer of soft contact lenses, was treated after the initial examination, and follow-up laboratory results led to the diagnosis of Acanthamoeba keratitis. The patient recovered under the targeted therapy, demonstrating that the acanthamoebae were the etiological agents of the keratitis in this case. The acanthamoebae belonged morphologically to group II. Genotyping of the causative Acanthamoeba strain based on sequences of the PCR amplimer ASA.S1 amplified from 18S ribosomal DNA by using the genus-specific primers JDP1 and JDP2 followed. The phylogenetic comparison of ASA.S1 confirmed that the isolated Acanthamoeba strain is closely related to genotype T13 supported by pairwise sequence identities of 97.1-98.0% and bootstrap support of 980 replicates with reference sequences of genotype T13. These results regarding the Acanthamoeba keratitis-causing isolate KaBo expands the number of known pathogenic genotypes to 12. To our knowledge, this is the first report of a T13 Acanthamoeba genotype being associated with keratitis in humans. PMID:24802868

  15. Acanthamoeba genotypes T3 and T4 as causative agents of amoebic keratitis in Mexico.

    PubMed

    Omaña-Molina, Maritza; Vanzzini-Zago, Virginia; Hernandez-Martinez, Dolores; Gonzalez-Robles, Arturo; Salazar-Villatoro, Lizbeth; Ramirez-Flores, Elizabeth; Oregon-Miranda, Eric; Lorenzo-Morales, Jacob; Martinez-Palomo, Adolfo

    2016-02-01

    Free-living amoebae (FLA) are widely distributed worldwide. Some genera included in this group act as opportunistic pathogens causing fatal encephalitis and Acanthamoeba keratitis (AK), a sight-threatening infection of the cornea associated with the use of soft contact lenses that could even end in blindness if an early diagnosis and treatment are not achieved. Furthermore, the numbers of AK cases keep rising worldwide mainly due to an increase of contact lens wearers and lack of hygiene in the maintenance of lenses and their cases. In Mexico, no cases of AK have been described so far although the isolation of other pathogenic FLA such as Naegleria fowleri and Balamuthia mandrillaris from both clinical and environmental sources has been reported. The present study reports two cases of Acanthamoeba keratitis diagnosed in two patients admitted to the Hospital "Luis Sánchez Bulnes" for Blindness Prevention in Mexico City, Mexico. Corneal scrapes and contact lenses were checked for the presence of Acanthamoeba strains in both patients. Strains were axenized after initial isolation to classify at the genotype level. After sequencing the diagnostic fragment 3 (DF3) region located on the 18S ribosomal DNA (rDNA) gene of Acanthamoeba, genotype T3 and genotype T4 were identified in clinical case 1 and 2, respectively. To our knowledge, these are the first reported cases of AK in Mexico in the literature and the first description of Acanthamoeba genotypes T3 and T4 as causative agents of amoebic infection. PMID:26581373

  16. Advances in the Diagnosis and Treatment of Acanthamoeba Keratitis

    PubMed Central

    Clarke, Benjamin; Sinha, Arti; Parmar, Dipak N.

    2012-01-01

    This paper aims to review the recent literature describing Acanthamoeba keratitis and outline current thoughts on pathogenesis, diagnosis, and treatment as well as currently emerging diagnostic and treatment modalities. PMID:23304449

  17. The role of domestic tap water on Acanthamoeba keratitis in non-contact lens wearers and validation of laboratory methods.

    PubMed

    Koltas, Ismail Soner; Eroglu, Fadime; Erdem, Elif; Yagmur, Meltem; Tanır, Ferdi

    2015-09-01

    Acanthamoeba is increasingly recognized as an important cause of keratitis in non-contact lens wearers while contact lens wear is the leading risk factor for Acanthamoeba keratitis (AK). It is unlikely that the Acanthamoeba colonization is a feature which is effective only in patient's homes with infectious keratitis since the organism has been isolated from domestic tap water. Two hundred and thirty-one (231) corneal scrapings were taken from infectious keratitis cases, and four contact lens solutions and domestic tap waters were taken from 22 out of 44 AK-diagnosed patient's homes. Microscopic examination, culture, PCR, real-time PCR and DNA sequencing analyses were used for AK-diagnosed samples. The real-time PCR was the most sensitive (100 %) one among the methods used in diagnosis of AK. The 44 (19.0 %) out of 231 corneal scrapings, 4/4 (100 %) contact lens solution and 11/22 (50 %) of domestic tap water samples were found to be positive by real-time PCR for Acanthamoeba. A. griffini (T3), A. castellanii (T4) and A. jacobsi (T15) genotypes were obtained from corneal scrapings, contact lens solutions and domestic tap water samples taken from the patient's homes diagnosed with AK. The isolation of Acanthamoeba containing 6/22 (27.3 %) A. griffini (T3), 14/22 (63.6 %) A. castellanii (T4) and 2/22 (9.1 %) A. jacobsi (T15) from the domestic tap water outlets of 22 of 44 (50 %) of patient's homes revealed that is a significant source of these organisms. A. griffini (T3) and A. jacobsi (T15) genotypes have not been determined from AK cases in Turkey previously. Thus, we conclude that Acanthamoeba keratitis is associated with exposition of patients who has ocular trauma or ocular surface disease to domestic tap water in endemic or potentially endemic countries. PMID:26017346

  18. The role of domestic tap water on Acanthamoeba keratitis in non-contact lens wearers and validation of laboratory methods.

    PubMed

    Koltas, Ismail Soner; Eroglu, Fadime; Erdem, Elif; Yagmur, Meltem; Tanır, Ferdi

    2015-09-01

    Acanthamoeba is increasingly recognized as an important cause of keratitis in non-contact lens wearers while contact lens wear is the leading risk factor for Acanthamoeba keratitis (AK). It is unlikely that the Acanthamoeba colonization is a feature which is effective only in patient's homes with infectious keratitis since the organism has been isolated from domestic tap water. Two hundred and thirty-one (231) corneal scrapings were taken from infectious keratitis cases, and four contact lens solutions and domestic tap waters were taken from 22 out of 44 AK-diagnosed patient's homes. Microscopic examination, culture, PCR, real-time PCR and DNA sequencing analyses were used for AK-diagnosed samples. The real-time PCR was the most sensitive (100 %) one among the methods used in diagnosis of AK. The 44 (19.0 %) out of 231 corneal scrapings, 4/4 (100 %) contact lens solution and 11/22 (50 %) of domestic tap water samples were found to be positive by real-time PCR for Acanthamoeba. A. griffini (T3), A. castellanii (T4) and A. jacobsi (T15) genotypes were obtained from corneal scrapings, contact lens solutions and domestic tap water samples taken from the patient's homes diagnosed with AK. The isolation of Acanthamoeba containing 6/22 (27.3 %) A. griffini (T3), 14/22 (63.6 %) A. castellanii (T4) and 2/22 (9.1 %) A. jacobsi (T15) from the domestic tap water outlets of 22 of 44 (50 %) of patient's homes revealed that is a significant source of these organisms. A. griffini (T3) and A. jacobsi (T15) genotypes have not been determined from AK cases in Turkey previously. Thus, we conclude that Acanthamoeba keratitis is associated with exposition of patients who has ocular trauma or ocular surface disease to domestic tap water in endemic or potentially endemic countries.

  19. An update on Acanthamoeba keratitis: diagnosis, pathogenesis and treatment

    PubMed Central

    Lorenzo-Morales, Jacob; Khan, Naveed A.; Walochnik, Julia

    2015-01-01

    Free-living amoebae of the genus Acanthamoeba are causal agents of a severe sight-threatening infection of the cornea known as Acanthamoeba keratitis. Moreover, the number of reported cases worldwide is increasing year after year, mostly in contact lens wearers, although cases have also been reported in non-contact lens wearers. Interestingly, Acanthamoeba keratitis has remained significant, despite our advances in antimicrobial chemotherapy and supportive care. In part, this is due to an incomplete understanding of the pathogenesis and pathophysiology of the disease, diagnostic delays and problems associated with chemotherapeutic interventions. In view of the devastating nature of this disease, here we present our current understanding of Acanthamoeba keratitis and molecular mechanisms associated with the disease, as well as virulence traits of Acanthamoeba that may be potential targets for improved diagnosis, therapeutic interventions and/or for the development of preventative measures. Novel molecular approaches such as proteomics, RNAi and a consensus in the diagnostic approaches for a suspected case of Acanthamoeba keratitis are proposed and reviewed based on data which have been compiled after years of working on this amoebic organism using many different techniques and listening to many experts in this field at conferences, workshops and international meetings. Altogether, this review may serve as the milestone for developing an effective solution for the prevention, control and treatment of Acanthamoeba infections. PMID:25687209

  20. An update on Acanthamoeba keratitis: diagnosis, pathogenesis and treatment.

    PubMed

    Lorenzo-Morales, Jacob; Khan, Naveed A; Walochnik, Julia

    2015-01-01

    Free-living amoebae of the genus Acanthamoeba are causal agents of a severe sight-threatening infection of the cornea known as Acanthamoeba keratitis. Moreover, the number of reported cases worldwide is increasing year after year, mostly in contact lens wearers, although cases have also been reported in non-contact lens wearers. Interestingly, Acanthamoeba keratitis has remained significant, despite our advances in antimicrobial chemotherapy and supportive care. In part, this is due to an incomplete understanding of the pathogenesis and pathophysiology of the disease, diagnostic delays and problems associated with chemotherapeutic interventions. In view of the devastating nature of this disease, here we present our current understanding of Acanthamoeba keratitis and molecular mechanisms associated with the disease, as well as virulence traits of Acanthamoeba that may be potential targets for improved diagnosis, therapeutic interventions and/or for the development of preventative measures. Novel molecular approaches such as proteomics, RNAi and a consensus in the diagnostic approaches for a suspected case of Acanthamoeba keratitis are proposed and reviewed based on data which have been compiled after years of working on this amoebic organism using many different techniques and listening to many experts in this field at conferences, workshops and international meetings. Altogether, this review may serve as the milestone for developing an effective solution for the prevention, control and treatment of Acanthamoeba infections.

  1. A Rabbit Model of Acanthamoeba Keratitis That Better Reflects the Natural Human Infection.

    PubMed

    Feng, Xianmin; Zheng, Wenyu; Wang, Yuehua; Zhao, Donghai; Jiang, Xiaoming; Lv, Shijie

    2015-08-01

    Acanthamoeba species are ubiquitous, free-living protozoa that can invade the cornea and result in Acanthamoeba keratitis (AK), a painful progressive sight-threatening corneal disease. Disease progression in current animal models is too rapid to mimic AK in humans accurately. This study provides a novel method for establishing AK in rabbits and compared it with the conventional method with regard to pathogenesis and immune response in humans. The New Zealand white rabbits were randomly divided into two experimental groups (Groups A and B). Rabbits in the Group A (n = 14) received intrastromal injections of 1 × 10(4) /100 µL Acanthamoeba healyi trophozoites (conventional AK model). The Group B animals (n = 14) received microinjections of 1 × 10(4) /10 µL A. healyi trophozoites between the corneal epithelium and Bowman's layer, anterior to the corneal stroma (novel AK model). In addition, two rabbits were left untreated as normal controls. AK in the treated rabbits was evaluated clinically, histopathologically, and immunologically for 35 days. AK was successfully established in both the conventional and novel model groups. Compared with the Group A, AK in the Group B displayed an efficient immune response with less severe pathology. Moreover, the self-limiting but chronic nature of the infection in the Group B was strikingly similar to that of AK in humans. The novel animal model for AK described here more closely simulates the pathogenesis and immune response of Acanthamoeba corneal infection in humans than the animal models currently in use.

  2. Autophagy inhibitors as a potential antiamoebic treatment for Acanthamoeba keratitis.

    PubMed

    Moon, Eun-Kyung; Kim, So-Hee; Hong, Yeonchul; Chung, Dong-Il; Goo, Youn-Kyoung; Kong, Hyun-Hee

    2015-07-01

    Acanthamoeba cysts are resistant to extreme physical and chemical conditions. Autophagy is an essential pathway for encystation of Acanthamoeba cells. To evaluate the possibility of an autophagic Acanthamoeba encystation mechanism, we evaluated autophagy inhibitors, such as 3-methyladenine (3MA), LY294002, wortmannin, bafilomycin A, and chloroquine. Among these autophagy inhibitors, the use of 3MA and chloroquine showed a significant reduction in the encystation ratio in Acanthamoeba cells. Wortmannin also inhibited the formation of mature cysts, while LY294002 and bafilomycin A did not affect the encystation of Acanthamoeba cells. Transmission electron microscopy revealed that 3MA and wortmannin inhibited autophagy formation and that chloroquine interfered with the formation of autolysosomes. Inhibition of autophagy or autolysosome formation resulted in a significant block in the encystation in Acanthamoeba cells. Clinical treatment with 0.02% polyhexamethylene biguanide (PHMB) showed high cytopathic effects on Acanthamoeba trophozoites and cysts; however, it also revealed high cytopathic effects on human corneal epithelial cells. In this study, we investigated effects of the combination of a low (0.00125%) concentration of PHMB with each of the autophagy inhibitors 3MA, wortmannin, and chloroquine on Acanthamoeba and human corneal epithelial cells. These new combination treatments showed low cytopathic effects on human corneal cells and high cytopathic effects on Acanthamoeba cells. Taken together, these results provide fundamental information for optimizing the treatment of Acanthamoeba keratitis.

  3. Acanthamoeba, fungal, and bacterial keratitis: a comparison of risk factors and clinical features

    PubMed Central

    Mascarenhas, Jeena; Lalitha, Prajna; Prajna, N. Venkatesh; Srinivasan, Muthiah; Das, Manoranjan; D’Silva, Sean S.; Oldenburg, Catherine E.; Borkar, Durga S.; Esterberg, Elizabeth J.; Lietman, Thomas M.; Keenan, Jeremy D.

    2013-01-01

    Purpose To determine risk factors and clinical signs that may differentiate between bacterial, fungal, and acanthamoeba keratitis among patients presenting with presumed infectious keratitis. Design Hospital-based cross-sectional study. Methods We examined the medical records of 115 patients with laboratory-proven bacterial keratitis, 115 patients with laboratory-proven fungal keratitis, and 115 patients with laboratory-proven acanthamoeba keratitis seen at Aravind Eye Hospital, Madurai, India, from 2006–2011. Risk factors and clinical features of the three organisms were compared using multinomial logistic regression. Results Of 95 patients with bacterial keratitis, 103 patients with fungal keratitis, and 93 patients with acanthamoeba keratitis who had medical records available for review, 287 (99%) did not wear contact lenses. Differentiating features were more common for acanthamoeba keratitis than for bacterial or fungal keratitis. Compared to patients with bacterial or fungal keratitis, patients with acanthamoeba keratitis were more likely to be younger and to have a longer duration of symptoms, and to have a ring infiltrate or disease confined to the epithelium. Conclusions Risk factors and clinical examination findings can be useful for differentiating acanthamoeba keratitis from bacterial and fungal keratitis. PMID:24200232

  4. Raman Microspectroscopy Analysis in the Treatment of Acanthamoeba Keratitis

    PubMed Central

    Rusciano, Giulia; Capriglione, Paola; Pesce, Giuseppe; Del Prete, Salvatore; Cennamo, Gilda; Di Cave, David; Cerulli, Luciano; Sasso, Antonio

    2013-01-01

    Acanthamoeba keratitis is a rare but serious corneal disease, often observed in contact lens wearers. Clinical treatment of infected patients frequently involves the use of polyhexamethylene biguanide (PHMB), a polymer used as a disinfectant and antiseptic, which is toxic also for the epithelial cells of the cornea. Prompt and effective diagnostic tools are hence highly desiderable for both starting early therapy and timely suspension of the treatment. In this work we use Raman microspectroscopy to analyse in vitro a single Acanthamoeba cell in cystic phase. In particular, we investigate the effect of PHMB at the single-cell level, providing useful information on both the underlying biochemical mechanism and the time frame for Acanthamoeba eradication in ocular infections. Furthermore, we demonstrate that Raman spectroscopy, in conjunction with standard multivariate analysis methods, allows discriminating between live and dead Acanthamoebas, which is fundamental to optimizing patients’ treatment. PMID:23977228

  5. Reevaluation of an Acanthamoeba Molecular Diagnostic Algorithm following an Atypical Case of Amoebic Keratitis

    PubMed Central

    Lau, Rachel; Cunanan, Marlou; Jackson, Jonathan; Ali, Ibne Karim M.; Chong-Kit, Ann; Gasgas, Jason; Tian, Jinfang; Ralevski, Filip

    2015-01-01

    Amoebic keratitis (AK) is a potentially blinding infection, the prompt diagnosis of which is essential for limiting ocular morbidity. We undertook a quality improvement initiative with respect to the molecular detection of acanthamoebae in our laboratory because of an unusual case of discordance. Nine ATCC strains of Acanthamoeba and 40 delinked, biobanked, surplus corneal scraping specimens were analyzed for the presence of acanthamoebae with four separate real-time PCR assays. The assay used by the Free-Living and Intestinal Amebas Laboratory of the CDC was considered the reference standard, and the performance characteristics of each individual assay and pairs of assays were calculated. Outcome measures were sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV). Of 49 included specimens, 14 (28.6%) were positive by the gold standard assay, and 35 (71.4%) were negative. The sensitivities of the individual assays ranged from 64.3% to 92.9%, compared to the gold standard, while the specificities ranged from 88.6% to 91.4%. The PPVs and NPVs ranged from 69.2% to 78.6% and from 86.1% to 96.9%, respectively. Combinations of assay pairs led to improved performance, with sensitivities ranging from 92.9% to 100% and specificities ranging from 97.1% to 100%. ATCC and clinical strains of Acanthamoeba that failed to be detected by certain individual assays included Acanthamoeba castellanii, Acanthamoeba culbertsoni, and Acanthamoeba lenticulata. For three clinical specimens, false negativity of the gold standard assay could not be excluded. Molecular diagnostic approaches, especially combinations of highly sensitive and specific assays, offer a reasonably performing, operator-independent, rapid strategy for the detection of acanthamoebae in clinical specimens and are likely to be more practical than either culture or direct microscopic detection. PMID:26202123

  6. Reevaluation of an Acanthamoeba Molecular Diagnostic Algorithm following an Atypical Case of Amoebic Keratitis.

    PubMed

    Lau, Rachel; Cunanan, Marlou; Jackson, Jonathan; Ali, Ibne Karim M; Chong-Kit, Ann; Gasgas, Jason; Tian, Jinfang; Ralevski, Filip; Boggild, Andrea K

    2015-10-01

    Amoebic keratitis (AK) is a potentially blinding infection, the prompt diagnosis of which is essential for limiting ocular morbidity. We undertook a quality improvement initiative with respect to the molecular detection of acanthamoebae in our laboratory because of an unusual case of discordance. Nine ATCC strains of Acanthamoeba and 40 delinked, biobanked, surplus corneal scraping specimens were analyzed for the presence of acanthamoebae with four separate real-time PCR assays. The assay used by the Free-Living and Intestinal Amebas Laboratory of the CDC was considered the reference standard, and the performance characteristics of each individual assay and pairs of assays were calculated. Outcome measures were sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV). Of 49 included specimens, 14 (28.6%) were positive by the gold standard assay, and 35 (71.4%) were negative. The sensitivities of the individual assays ranged from 64.3% to 92.9%, compared to the gold standard, while the specificities ranged from 88.6% to 91.4%. The PPVs and NPVs ranged from 69.2% to 78.6% and from 86.1% to 96.9%, respectively. Combinations of assay pairs led to improved performance, with sensitivities ranging from 92.9% to 100% and specificities ranging from 97.1% to 100%. ATCC and clinical strains of Acanthamoeba that failed to be detected by certain individual assays included Acanthamoeba castellanii, Acanthamoeba culbertsoni, and Acanthamoeba lenticulata. For three clinical specimens, false negativity of the gold standard assay could not be excluded. Molecular diagnostic approaches, especially combinations of highly sensitive and specific assays, offer a reasonably performing, operator-independent, rapid strategy for the detection of acanthamoebae in clinical specimens and are likely to be more practical than either culture or direct microscopic detection.

  7. Identifying differentially expressed genes in trophozoites and cysts of Acanthamoeba T4 genotype: Implications for developing new treatments for Acanthamoeba keratitis.

    PubMed

    Abedkhojasteh, Hoda; Niyyati, Maryam; Rezaei, Sasan; Mohebali, Mehdi; Farnia, Shohreh; Kazemi-Rad, Elham; Roozafzoon, Reza; Sianati, Hamed; Rezaeian, Mostafa; Heidari, Mansour

    2015-02-01

    Acanthamoeba T4 genotype is the most prevalent genotype associated with amoebic keratitis. Acanthamoeba keratitis therapy is difficult due to transformation of trophozoite to cyst stage, which hinders the treatment of the disease. Although encystation assists the organism to survive against the chemotherapeutic compounds, the precise mechanism of encystation remains poorly understood. The purpose of this work was to identify differentially expressed genes in Acanthamoeba T4 genotype which might be useful for understanding of the encystment process and may thus help develop more efficient treatment. The mRNA profile of trophozoite and cyst of Acanthamoeba T4 genotype isolated from a soft contact lens wearer were analyzed using a cDNA amplified fragment length polymorphism (cDNA-AFLP) technique. Subsequently, a real time reverse transcriptase-PCR was performed to validate the cDNA-AFLP results. Three genes, heat shock protein70 (hsp70), actin-I and elongation factor-1alpha (EF-1α) were differentially expressed during Acanthamoeba differentiation. An in silico result predicted that transformation of trophozoite to cyst could be mediated through their cooperation with the protein partners interaction. Taken together, our experimental and bioinformatics findings suggested potential functions of hsp70, EF-1α and actin-I in differentiation of Acanthamoeba T4 genotype which may be useful in the design of an efficient therapeutic strategy in AK.

  8. Acanthamoeba keratitis: improving the Scottish diagnostic service for the rapid molecular detection of Acanthamoeba species.

    PubMed

    Alexander, Claire Low; Coyne, Michael; Jones, Brian; Anijeet, Deepa

    2015-07-01

    Acanthamoeba species are responsible for causing the potentially sight-threatening condition, Acanthamoeba keratitis, which is commonly associated with contact lens use. In this report, we highlight the challenges faced using conventional laboratory identification methods to identify this often under-reported pathogen, and discuss the reasons for introducing the first national service in Scotland for the rapid and sensitive molecular identification of Acanthamoeba species. By comparing culture and molecular testing data from a total of 63 patients (n = 80 samples) throughout Scotland presenting with ocular eye disease, we describe the improvement in detection rates where an additional four positive cases were identified using a molecular assay versus culture. The testing of a further ten patients by confocal imaging is also presented. This report emphasizes the importance of continuing to improve clinical laboratory services to ensure a prompt, correct diagnosis and better prognosis, in addition to raising awareness of this potentially debilitating opportunistic pathogen.

  9. Risk factors for acanthamoeba keratitis in contact lens users: a case-control study.

    PubMed Central

    Radford, C. F.; Bacon, A. S.; Dart, J. K.; Minassian, D. C.

    1995-01-01

    OBJECTIVE--To investigate reasons for an increase in cases of Acanthamoeba keratitis related to contact lenses. DESIGN--Case-control study. Cases were contact lens related acanthamoeba keratitis patients treated between 1 September 1989 and 31 August 1992. Controls were lens users without lens related disease who presented as new patients to the casualty department from 1 March 1992 to 31 August 1992. All subjects completed a questionnaire detailing lens use and hygiene practices. SETTING--Eye hospital. SUBJECTS--35 cases with acanthamoeba keratitis and 378 controls. MAIN OUTCOME MEASURES--Relative risks comparing different contact lens types, socioeconomic classification, age, sex, lens use, lens wearing experience, hygiene compliance, and hygiene systems. RESULTS--The crude relative risk for developing acanthamoeba keratitis with the use of daily wear disposable lenses was 49.45 (95% confidence interval 6.53 to 2227; P < 0.001) compared with conventional soft lenses (the referent). Multivariable analysis showed that this increased risk could be largely attributed to lack of disinfection (relative risk 55.86 (10 to 302); P < 0.001) and use of chlorine based disinfection (14.63 (2.8 to 76); P = 0.001) compared with other chemical systems (the referent). None of the other outcome measures showed a significant association. CONCLUSIONS--Both failure to disinfect daily wear soft contact lenses and the use of chlorine release lens disinfection systems, which have little protective effect against the organism, are major risk factors for acanthamoeba keratitis. These risks have been particularly common in disposable lens use. Over 80% of acanthamoeba keratitis could be avoided by the use of lens disinfection systems that are effective against the organism. PMID:7787645

  10. Emerging Threats for Human Health in Poland: Pathogenic Isolates from Drug Resistant Acanthamoeba Keratitis Monitored in terms of Their In Vitro Dynamics and Temperature Adaptability

    PubMed Central

    Chomicz, Lidia; Conn, David Bruce; Padzik, Marcin; Szaflik, Jacek P.; Walochnik, Julia; Zawadzki, Paweł J.; Pawłowski, Witold; Dybicz, Monika

    2015-01-01

    Amphizoic amoebae generate a serious human health threat due to their pathogenic potential as facultative parasites, causative agents of vision-threatening Acanthamoeba keratitis (AK). Recently, AK incidences have been reported with increasing frequency worldwide, particularly in contact lens wearers. In our study, severe cases of AK in Poland and respective pathogenic isolates were assessed at clinical, morphological, and molecular levels. Misdiagnoses and the unsuccessful treatment in other ophthalmic units delayed suitable therapy, and resistance to applied chemicals resulted in severe courses and treatment difficulties. Molecular assessment indicated that all sequenced pathogenic corneal isolates deriving from Polish patients with AK examined by us showed 98–100% homology with Acanthamoeba genotype T4, the most prevalent genotype in this human ocular infection worldwide. In vitro assays revealed that the pathogenic strains are able to grow at elevated temperature and have a wide adaptive capability. This study is our subsequent in vitro investigation on pathogenic Acanthamoeba strains of AK originating from Polish patients. Further investigations designed to foster a better understanding of the factors leading to an increase of AK observed in the past years in Poland may help to prevent or at least better cope with future cases. PMID:26682216

  11. Failure of chemotherapy in the first reported cases of Acanthamoeba keratitis in Pakistan.

    PubMed

    Siddiqui, Ruqaiyyah; Chaudhry, Tanveer; Lakhundi, Sahreena; Ahmad, Khabir; Khan, Naveed Ahmed

    2014-01-01

    Acanthamoeba keratitis is a painful and progressive infection of the cornea that can result in loss of vision. Here, for the first time in Pakistan, we report two cases of Acanthamoeba keratitis. The first patient was a 37-year-old female who presented with severe itching, redness, pain, along with loss of vision. The patient was a regular soft contact lens wearer. The second patient was a 25-year-old female who had been using soft contact lenses for the past two years. She presented with a burning sensation and extreme pain, along with loss of vision. Both patients were treated for a possible microbial keratitis with topical moxifloxacin hydrochloride drops, vancomycin drops, propamidine isethionate ointment, amphotericin B drops, and amikacin drops. However, the response was inadequate and both patients were referred for corneal transplant. Acanthamoeba castellanii was isolated by placing contact lenses and contact lens cases on non-nutrient agar plates containing a lawn of non-invasive Escherichia coli K-12 HB101 bacteria. The polymerase chain reaction (PCR) using genus-specific probes confirmed the identity of Acanthamoeba spp., whereas the morphological characteristics of trophozoites and cysts were suggestive of A. castellanii in both cases. With growing use of contact lenses for vision correction/cosmetic use coupled with sub-standard lens care in this region and the possibility of non-contact lens-associated Acanthamoeba keratitis, a need for increased awareness of this sight-threatening infection is discussed further. PMID:24548160

  12. Failure of chemotherapy in the first reported cases of Acanthamoeba keratitis in Pakistan

    PubMed Central

    Siddiqui, Ruqaiyyah; Chaudhry, Tanveer; Lakhundi, Sahreena; Ahmad, Khabir; Khan, Naveed Ahmed

    2014-01-01

    Acanthamoeba keratitis is a painful and progressive infection of the cornea that can result in loss of vision. Here, for the first time in Pakistan, we report two cases of Acanthamoeba keratitis. The first patient was a 37-year-old female who presented with severe itching, redness, pain, along with loss of vision. The patient was a regular soft contact lens wearer. The second patient was a 25-year-old female who had been using soft contact lenses for the past two years. She presented with a burning sensation and extreme pain, along with loss of vision. Both patients were treated for a possible microbial keratitis with topical moxifloxacin hydrochloride drops, vancomycin drops, propamidine isethionate ointment, amphotericin B drops, and amikacin drops. However, the response was inadequate and both patients were referred for corneal transplant. Acanthamoeba castellanii was isolated by placing contact lenses and contact lens cases on non-nutrient agar plates containing a lawn of non-invasive Escherichia coli K-12 HB101 bacteria. The polymerase chain reaction (PCR) using genus-specific probes confirmed the identity of Acanthamoeba spp., whereas the morphological characteristics of trophozoites and cysts were suggestive of A. castellanii in both cases. With growing use of contact lenses for vision correction/cosmetic use coupled with sub-standard lens care in this region and the possibility of non-contact lens-associated Acanthamoeba keratitis, a need for increased awareness of this sight-threatening infection is discussed further. PMID:24548160

  13. Use of Subgenic 18S Ribosomal DNA PCR and Sequencing for Genus and Genotype Identification of Acanthamoebae from Humans with Keratitis and from Sewage Sludge

    PubMed Central

    Schroeder, Jill M.; Booton, Gregory C.; Hay, John; Niszl, Ingrid A.; Seal, David V.; Markus, Miles B.; Fuerst, Paul A.; Byers, Thomas J.

    2001-01-01

    This study identified subgenic PCR amplimers from 18S rDNA that were (i) highly specific for the genus Acanthamoeba, (ii) obtainable from all known genotypes, and (iii) useful for identification of individual genotypes. A 423- to 551-bp Acanthamoeba-specific amplimer ASA.S1 obtained with primers JDP1 and JDP2 was the most reliable for purposes i and ii. A variable region within this amplimer also identified genotype clusters, but purpose iii was best achieved with sequencing of the genotype-specific amplimer GTSA.B1. Because this amplimer could be obtained from any eukaryote, axenic Acanthamoeba cultures were required for its study. GTSA.B1, produced with primers CRN5 and 1137, extended between reference bp 1 and 1475. Genotypic identification relied on three segments: bp 178 to 355, 705 to 926, and 1175 to 1379. ASA.S1 was obtained from single amoeba, from cultures of all known 18S rDNA genotypes, and from corneal scrapings of Scottish patients with suspected Acanthamoeba keratitis (AK). The AK PCR findings were consistent with culture results for 11 of 15 culture-positive specimens and detected Acanthamoeba in one of nine culture-negative specimens. ASA.S1 sequences were examined for 6 of the 11 culture-positive isolates and were most closely associated with genotypic cluster T3-T4-T11. A similar distance analysis using GTSA.B1 sequences identified nine South African AK-associated isolates as genotype T4 and three isolates from sewage sludge as genotype T5. Our results demonstrate the usefulness of 18S ribosomal DNA PCR amplimers ASA.S1 and GTSA.B1 for Acanthamoeba-specific detection and reliable genotyping, respectively, and provide further evidence that T4 is the predominant genotype in AK. PMID:11326011

  14. Rapid diagnosis of Acanthamoeba keratitis from corneal scrapings using indirect fluorescent antibody staining.

    PubMed

    Epstein, R J; Wilson, L A; Visvesvara, G S; Plourde, E G

    1986-09-01

    Two soft contact lens wearers using a homemade saline solution developed corneal stromal inflammation and epithelial ulceration and were both treated for months with a presumptive diagnosis of herpes simplex keratitis. Subsequently, corneal scrapings revealed refractile, cystic structures consistent with the appearance of Acanthamoeba. This was rapidly confirmed by indirect fluorescent antibody studies, and Acanthamoeba castellani was later identified by growth in culture in both cases. Acanthamoeba is being reported with increasing frequency as a pathogen responsible for chronic stromal keratitis and ulceration in contact lens wearers. Since specific therapy is required to control this organism, rapid diagnosis is essential. Indirect fluorescent antibody staining of corneal scrapings provides a simple means of accomplishing this goal with a high degree of accuracy. PMID:2428344

  15. The use of dimethyl sulfoxide in contact lens disinfectants is a potential preventative strategy against contracting Acanthamoeba keratitis.

    PubMed

    Siddiqui, Ruqaiyyah; Aqeel, Yousuf; Khan, Naveed Ahmed

    2016-10-01

    Acanthamoeba castellanii is the causative agent of blinding keratitis. Though reported in non-contact lens wearers, it is most frequently associated with improper use of contact lens. For contact lens wearers, amoebae attachment to the lens is a critical first step, followed by amoebae binding to the corneal epithelial cells during extended lens wear. Acanthamoeba attachment to surfaces (biological or inert) and migration is an active process and occurs during the trophozoite stage. Thus retaining amoebae in the cyst stage (dormant form) offers an added preventative measure in impeding parasite traversal from the contact lens onto the cornea. Here, we showed that as low as 3% DMSO, abolished A. castellanii excystation. Based on the findings, it is proposed that DMSO should be included in the contact lens disinfectants as an added preventative strategy against contracting Acanthamoeba keratitis.

  16. Use of 18S rRNA Gene-Based PCR Assay for Diagnosis of Acanthamoeba Keratitis in Non-Contact Lens Wearers in India

    PubMed Central

    Pasricha, Gunisha; Sharma, Savitri; Garg, Prashant; Aggarwal, Ramesh K.

    2003-01-01

    Identification of Acanthamoeba cysts and trophozoites in ocular tissues requires considerable expertise and is often time-consuming. An 18S rRNA gene-based PCR test, highly specific for the genus Acanthamoeba, has recently been reported in the molecular diagnosis of Acanthamoeba keratitis. This PCR assay was compared with conventional microbiological tests for the diagnosis of Acanthamoeba keratitis. In a pilot study, the PCR conditions with modifications were first tested on corneal scrapings from patients with culture-proven non-contact lens-related Acanthamoeba, bacterial, and fungal keratitis. This was followed by testing of corneal scrapings from 53 consecutive cases of microbial keratitis to determine sensitivity, specificity, and predictive values of the assay. All corneal scrapings from patients with proven Acanthamoeba keratitis showed a 463-bp amplicon, while no amplicon was obtained from patients with bacterial or fungal keratitis. Some of these amplified products were sequenced and compared with EMBL database reference sequences to validate these to be of Acanthamoeba origin. Out of 53 consecutive cases of microbial keratitis included for evaluating the PCR, 10 (18.9%) cases were diagnosed as Acanthamoeba keratitis on the basis of combined results of culture, smear, and PCR of corneal scrapings. Based on culture results as the “gold standard,” the sensitivity of PCR was the same as that of the smear (87.5%); however, the specificity and the positive and negative predictive values of PCR were marginally higher than the smear examination (97.8 versus 95.6%, 87.5 versus 77.8%, and 97.8 versus 97.7%) although the difference was not significant. This study confirms the efficacy of the PCR assay and is the first study to evaluate a PCR-based assay against conventional methods of diagnosis in a clinical setting. PMID:12843065

  17. Temperature limitation may explain the containment of the trophozoites in the cornea during Acanthamoeba castellanii keratitis.

    PubMed

    Nielsen, Mattias Kiel; Nielsen, Kim; Hjortdal, Jesper; Sørensen, Uffe B Skov

    2014-12-01

    Acanthamoeba keratitis is a serious sight-threatening disease. The relatively low temperature of the cornea may explain why amoebic infections usually are localized in this tissue and rarely spread to other parts of the eye. In this study, the growth rate of the amoeba Acanthamoeba castellanii was examined at different temperatures. The aim was to establish the optimal growth temperature for A. castellanii and to examine the growth within the vicinity of the core body temperature. The growth rates of four clinical and two environmental strains of A. castellanii were estimated at different temperatures, and temperature limitations for the trophozoite stage was established. Movements influenced by temperature gradients were monitored for two clinical strains of A. castellanii. The highest growth rate for each of the six amoebic strains tested was found to be close to 32 °C. The growth of the trophozoites of all examined strains was greatly reduced or completely halted at temperatures above 36 °C and encysted at the elevated temperature. Thus, the optimal growth temperature for the four strains of A. castellanii is close to the surface temperature of the human cornea, while the higher body core-temperature induced encysting of the amoebae. This may explain why most amoebic eye infections are confined to the cornea. PMID:25204727

  18. A novel antiamoebic agent against Acanthamoeba sp. - A causative agent for eye keratitis infection

    NASA Astrophysics Data System (ADS)

    Kusrini, Eny; Hashim, Fatimah; Azmi, Wan Nor Nadhirah Wan Noor; Amin, Nakisah Mat; Estuningtyas, Ari

    2016-01-01

    The terbium trinitrate.trihydrate.18-crown ether-6, Tb(NO3)3(OH2)3.(18C6) complex has been characterized by elemental analysis, photoluminescence and single X-ray diffraction. The IC50 values were determined based on MTT assay while light and fluorescence microscopy imaging were employed to evaluate the cellular morphological changes. Alkaline comet assay was performed to analyze the DNA damage. The photoluminescence spectrum of the Tb complex excited at 325 nm displayed seven luminescence peaks corresponding to the 5D4 → 7F0, 1, 2, 3, 4, 5, 6 transitions. The cytotoxicity and genotoxicity studies indicated that the Tb(NO3)3(OH2)3.(18C6) complex and its salt form as well as the 18C6 molecule have excellent anti-amoebic activity with very low IC50 values are 7, 2.6 and 1.2 μg/mL, respectively, with significant decrease (p < 0.05) in Acanthamoeba viability when the concentration was increased from 0 to 30 μg/mL. The mode of cell death in Acanthamoeba cells following treatment with the Tb complex was apoptosis. This is in contrast to the Tb(NO3)3.6H2O salt- and 18C6 molecule-treated Acanthamoeba, which exhibited necrotic type cells. The percentage of DNA damage following treatment with all the compounds at the IC25 values showed high percentage of type 1 with the % nuclei damage are 14.15 ± 2.4; 46.00 ± 4.2; 36.36 ± 2.4; 45.16 ± 0.6%, respectively for untreated, treated with Tb complex, Tb salt and 18C6 molecule. The work features promising potential of Tb(NO3)3(OH2)3.(18C6) complex as anti-amoebic agent, representing a therapeutic option for Acanthamoeba keratitis infection.

  19. A novel antiamoebic agent against Acanthamoeba sp.--A causative agent for eye keratitis infection.

    PubMed

    Kusrini, Eny; Hashim, Fatimah; Azmi, Wan Nor Nadhirah Wan Noor; Amin, Nakisah Mat; Estuningtyas, Ari

    2016-01-15

    The terbium trinitrate.trihydrate.18-crown ether-6, Tb(NO3)3(OH2)3.(18C6) complex has been characterized by elemental analysis, photoluminescence and single X-ray diffraction. The IC50 values were determined based on MTT assay while light and fluorescence microscopy imaging were employed to evaluate the cellular morphological changes. Alkaline comet assay was performed to analyze the DNA damage. The photoluminescence spectrum of the Tb complex excited at 325 nm displayed seven luminescence peaks corresponding to the (5)D4→(7)F(0, 1, 2, 3, 4, 5, 6) transitions. The cytotoxicity and genotoxicity studies indicated that the Tb(NO3)3(OH2)3.(18C6) complex and its salt form as well as the 18C6 molecule have excellent anti-amoebic activity with very low IC50 values are 7, 2.6 and 1.2 μg/mL, respectively, with significant decrease (p<0.05) in Acanthamoeba viability when the concentration was increased from 0 to 30 μg/mL. The mode of cell death in Acanthamoeba cells following treatment with the Tb complex was apoptosis. This is in contrast to the Tb(NO3)3.6H2O salt- and 18C6 molecule-treated Acanthamoeba, which exhibited necrotic type cells. The percentage of DNA damage following treatment with all the compounds at the IC25 values showed high percentage of type 1 with the % nuclei damage are 14.15±2.4; 46.00±4.2; 36.36±2.4; 45.16±0.6%, respectively for untreated, treated with Tb complex, Tb salt and 18C6 molecule. The work features promising potential of Tb(NO3)3(OH2)3.(18C6) complex as anti-amoebic agent, representing a therapeutic option for Acanthamoeba keratitis infection.

  20. Monitoring of in vitro dynamics of Acanthamoeba strains isolated from infected eyes as a useful tool in keratitis management.

    PubMed

    Chomicz, Lidia; Padzik, Marcin; Szaflik, Jacek P; Nahorski, Wacław L; Kryczka, Tomasz; Szaflik, Jerzy

    2014-11-01

    Free-living amoebae of Acanthamoeba genus are ubiquitous in various parts of the world. Some species of these amoebozoans present a serious risk to human health as the causative agents of vision-threatening diseases, Acanthamoeba keratitis. Correct diagnosis requires both a clinical examination of the cornea and amoebic form identification in affected eyes. Despite advances in pharmacotherapy, the infection is difficult to diagnose and to threat. Population dynamics of five different Acanthamoeba strains cultured in vitro under bacteria-free condition in BSC medium, was monitored in terms of diagnostic and therapeutic management. The range of protozoan number in the exponential growth phase, the morpho-physiological status of amoeba forms and their ability to multiply were evaluated. Results of the studies revealed that early and continued monitoring of the strains maintained in an axenic culture showed correlation between the dynamics of cultivated amoebae and the course of the disease, differences in response to pharmacotherapy and the surgical management efficacy. Concluding, the in vitro monitoring of dynamics of Acanthamoeba strains isolated from infected corneas may be important not only for proper diagnosis but also as a useful tool in keratitis management and therapeutic prognosis.

  1. [Acanthamoebal keratitis].

    PubMed

    Sarparanta, Katri; Lindbohm, Nina; Tervo, Timo; Tuisku, Ilpo; Jokiranta, Sakari

    2009-01-01

    Acanthamoebae are protists causing opportunistic infections to immunocompromised patients, and refractory keratitis also to others. Factors predisposing to Acanthamoebae keratitis include conditions that impair corneal defence mechanisms, such as traumas or use of contact lenses combined with exposure to contaminated water, for example. Delays in diagnosis and therapy will worsen the prognosis of acanthamoebal keratitis. In recent years, noninvasive in vivo confocal microscopy that enables an early diagnosis and is quicker than before, has become more common alongside conventional diagnostic methods, i.e. culture and smears made from corneal specimens.

  2. Twenty years of acanthamoeba diagnostics in Austria.

    PubMed

    Walochnik, Julia; Scheikl, Ute; Haller-Schober, Eva-Maria

    2015-01-01

    Acanthamoebae are the causative agents of an often seriously progressing keratitis (AK) occurring predominantly in contact lens wearers and can cause several disseminating infections potentially resulting in granulomatous amoebic encephalitis (GAE) in the immunocompromised host. Our institution is the Austrian reference laboratory for Acanthamoeba diagnostics and the aim of this study was to give an overview of proven cases of Acanthamoeba infections in Austria during the past 20 yr. All samples of patients with suspected AK or GAE were screened for Acanthamoeba spp. by culture and/or PCR and the detected amoebae were genotyped. Altogether, 154 cases of AK and three cases of GAE were diagnosed. Age of the AK patients ranged from 8 to 82 yr (mean 37.8) and 58% of the patients were female. Approximately 89% of the AK patients were contact lens wearers, almost all cases were unilateral and 19% of the patients required a keratoplasty. Age of the GAE patients ranged from 2 to 25 yr (mean 14.7), all were HIV-negative, but two were severely immunosuppressed at the time of diagnosis. The predominant genotype in the AK cases was T4, other genotypes found were T3, T5, T6, T10 and T11. The three GAE cases involved genotypes T2, T4 and T5.

  3. Twenty Years of Acanthamoeba Diagnostics in Austria

    PubMed Central

    Walochnik, Julia; Scheikl, Ute; Haller-Schober, Eva-Maria

    2015-01-01

    Acanthamoebae are the causative agents of an often seriously progressing keratitis (AK) occurring predominantly in contact lens wearers and can cause several disseminating infections potentially resulting in granulomatous amoebic encephalitis (GAE) in the immunocompromised host. Our institution is the Austrian reference laboratory for Acanthamoeba diagnostics and the aim of this study was to give an overview of proven cases of Acanthamoeba infections in Austria during the past 20 yr. All samples of patients with suspected AK or GAE were screened for Acanthamoeba spp. by culture and/or PCR and the detected amoebae were genotyped. Altogether, 154 cases of AK and three cases of GAE were diagnosed. Age of the AK patients ranged from 8 to 82 yr (mean 37.8) and 58% of the patients were female. Approximately 89% of the AK patients were contact lens wearers, almost all cases were unilateral and 19% of the patients required a keratoplasty. Age of the GAE patients ranged from 2 to 25 yr (mean 14.7), all were HIV-negative, but two were severely immunosuppressed at the time of diagnosis. The predominant genotype in the AK cases was T4, other genotypes found were T3, T5, T6, T10 and T11. The three GAE cases involved genotypes T2, T4 and T5. PMID:25047131

  4. Cytotoxic effect of acriflavine against clinical isolates of Acanthamoeba spp.

    PubMed

    Polat, Zubeyda Akin; Karakus, Gulderen

    2013-02-01

    Acanthamoeba keratitis (AK) is a potentially devastating and sight-threatening infection of the cornea caused by the ubiquitous free-living amoebae, Acanthamoeba species. Its eradication is difficult because the amoebas encyst, making it highly resistant to anti-amoebic drugs. Acriflavine neutral (ACF) has been used for treatment of microbial infections for humans and fishes. The aim of our study was to evaluate the time-dependent cytotoxicities of ACF against Acanthamoeba spp. Trophozoites and cysts of three different strains (strain PAT06 Acanthamoeba castellanii, strain 2HH Acanthamoeba hatchetti, and strain 11DS A. hatchetti) of Acanthamoeba spp. were tested. All strains had been isolated from patients suffering from a severe AK. The effects of the ACF with the concentrations ranging from 15 to 500 mg mL(-1) on the cytotoxicity of Acanthamoeba strains were examined. ACF showed a time- and dose-dependent amebicidal action on the trophozoites and cysts. Pat06 (A. castellanii) was the most resistant, while strain 11DS (A. hatchetti) was the most sensitive. As a result, ACF could be concluded as a new agent for the treatment of Acanthamoeba infections. On the other hand, it still needs to be further evaluated by in vivo test systems to confirm the efficiency of its biological effect. PMID:23052789

  5. Comparative proteomic analysis of extracellular secreted proteins expressed by two pathogenic Acanthamoeba castellanii clinical isolates and a non-pathogenic ATCC strain.

    PubMed

    Huang, Jian-Ming; Lin, Wei-Chen; Li, Sung-Chou; Shih, Min-Hsiu; Chan, Wen-Ching; Shin, Jyh-Wei; Huang, Fu-Chin

    2016-07-01

    Acanthamoeba keratitis (AK) is a serious ocular disease caused by pathogenic Acanthamoeba gaining entry through wounds in the corneal injury; generally, patients at risk for contracting AK wear contact lenses, usually over a long period of time. Moreover, pathogenic Acanthamoeba causes serious consequences: it makes the cornea turbid and difficult to operate on, including procedures such as enucleation of the eyeball. At present, diagnosis of this disease is not straightforward, and treatment is very demanding. We have established the comparative transcriptome and extracellular secreted proteomic database according to the non-pathogenic strain ATCC 30010 and the pathogenic strains NCKU_B and NCKU_D. We identified 44 secreted proteins successfully, 10 consensus secreted proteins and 34 strain-specific secreted proteins. These proteins may provide targets for therapy and immuno-diagnosis of Acanthamoeba infections. This study shows a suitable approach to identify secreted proteins in Acanthamoeba and provides new perspectives for the study of molecules potentially involved in the AK.

  6. Acanthamoeba spp. in Contact Lenses from Healthy Individuals from Madrid, Spain

    PubMed Central

    Gomes, Thiago dos Santos; Magnet, Angela; Izquierdo, Fernando; Vaccaro, Lucianna; Redondo, Fernando; Bueno, Sara; Sánchez, Maria Luisa; Angulo, Santiago; Fenoy, Soledad; Hurtado, Carolina; del Aguila, Carmen

    2016-01-01

    Purpose Acanthamoeba keratitis (AK) is a painful and potentially blinding corneal infection caused by Acanthamoeba spp. In Madrid, environmental studies have demonstrated a high presence of these free-living amoebae in tap water. Since most of AK cases occur in contact lenses (CL) wearers with inadequate hygiene habits, the presence of Acanthamoeba in discarded CL has been studied and compared with other common etiological agents of keratitis, such as Pseudomonas aeruginosa and Staphylococcus aureus. Methods One hundred and seventy-seven healthy individuals from Madrid contributed their discarded CL and answered a questionnaire on hygiene habits. DNA was extracted from the CL solution and analyzed by real-time PCR for Acanthamoeba, Pseudomonas aeruginosa and Staphylococcus aureus. These CL and their solutions were also cultured on non-nutrient agar to isolate Acanthamoeba. Results Among the 177 samples, Acanthamoeba DNA was detected in 87 (49.2%), P. aeruginosa DNA in 14 (7.9%) and S. aureus DNA in 19 (10.7%). Cultivable amoebae, however, were observed in only one sample (0.6%). This isolate was genotyped as T4. The habits reported by this CL owner included some recognized risk factors for AK, but in this study only the practice of “not cleaning the CL case” presented some statistical significant association with Acanthamoeba DNA presence. Detection of the investigated bacterial DNA did not demonstrate statistical significant association with the studied practices, but the presence of P. aeruginosa revealed a possible inhibition of Acanthamoeba in these samples. Conclusions The PCR results suggest a high presence of Acanthamoeba spp. in healthy CL wearers from Madrid, but we can assume that CL solutions are properly disinfecting the CL since only 1.1% of the positive PCR samples correspond to viable amoebae and, after four years, only one participant reported stronger ocular problems. Nevertheless, more studies are necessary to corroborate this hypothesis. PMID

  7. In vitro efficacies of clinically available drugs against growth and viability of an Acanthamoeba castellanii keratitis isolate belonging to the T4 genotype.

    PubMed

    Baig, Abdul Mannan; Iqbal, Junaid; Khan, Naveed Ahmed

    2013-08-01

    The effects of clinically available drugs targeting muscarinic cholinergic, adrenergic, dopaminergic, and serotonergic receptors; intracellular calcium levels and/or the function of calcium-dependent biochemical pathways; ion channels; and cellular pumps were tested against a keratitis isolate of Acanthamoeba castellanii belonging to the T4 genotype. In vitro growth inhibition (amoebistatic) assays were performed by incubating A. castellanii with various concentrations of drugs in the growth medium for 48 h at 30°C. To determine amoebicidal effects, amoebae were incubated with drugs in phosphate-buffered saline for 24 h, and viability was determined using trypan blue exclusion staining. For controls, amoebae were incubated with the solvent alone. Of the eight drugs tested, amlodipine, prochlorperazine, and loperamide showed potent amoebicidal effects, as no viable trophozoites were observed (>95% kill rate), while amiodarone, procyclidine, digoxin, and apomorphine exhibited up to 50% amoebicidal effects. In contrast, haloperidol did not affect viability, but all the drugs tested inhibited A. castellanii growth. Importantly, amlodipine, prochlorperazine, and loperamide showed compelling cysticidal effects. The cysticidal effects were irreversible, as cysts treated with the aforementioned drugs did not reemerge as viable amoebae upon inoculation in the growth medium. Except for apomorphine and haloperidol, all the tested drugs blocked trophozoite differentiation into cysts in encystation assays. Given the limited availability of effective drugs to treat amoebal infections, the clinically available drugs tested in this study represent potential agents for managing keratitis and granulomatous amoebic encephalitis caused by Acanthamoeba spp. and possibly against other meningoencephalitis-causing amoebae, such as Balamuthia mandrillaris and Naegleria fowleri. PMID:23669391

  8. Acanthamoeba Keratitis FAQs

    MedlinePlus

    ... lenses improperly Disinfecting lenses improperly (such as using tap water or homemade solutions to clean the lenses) Swimming, ... rinsed with sterile contact lens solution (never use tap water), emptied, and left open to dry after each ...

  9. Cytotoxic activity of N-chlorotaurine on Acanthamoeba spp.

    PubMed

    Fürnkranz, Ursula; Nagl, Markus; Gottardi, Waldemar; Köhsler, Martina; Aspöck, Horst; Walochnik, Julia

    2008-02-01

    Acanthamoeba spp. are the causative agents of Acanthamoeba keratitis (AK), which mainly occurs in contact lens wearers, and of skin lesions, granulomatous amoebic encephalitis (GAE), and disseminating diseases in the immunocompromised host. AK therapy is complex and irritating for the eye, skin lesions are difficult to treat, and there is no effective treatment for GAE. Therefore, new anti-Acanthamoeba drugs are needed. We investigated the anti-Acanthamoeba activity of N-chlorotaurine (NCT), an endogenous mild antiseptic. It was shown that NCT has amoebicidal qualities, both in phosphate-buffered saline (PBS) and in amoebic culture medium. After 6 h of treatment with 10 mM NCT in PBS, the levels of trophozoites of all strains investigated already showed at least a 2-log reduction. When the trophozoites were treated with 20 mM NCT in culture medium, they showed a 2-log reduction after 24 h. The addition of NH(4)Cl to NCT led to a faster decrease in the numbers of living cells, if tests were carried out in PBS. A delay of excystation was observed when cysts were treated with 55 mM (1%) NCT in culture medium. A complete failure of excystment was the result of treatment with 1% NCT plus 1% NH(4)Cl in PBS. Altogether, NCT clearly demonstrated amoebicidal activity at concentrations well tolerated by human tissues and might be useful as a topical drug for the treatment of Acanthamoeba infections. The addition of ammonium chloride can be considered to enhance the activity.

  10. Ecology of Acanthamoeba.

    PubMed

    De Jonckheere, J F

    1991-01-01

    Acanthamoeba is a free-living ameba that is present in all types of environments throughout the world. The recent increase in cases of keratitis, especially in relation to an increase in the use of contact lenses, is probably due to the omnipresence of the organism as a result of the pronounced resistance of its cysts to disinfection and desiccation. The temperature of the eye is lower than that of the rest of the human body. Therefore, the presence of Acanthamoeba strains that grow at lower temperatures may also contribute to infection, thereby increasing the number of possibly infectious amebas. Recent evidence, however, indicates that perhaps only a limited number of species cause ocular disease. Delineation of the exact species of Acanthamoeba that cause keratitis is a prerequisite for the study of the ecology of the keratitis-producing amebas. PMID:2047667

  11. Abietane diterpenoids from Salvia sclarea transformed roots as growth inhibitors of pathogenic Acanthamoeba spp.

    PubMed

    Kuźma, Łukasz; Derda, Monika; Hadaś, Edward; Wysokińska, Halina

    2015-01-01

    Amoebae from the genus Acanthamoeba are known agents leading to various diseases such as granulomatous amoebic encephalitis (GAE), a chronic progressive disease of the central nervous system, amoebic keratitis (AK), chronic eye infection, amoebic pneumitis (AP), chronic lung infection, and skin infections. It is known that various synthetic anti-Acanthamoeba substances are ineffective. Therefore, other substances, e.g., natural plant compounds, are the focus of biological investigations regarding anti-parasite activity. In this work, the ability of four abietane diterpenoids (ferruginol, salvipisone, aethiopinone, and 1-oxo-aethiopinone) to inhibit Acanthamoeba growth is reported. All investigated compounds were active against Acanthamoeba growing in vitro. Among them, ferruginol demonstrated the highest activity against Acanthamoeba. This compound inhibited Acanthamoeba growth by about 72% in a 3-day exposure period (IC50 17.45 μM), while aethiopinone and 1-oxo-aethiopinone demonstrated this activity at the level of 55-56%. Salvipisone reduced the growth of Acanthamoeba in vitro culture by 39%. For this compound, the value of IC50 was 701.94 μM after 72 h of exposure.

  12. Characterization of a human-pathogenic Acanthamoeba griffini isolated from a contact lens-wearing keratitis patient in Spain.

    PubMed

    Heredero-Bermejo, I; Criado-Fornelio, A; De Fuentes, I; Soliveri, J; Copa-Patiño, J L; Pérez-Serrano, J

    2015-02-01

    Amoebae were isolated from contact lenses of a symptomatic lens wearer in Spain. Protozoa were characterized by studying their morphology, biology, protease activity and the 18S rRNA gene sequence. Morphology of the organism was observed by light microscopy, scanning electron microscopy and transmission electron microscopy. Its structure corresponded to an amphizoic amoeba. The protozoa grew well at 37 °C and poorly at lower temperatures. In addition, it was capable of lysing mammalian cells in vitro. A major 56 kDa proteolytic enzyme was observed in amoeba crude extracts by gelatin-sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Most proteolytic enzymes in protozoa extracts showed significant activity over a wide range of pH (3-9) and temperature (8-45 °C) values. The assays on inhibition of protease activity indicated strongly that enzymes detected in amoeba extracts corresponded to serine proteases and, to a lesser extent, cysteine proteases. The use of proteinase inhibitors on a tissue culture model proved that the proteinase activity is critical for developing focal lesions in HeLa cell monolayers. Finally, partial sequencing of the 18S ribosomal RNA gene and phylogenetic analyses indicated that the isolate is closely related to Acanthamoeba griffini H37 from the UK (T3 genotype).

  13. Pathogenic Strains of Acanthamoeba Are Recognized by TLR4 and Initiated Inflammatory Responses in the Cornea

    PubMed Central

    Alizadeh, Hassan; Tripathi, Trivendra; Abdi, Mahshid; Smith, Ashley Dawn

    2014-01-01

    Free-living amoebae of the Acanthamoeba species are the causative agent of Acanthamoeba keratitis (AK), a sight-threatening corneal infection that causes severe pain and a characteristic ring-shaped corneal infiltrate. Innate immune responses play an important role in resistance against AK. The aim of this study is to determine if Toll-like receptors (TLRs) on corneal epithelial cells are activated by Acanthamoeba, leading to initiation of inflammatory responses in the cornea. Human corneal epithelial (HCE) cells constitutively expressed TLR1, TLR2, TLR3, TLR4, and TLR9 mRNA, and A. castellanii upregulated TLR4 transcription. Expression of TLR1, TLR2, TLR3, and TLR9 was unchanged when HCE cells were exposed to A. castellanii. IL-8 mRNA expression was upregulated in HCE cells exposed to A. castellanii. A. castellanii and lipopolysaccharide (LPS) induced significant IL-8 production by HCE cells as measured by ELISA. The percentage of total cells positive for TLR4 was higher in A. castellanii stimulated HCE cells compared to unstimulated HCE cells. A. castellanii induced upregulation of IL-8 in TLR4 expressing human embryonic kidney (HEK)-293 cells, but not TLR3 expressing HEK-293 cells. TLR4 neutralizing antibody inhibited A. castellanii-induced IL-8 by HCE and HEK-293 cells. Clinical strains but not soil strains of Acanthamoeba activated TLR4 expression in Chinese hamster corneas in vivo and in vitro. Clinical isolates but not soil isolates of Acanthamoeba induced significant (P< 0.05) CXCL2 production in Chinese hamster corneas 3 and 7 days after infection, which coincided with increased inflammatory cells in the corneas. Results suggest that pathogenic species of Acanthamoeba activate TLR4 and induce production of CXCL2 in the Chinese hamster model of AK. TLR4 may be a potential target in the development of novel treatment strategies in Acanthamoeba and other microbial infections that activate TLR4 in corneal cells. PMID:24633052

  14. Biology and pathogenesis of Acanthamoeba.

    PubMed

    Siddiqui, Ruqaiyyah; Khan, Naveed Ahmed

    2012-01-01

    Acanthamoeba is a free-living protist pathogen, capable of causing a blinding keratitis and fatal granulomatous encephalitis. The factors that contribute to Acanthamoeba infections include parasite biology, genetic diversity, environmental spread and host susceptibility, and are highlighted together with potential therapeutic and preventative measures. The use of Acanthamoeba in the study of cellular differentiation mechanisms, motility and phagocytosis, bacterial pathogenesis and evolutionary processes makes it an attractive model organism. There is a significant emphasis on Acanthamoeba as a Trojan horse of other microbes including viral, bacterial, protists and yeast pathogens.

  15. Biology and pathogenesis of Acanthamoeba

    PubMed Central

    2012-01-01

    Acanthamoeba is a free-living protist pathogen, capable of causing a blinding keratitis and fatal granulomatous encephalitis. The factors that contribute to Acanthamoeba infections include parasite biology, genetic diversity, environmental spread and host susceptibility, and are highlighted together with potential therapeutic and preventative measures. The use of Acanthamoeba in the study of cellular differentiation mechanisms, motility and phagocytosis, bacterial pathogenesis and evolutionary processes makes it an attractive model organism. There is a significant emphasis on Acanthamoeba as a Trojan horse of other microbes including viral, bacterial, protists and yeast pathogens. PMID:22229971

  16. Acanthamoeba infection in lungs of mice expressed by toll-like receptors (TLR2 and TLR4).

    PubMed

    Derda, Monika; Wojtkowiak-Giera, Agnieszka; Kolasa-Wołosiuk, Agnieszka; Kosik-Bogacka, Danuta; Hadaś, Edward; Jagodziński, Paweł P; Wandurska-Nowak, Elżbieta

    2016-06-01

    Toll-like receptors (TLRs) play a key role in the innate immune responses to a variety of pathogens including parasites. TLRs are among the most highly conserved in the evolution of the receptor family, localized mainly on cells of the immune system and on other cells such as lung cells. The aim of this study was to determine for the first time the expression of TLR2 and TLR4 in the lung of Acanthamoeba spp. infected mice using quantitative real-time polymerase chain reaction (Q-PCR) and immunohistochemical (IHC) staining. The Acanthamoeba spp. were isolated from a patient with Acanthamoeba keratitis (AK) (strain Ac 55) and from environmental samples of water from Malta Lake (Poznań, Poland - strain Ac 43). We observed a significantly increased level of expression of TLR2 as well as TLR4 mRNA from 2 to 30 days post Acanthamoeba infection (dpi) in the lungs of mice infected with Ac55 (KP120880) and Ac43 (KP120879) strains. According to our observations, increased TLR2 and TLR4 expression in the pneumocytes, interstitial cells and epithelial cells of the bronchial tree may suggest an important role of these receptors in protective immunity against Acanthamoeba infection in the lung. Moreover, increased levels of TLR2 and TLR4 mRNA expression in infected Acanthamoeba mice may suggest the involvement of these TLRs in the recognition of this amoeba pathogen-associated molecular pattern (PAMP). PMID:26940205

  17. Acanthamoeba infection in lungs of mice expressed by toll-like receptors (TLR2 and TLR4).

    PubMed

    Derda, Monika; Wojtkowiak-Giera, Agnieszka; Kolasa-Wołosiuk, Agnieszka; Kosik-Bogacka, Danuta; Hadaś, Edward; Jagodziński, Paweł P; Wandurska-Nowak, Elżbieta

    2016-06-01

    Toll-like receptors (TLRs) play a key role in the innate immune responses to a variety of pathogens including parasites. TLRs are among the most highly conserved in the evolution of the receptor family, localized mainly on cells of the immune system and on other cells such as lung cells. The aim of this study was to determine for the first time the expression of TLR2 and TLR4 in the lung of Acanthamoeba spp. infected mice using quantitative real-time polymerase chain reaction (Q-PCR) and immunohistochemical (IHC) staining. The Acanthamoeba spp. were isolated from a patient with Acanthamoeba keratitis (AK) (strain Ac 55) and from environmental samples of water from Malta Lake (Poznań, Poland - strain Ac 43). We observed a significantly increased level of expression of TLR2 as well as TLR4 mRNA from 2 to 30 days post Acanthamoeba infection (dpi) in the lungs of mice infected with Ac55 (KP120880) and Ac43 (KP120879) strains. According to our observations, increased TLR2 and TLR4 expression in the pneumocytes, interstitial cells and epithelial cells of the bronchial tree may suggest an important role of these receptors in protective immunity against Acanthamoeba infection in the lung. Moreover, increased levels of TLR2 and TLR4 mRNA expression in infected Acanthamoeba mice may suggest the involvement of these TLRs in the recognition of this amoeba pathogen-associated molecular pattern (PAMP).

  18. Isolation and Genotyping of Acanthamoeba Strains from Environmental Sources in Ahvaz City, Khuzestan Province, Southern Iran

    PubMed Central

    Rahdar, M; Niyyati, M; Salehi, M; Feghhi, M; Makvandi, M; Pourmehdi, M; Farnia, S

    2012-01-01

    Background Acanthamoeba spp. are free-living amoebae commonly found in the environmental sources such as water, soil, and air. This ubiquitous amoeba is the causative agent of amoebic keratitis (AK). The objective of the present study was to investigate the presence of Acanthamoeba spp. in water and soil sources in Ahvaz City, Khuzestan Province, southern Iran. Methods In general, 110 samples of water and soil were taken from different localities of Ahvaz including agricultural canals, rivers, and swimming pools. Filtration and cultivation were carried out on non-nutrient agar medium (NNA). Axenic cultivation was performed for all of positive isolates. PCR analysis was conducted on positive samples. Sequencing was done for 15 PCR products. Genotypes were identified by Blast search and homology analysis. Results Acanthamoeba spp. was found in 43 (71.6%) of samples of water and 13 (26%) soil samples. Genotyping of 15 samples proved that Acanthamoeba belonged to T4 (86.6%), T2 (6.6%), and T5 (6.6%) genotypes. Conclusion TYI-S-33 medium could be better than PYG medium for Acanthamoeba axenic culture. PMID:23323088

  19. Genetic Characterization of Clinical Acanthamoeba Isolates from Japan using Nuclear and Mitochondrial Small Subunit Ribosomal RNA

    PubMed Central

    Rahman, Md Moshiur; Yagita, Kenji; Kobayashi, Akira; Oikawa, Yosaburo; Hussein, Amjad I.A.; Matsumura, Takahiro

    2013-01-01

    Because of an increased number of Acanthamoeba keratitis (AK) along with associated disease burdens, medical professionals have become more aware of this pathogen in recent years. In this study, by analyzing both the nuclear 18S small subunit ribosomal RNA (18S rRNA) and mitochondrial 16S rRNA gene loci, 27 clinical Acanthamoeba strains that caused AK in Japan were classified into 3 genotypes, T3 (3 strains), T4 (23 strains), and T5 (one strain). Most haplotypes were identical to the reference haplotypes reported from all over the world, and thus no specificity of the haplotype distribution in Japan was found. The T4 sub-genotype analysis using the 16S rRNA gene locus also revealed a clear sub-conformation within the T4 cluster, and lead to the recognition of a new sub-genotype T4i, in addition to the previously reported sub-genotypes T4a-T4h. Furthermore, 9 out of 23 strains in the T4 genotype were identified to a specific haplotype (AF479533), which seems to be a causal haplotype of AK. While heterozygous nuclear haplotypes were observed from 2 strains, the mitochondrial haplotypes were homozygous as T4 genotype in the both strains, and suggested a possibility of nuclear hybridization (mating reproduction) between different strains in Acanthamoeba. The nuclear 18S rRNA gene and mitochondrial 16S rRNA gene loci of Acanthamoeba spp. possess different unique characteristics usable for the genotyping analyses, and those specific features could contribute to the establishment of molecular taxonomy for the species complex of Acanthamoeba. PMID:24039282

  20. Comparison of specific activity and cytopathic effects of purified 33 kDa serine proteinase from Acanthamoeba strains with different degree of virulence

    PubMed Central

    Kim, Won-Tae; Kong, Hyun-Hee; Ha, Young-Ran; Hong, Yeon-Chul; Jeong, Hae Jin; Yu, Hak Sun

    2006-01-01

    The pathogenic mechanism of granulomatous amebic encephalitis (GAE) and amebic keratitis (AK) by Acanthamoeba has yet to be clarified. Protease has been recognized to play an important role in the pathogenesis of GAE and AK. In the present study, we have compared specific activity and cytopathic effects (CPE) of purified 33 kDa serine proteinases from Acanthamoeba strains with different degree of virulence (A. healyi OC-3A, A. lugdunensis KA/E2, and A. castellanii Neff). Trophozoites of the 3 strains revealed different degrees of CPE on human corneal epithelial (HCE) cells. The effect was remarkably reduced by adding phenylmethylsulfonylfluoride (PMSF), a serine proteinase inhibitor. This result indicated that PMSF-susceptible proteinase is the main component causing cytopathy to HCE cells by Acanthamoeba. The purified 33 kDa serine proteinase showed strong activity toward HCE cells and extracellular matrix proteins. The purified proteinase from OC-3A, the most virulent strain, demonstrated the highest enzyme activity compared to KA/E2, an ocular isolate, and Neff, a soil isolate. Polyclonal antibodies against the purified 33 kDa serine proteinase inhibit almost completely the proteolytic activity of culture supernatant of Acanthamoeba. In line with these results, the 33 kDa serine proteinase is suggested to play an important role in pathogenesis and to be the main component of virulence factor of Acanthamoeba. PMID:17170574

  1. Hypoxia attenuates inflammatory mediators production induced by Acanthamoeba via Toll-like receptor 4 signaling in human corneal epithelial cells

    SciTech Connect

    Pan, Hong; Wu, Xinyi

    2012-04-13

    Highlights: Black-Right-Pointing-Pointer Hypoxia attenuates Acanthamoeba-induced the production of IL-8 and IFN-{beta}. Black-Right-Pointing-Pointer Hypoxia inhibits TLR4 expression in a time-dependent manner in HCECs. Black-Right-Pointing-Pointer Hypoxia inhibits Acanthamoeba-induced the activation of NF-{kappa}B and ERK1/2 in HCECs. Black-Right-Pointing-Pointer Hypoxia decreases Acanthamoeba-induced inflammatory response via TLR4 signaling. Black-Right-Pointing-Pointer LPS-induced the secretion of IL-6 and IL-8 is abated by hypoxia via TLR4 signaling. -- Abstract: Acanthamoeba keratitis (AK) is a vision-threatening corneal infection that is intimately associated with contact lens use which leads to hypoxic conditions on the corneal surface. However, the effect of hypoxia on the Acanthamoeba-induced host inflammatory response of corneal epithelial cells has not been studied. In the present study, we investigated the effect of hypoxia on the Acanthamoeba-induced production of inflammatory mediators interleukin-8 (IL-8) and interferon-{beta} (IFN-{beta}) in human corneal epithelial cells and then evaluated its effects on the Toll-like receptor 4 (TLR4) signaling, including TLR4 and myeloid differentiation primary response gene (88) (MyD88) expression as well as the activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-{kappa}B) and extracellular signal-regulated kinases 1/2 (ERK1/2). We then studied the effect of hypoxia on a TLR4-specific inflammatory response triggered by the TLR4 ligand lipopolysaccharide (LPS). Our data showed that hypoxia significantly decreased the production of IL-8 and IFN-{beta}. Furthermore, hypoxia attenuated Acanthamoeba-triggered TLR4 expression as well as the activation of NF-{kappa}B and ERK1/2, indicating that hypoxia abated Acanthamoeba-induced inflammatory responses by affecting TLR4 signaling. Hypoxia also inhibited LPS-induced IL-6 and IL-8 secretion, myeloid differentiation primary response gene (88

  2. Fluorescent Oligonucleotide Probes for Clinical and Environmental Detection of Acanthamoeba and the T4 18S rRNA Gene Sequence Type

    PubMed Central

    Stothard, Diane R.; Hay, John; Schroeder-Diedrich, Jill M.; Seal, David V.; Byers, Thomas J.

    1999-01-01

    The first genus- and subgenus-specific fluorescent oligonucleotide probes for in situ staining of Acanthamoeba are described. Sequences of these phylogeny-based probes complement the 18S rRNA and the gene encoding it (18S rDNA). The genus-specific probe (GSP) is a fluorescein-labeled 22-mer specific for Acanthamoeba as shown here by its hybridization to growing trophozoites of all 12 known Acanthamoeba 18S rDNA sequence types and by its failure to hybridize with amoebae of two other genera (Hartmannella vermiformis and Balamuthia mandrillaris), two human cell lines, and two bacteria (Pseudomonas aeruginosa and Escherichia coli). The sequence type T4-specific probe (ST4P) is a rhodamine-labeled 30-mer specific for Acanthamoeba 18S rDNA sequence type T4, as shown here in hybridization tests with trophozoites of all 12 sequence types. T4 is the subgenus group associated most closely with Acanthamoeba keratitis (AK). GSP also was tested with corneal scrapings from 17 patients with a high index of clinical suspicion of AK plus 5 patient controls. GSP stained both trophozoites and cysts, although nonspecific cyst wall autofluorescence also was observed. Results could be obtained with GSP in 1 to 2 days, and based on results from cell culture tests, the probe correctly detected the presence or absence of Acanthamoeba in 21 of 24 specimens from the 22 patients. The use of GSP with cultured trophozoites and cysts from corneal scrapings has illustrated the suitability of using fluorescent oligonucleotide probes for identification of the genus Acanthamoeba in both environmental and clinical samples. In addition, the use of ST4P with cultured amoebae has indicated the potential of oligonucleotide probes for use in subgenus classification. PMID:10405422

  3. Isolation of Acanthamoeba from the rhizosphere of maize and lucerne plants

    NASA Astrophysics Data System (ADS)

    Orosz, Erika; Farkas, Ágnes; Ködöböcz, László; Becsak, Péter; Danka, József; Kucsera, István; Füleky, György

    2013-04-01

    Acanthamoeba species are free-living amoebae that can be found in almost every range of environments. Within this genus, a number of species are recognized as human pathogens, potentially causing Acanthamoeba keratitis, granulomatous amoebic encephalitis, and chronic granulomatous lesions. Soil and water samples were taken from experimental station at Julianna Major of Plant Protection Institute of Centre for Agricultural Research, Hungarian Academy of Sciences. We detected living Acanthamoeba spp. based on culture- confirmed detection combined with the molecular taxonomic identification method. Living Acanthamoeba spp. were detected in thirteen (65%) samples. The presence of Acanthamoeba spp. in the samples depends significantly on the rhizosphere plants. The most frequently identified living Acanthamoeba genotype was T4 followed by T11, T2/T6 and T17. Genotypes T4 and T11 of Acanthamoeba, are responsible for Acanthamoeba keratitis as well as granulomatous amoebic encephalitis, and should therefore be considered as a potential health risk associated with human activities in the environment.

  4. Distinguishing infective versus noninfective keratitis

    PubMed Central

    Srinivasan, M; Mascarenhas, Jeena

    2008-01-01

    For the purpose of this symposium, the term ″keratitis″ implies suppurative nonviral and viral keratitis. Corneal ulcers have been described in ancient literature. But even today, despite the availability of a wide range of newer antimicrobials and new diagnostic techniques, infective keratitis continues to pose a diagnostic and therapeutic challenge. This article focuses on the key diagnostic clinical features of the most common organisms causing infective keratitis - bacteria, fungi, viruses, nocardia and acanthamoeba - in India. While the clinical features in some cases are fairly straightforward, most cases challenge the clinician. We describe the salient clinical features which can help arrive at a diagnosis to begin appropriate treatment immediately, prior to the laboratory report. PMID:18417820

  5. Phylogenetic evidence for a new genotype of Acanthamoeba (Amoebozoa, Acanthamoebida).

    PubMed

    Corsaro, Daniele; Venditti, Danielle

    2010-06-01

    Acanthamoeba are widespread free-living amoebae, able to cause infection in animals, with keratitis and granulomatous encephalitis as major diseases in humans. Recent developments in the subgenus classification are based on the determination of the nucleotide sequence of the 18S rDNA. By this mean, Acanthamoeba have been clustered into 15 sequence types or genotypes, called T1 to T15. In this study, we analysed near full 18S rDNA of an Acanthamoeba recovered from an environmental sample and various unidentified Acanthamoeba sequences retrieved from GenBank. We provided phylogenetic evidence for a new genotype, which we proposed to name T16.

  6. Acanthamoeba Migration in an Electric Field

    PubMed Central

    Rudell, Jolene Chang; Gao, Jing; Sun, Yuxin; Sun, Yaohui; Chodosh, James; Schwab, Ivan; Zhao, Min

    2013-01-01

    Purpose. We investigated the in vitro response of Acanthamoeba trophozoites to electric fields (EFs). Methods. Acanthamoeba castellanii were exposed to varying strengths of an EF. During EF exposure, cell migration was monitored using an inverted microscope equipped with a CCD camera and the SimplePCI 5.3 imaging system to capture time-lapse images. The migration of A. castellanii trophozoites was analyzed and quantified with ImageJ software. For analysis of cell migration in a three-dimensional culture system, Acanthamoeba trophozoites were cultured in agar, exposed to an EF, digitally video recorded, and analyzed at various Z focal planes. Results. Acanthamoeba trophozoites move at random in the absence of an EF, but move directionally in response to an EF. Directedness in the absence of an EF is 0.08 ± 0.01, while in 1200 mV/mm EF, directedness is significantly higher at −0.65 ± 0.01 (P < 0.001). We find that the trophozoite migration response is voltage-dependent, with higher directionality with higher voltage application. Acanthamoeba move directionally in a three-dimensional (3D) agar system as well when exposed to an EF. Conclusions. Acanthamoeba trophozoites move directionally in response to an EF in a two-dimensional and 3D culture system. Acanthamoeba trophozoite migration is also voltage-dependent, with increased directionality with increasing voltage. This may provide new treatment modalities for Acanthamoeba keratitis. PMID:23716626

  7. Allovahlkampfia spelaea Causing Keratitis in Humans

    PubMed Central

    Tolba, Mohammed Essa Marghany; Huseein, Enas Abdelhameed Mahmoud; Farrag, Haiam Mohamed Mahmoud; Mohamed, Hanan El Deek; Kobayashi, Seiki; Suzuki, Jun; Ali, Tarek Ahmed Mohamed; Sugano, Sumio

    2016-01-01

    Background Free-living amoebae are present worldwide. They can survive in different environment causing human diseases in some instances. Acanthamoeba sp. is known for causing sight-threatening keratitis in humans. Free-living amoeba keratitis is more common in developing countries. Amoebae of family Vahlkampfiidae are rarely reported to cause such affections. A new genus, Allovahlkampfia spelaea was recently identified from caves with no data about pathogenicity in humans. We tried to identify the causative free-living amoeba in a case of keratitis in an Egyptian patient using morphological and molecular techniques. Methods Pathogenic amoebae were culture using monoxenic culture system. Identification through morphological features and 18S ribosomal RNA subunit DNA amplification and sequencing was done. Pathogenicity to laboratory rabbits and ability to produce keratitis were assessed experimentally. Results Allovahlkampfia spelaea was identified as a cause of human keratitis. Whole sequence of 18S ribosomal subunit DNA was sequenced and assembled. The Egyptian strain was closely related to SK1 strain isolated in Slovenia. The ability to induce keratitis was confirmed using animal model. Conclusions This the first time to report Allovahlkampfia spelaea as a human pathogen. Combining both molecular and morphological identification is critical to correctly diagnose amoebae causing keratitis in humans. Use of different pairs of primers and sequencing amplified DNA is needed to prevent misdiagnosis. PMID:27415799

  8. Interstitial keratitis

    MedlinePlus

    ... cornea. This condition is often caused by infections. Syphilis is the most common cause of interstitial keratitis, ... Tuberculosis In the United States, most cases of syphilis are recognized and treated before this eye condition ...

  9. Bacterial Keratitis

    MedlinePlus

    ... very quickly, and if left untreated, can cause blindness. The bacteria usually responsible for this type of ... to intense ultraviolet radiation exposure, e.g. snow blindness or welder's arc eye). Next Bacterial Keratitis Symptoms ...

  10. Current Status of Acanthamoeba in Iran: A Narrative Review Article

    PubMed Central

    NIYYATI, Maryam; REZAEIAN, Mostafa

    2015-01-01

    Background: Free-living amoebae belonging to the genus Acanthamoeba have an environmental distribution. Amoebic keratitis due to these protozoan parasites continue to rise in Iran and worldwide. In Iran, there are various researches regarding both morphological and molecular identification of Acanthamoeba spp. in environmental and clinical samples. However, there is no thorough review about Acanthamoeba genotypes and their distribution in environmental sources such as water, dust and biofilm in Iran. Besides, according to increasing cases of Amoebic keratitis in the region awareness regarding the pathogenic potential of these sight-threatening amoebae is of utmost importance. Methods: We conducted a thorough review based on the database sources such as MEDLINE, PubMed and Google scholar. No restrictions were placed on study date, study design or language of publication. We searched all valuable and relevant information considering the occurrence of the Acanthamoeba in both environmental and clinical samples. Results: According to our thorough review Acanthamoeba belonging to T4 genotype is the most prevalent type strain in environmental and clinical samples in several regions in Iran and worldwide, however, there are reports regarding Acanthamoeba belonging to other genotypes such as T2, T3, T5, T6 and T11 and the mentioned point could leads us to more researches with the goal of presenting the real genotype dominance of Acanthamoeba and related disease in the country. Conclusion: Overall, the present review will focus on present status of genotypes of Acanthamoeba in Iran during recent years. PMID:26246812

  11. Genotypic heterogeneity based on 18S-rRNA gene sequences among Acanthamoeba isolates from clinical samples in Italy.

    PubMed

    Di Cave, David; D' Alfonso, Rossella; Dussey Comlavi, Kodjo A; D' Orazi, Carlo; Monno, Rosa; Berrilli, Federica

    2014-11-01

    Acanthamoeba keratitis (AK) is an ocular disease caused by members of a genus of free-living amoebae and it is associated predominantly with contact lens (CL) use. This study reports 55 cases of AK diagnosed in Italy. Genotype identification was carried out by PCR assay followed by sequence analysis of the 18S rRNA gene using the genus specific primers JDP1 and JDP2. Genotype assignment was based on phenetic analysis of the ASA.S1 subset of the small-subunit rRNA gene sequences. The material has been collected at the Polyclinic Tor Vergata of Rome for a total of 19 isolates and at the Polyclinic Hospital of Bari (36 isolates). Thirty-three out of the 55 genetically characterized isolates were assigned to the genotype T4. Ten isolates were identified as belonging to the genotype T15 thus confirming the first association between the genotype T15 and human amoebic keratitis previously described from the same area. We underline the occurrence of the genotype T3 and T11 identified for the first time in the country.

  12. Diversity and seasonal impact of Acanthamoeba species in a subtropical rivershed.

    PubMed

    Kao, Po-Min; Chou, Ming-Yuan; Tao, Chi-Wei; Huang, Wen-Chien; Hsu, Bing-Mu; Shen, Shu-Min; Fan, Cheng-Wei; Chiu, Yi-Chou

    2013-01-01

    This study evaluated the presence of Acanthamoeba species in the Puzih River watershed, which features typical subtropical monsoon climate and is located just above the Tropic of Cancer in Taiwan. The relationship between the seasonal and geographical distributions of Acanthamoeba species in this rivershed was also investigated. Acanthamoeba species were detected in water samples using the amoebal enrichment culture method and confirmed by PCR. A total of 136 water samples were included in this study, 16 (11.7%) of which contained Acanthamoeba species. Samples with the highest percentage of Acanthamoeba (32.4%) were obtained during the summer season, mainly from upstream areas. The identified species in the four seasons included Acanthamoeba palestinensis (T2), Acanthamoeba sp. IS2/T4 (T4), Acanthamoeba lenticulata (T5), Acanthamoeba hatchetti (T11), Acanthamoeba healyi (T12), and Acanthamoeba jacobsi (T15). The most frequently identified Acanthamoeba genotype was T4 (68.7%). Acanthamoeba genotype T4 is responsible for Acanthamoeba keratitis and should be considered for associated human health risk potential in the rivershed.

  13. Diversity and Seasonal Impact of Acanthamoeba Species in a Subtropical Rivershed

    PubMed Central

    Kao, Po-Min; Chou, Ming-Yuan; Tao, Chi-Wei; Huang, Wen-Chien; Hsu, Bing-Mu; Shen, Shu-Min; Fan, Cheng-Wei; Chiu, Yi-Chou

    2013-01-01

    This study evaluated the presence of Acanthamoeba species in the Puzih River watershed, which features typical subtropical monsoon climate and is located just above the Tropic of Cancer in Taiwan. The relationship between the seasonal and geographical distributions of Acanthamoeba species in this rivershed was also investigated. Acanthamoeba species were detected in water samples using the amoebal enrichment culture method and confirmed by PCR. A total of 136 water samples were included in this study, 16 (11.7%) of which contained Acanthamoeba species. Samples with the highest percentage of Acanthamoeba (32.4%) were obtained during the summer season, mainly from upstream areas. The identified species in the four seasons included Acanthamoeba palestinensis (T2), Acanthamoeba sp. IS2/T4 (T4), Acanthamoeba lenticulata (T5), Acanthamoeba hatchetti (T11), Acanthamoeba healyi (T12), and Acanthamoeba jacobsi (T15). The most frequently identified Acanthamoeba genotype was T4 (68.7%). Acanthamoeba genotype T4 is responsible for Acanthamoeba keratitis and should be considered for associated human health risk potential in the rivershed. PMID:24490160

  14. Acanthamoeba meningoencephalitis

    PubMed Central

    Chandra, S. R.; Adwani, Sikandar; Mahadevan, Anitha

    2014-01-01

    Report of a case of young immunocompetent male adult with autopsy proven acanthamoeba meningoencephalitis. The patient presented with a protracted febrile illness of 3 months duration with features of meningoencephalitis, this was followed by rapid deterioration while on anti tuberculous therapy and steroids and ended fatally. His magnetic resonance imaging showed features of hemorrhagic meningoencephalitis and magnetic resonance spectroscopy showed choline peak. Autopsy revealed necrotizing meningoencephalitis and intraocular colonization due to acanthamoeba. PMID:24753675

  15. [Observations on Acanthamoeba trophozoites in axenic cultures and their staining characteristics with different stains].

    PubMed

    Polat, Zübeyde Akin; Ozçelik, Semra; Vural, Ayşe; Saygi, Gülendame

    2007-01-01

    Acanthamoeba spp. are among the most prevalent protozoa found in the environment. The species of this genus are the causative agents of granulomatous amebic encephalitis (GAE), a fatal disease of the central nervous system (CNS), and amebic keratitis (AK), a painful sight-threatening disease of the eye. In this study we have used two species of Acanthamoeba, Acanthamoeba castellanii and A. hatchetti, both were obtained from Vienna, Austria. They were cultivated on non-nutritious agar seeded with Escherichia coli and PPYG (protease peptone-yeast extract-glucose) medium. Our aim was to concentrate on three points in relation to the trophozoites and cysts stages of these species as follows: (i) to observe their morphology, (ii). to confirm our previous observation of a canal between two trophozoites. The bridge-like connection between these trophozoites greatly resembled the one that can be observed in conjugation during an exchange of genetic material. Two tro-phozoites with a bridge-like extension between them keep their position for at least 200 minutes. (iii). to detect the reactions of trophozoites to various stains. According to our findings in regard to these three points: (i). trophozoites with more than one nucleus are often seen in axenic cultures. (ii). This resembles a type of conjugation with a transfer of genetic material between two trophozoites. Certainly, this needs further investigation using more sophisticated methods. (iii). trophozoites equally stained well with Heidenhain's iron haematoxylin, Giemsa, PAS, Masson Trichrome, and Toludin-O stains. However, our results with reticulin, PAP, Van Gison, Musicarmine and Orsein stains were not satisfactory.

  16. Fungal keratitis

    PubMed Central

    Tuli, Sonal S

    2011-01-01

    Clinical question: What is the most appropriate management of fungal keratitis? Results: Traditionally, topical Natamycin is the most commonly used medication for filamentous fungi while Amphotericin B is most commonly used for yeast. Voriconazole is rapidly becoming the drug of choice for all fungal keratitis because of its wide spectrum of coverage and increased penetration into the cornea. Implementation: Repeated debridement of the ulcer is recommended for the penetration of topical medications. While small, peripheral ulcers may be treated in the community, larger or central ulcers, especially if associated with signs suggestive of anterior chamber penetration should be referred to a tertiary center. Prolonged therapy for approximately four weeks is usually necessary. PMID:21468333

  17. Photochemotherapeutic strategy against Acanthamoeba infections.

    PubMed

    Aqeel, Yousuf; Siddiqui, Ruqaiyyah; Anwar, Ayaz; Shah, Muhammad Raza; Khoja, Shahrukh; Khan, Naveed Ahmed

    2015-01-01

    Acanthamoeba is a protist pathogen that can cause serious human infections, including blinding keratitis and a granulomatous amoebic encephalitis that almost always results in death. The current treatment for these infections includes a mixture of drugs, and even then, a recurrence can occur. Photochemotherapy has shown promise in the treatment of Acanthamoeba infections; however, the selective targeting of pathogenic Acanthamoeba has remained a major concern. The mannose-binding protein is an important adhesin expressed on the surface membranes of pathogenic Acanthamoeba organisms. To specifically target Acanthamoeba, the overall aim of this study was to synthesize a photosensitizing compound (porphyrin) conjugated with mannose and test its efficacy in vitro. The synthesis of mannose-conjugated porphyrin was achieved by mixing benzaldehyde and pyrrole, yielding tetraphenylporphyrin. Tetraphenylporphyrin was then converted into mono-nitrophenylporphyrin by selectively nitrating the para position of the phenyl rings, as confirmed by nuclear magnetic resonance (NMR) spectroscopy. The mono-nitrophenylporphyrin was reduced to mono-aminophenylporphyrin in the presence of tin dichloride and confirmed by a peak at m/z 629. Finally, mono-aminoporphyrin was conjugated with mannose, resulting in the formation of an imine bond. Mannose-conjugated porphyrin was confirmed through spectroscopic analysis and showed that it absorbed light of wavelengths ranging from 425 to 475 nm. To determine the antiacanthamoebic effects of the derived product, amoebae were incubated with mannose-conjugated porphyrin for 1 h and washed 3 times to remove extracellular compound. Next, the amoebae were exposed to light of the appropriate wavelength for 1 h. The results revealed that mannose-conjugated porphyrin produced potent trophicidal effects and blocked excystation. In contrast, Acanthamoeba castellanii incubated with mannose alone and porphyrin alone did not exhibit an antiamoebic effect

  18. Photochemotherapeutic Strategy against Acanthamoeba Infections

    PubMed Central

    Aqeel, Yousuf; Siddiqui, Ruqaiyyah; Anwar, Ayaz; Shah, Muhammad Raza; Khoja, Shahrukh

    2015-01-01

    Acanthamoeba is a protist pathogen that can cause serious human infections, including blinding keratitis and a granulomatous amoebic encephalitis that almost always results in death. The current treatment for these infections includes a mixture of drugs, and even then, a recurrence can occur. Photochemotherapy has shown promise in the treatment of Acanthamoeba infections; however, the selective targeting of pathogenic Acanthamoeba has remained a major concern. The mannose-binding protein is an important adhesin expressed on the surface membranes of pathogenic Acanthamoeba organisms. To specifically target Acanthamoeba, the overall aim of this study was to synthesize a photosensitizing compound (porphyrin) conjugated with mannose and test its efficacy in vitro. The synthesis of mannose-conjugated porphyrin was achieved by mixing benzaldehyde and pyrrole, yielding tetraphenylporphyrin. Tetraphenylporphyrin was then converted into mono-nitrophenylporphyrin by selectively nitrating the para position of the phenyl rings, as confirmed by nuclear magnetic resonance (NMR) spectroscopy. The mono-nitrophenylporphyrin was reduced to mono-aminophenylporphyrin in the presence of tin dichloride and confirmed by a peak at m/z 629. Finally, mono-aminoporphyrin was conjugated with mannose, resulting in the formation of an imine bond. Mannose-conjugated porphyrin was confirmed through spectroscopic analysis and showed that it absorbed light of wavelengths ranging from 425 to 475 nm. To determine the antiacanthamoebic effects of the derived product, amoebae were incubated with mannose-conjugated porphyrin for 1 h and washed 3 times to remove extracellular compound. Next, the amoebae were exposed to light of the appropriate wavelength for 1 h. The results revealed that mannose-conjugated porphyrin produced potent trophicidal effects and blocked excystation. In contrast, Acanthamoeba castellanii incubated with mannose alone and porphyrin alone did not exhibit an antiamoebic effect

  19. Pathogenic Free-Living Amoebae Isolated From Contact Lenses of Keratitis Patients

    PubMed Central

    HAJIALILO, Elham; NIYYATI, Maryam; SOLAYMANI, Mohammad; REZAEIAN, Mostafa

    2015-01-01

    Background: Free-living amoeba (FLA)-related keratitis is a progressive infection of the cornea with poor prognosis. The present study aimed to investigates the contact lenses of patients with keratitis for pathogenic free-living amoebae. Methods: Overall, 62 contact lenses and their paraphernalia of patients with keratitis cultured and tested for the presence of free-living amoebae using morphological criteria. Unusual plates including plates containing mix amoebae and Vermamoeba were submitted to molecular analysis. Results: Out of 62 plates, 11 revealed the outgrowth of free living amoeba of which 9 were Acanthamoeba, one plates contained mix amoebae including Acanthamoeba and Vermamoeba and one showed the presence of Vermamoeba. These two latter plates belonged to patients suffered from unilateral keratitis due to the misused of soft contact lenses. One of the patients had mix infection of Acanthamoeba (T4) and V. vermiformis meanwhile the other patient was infected with the V. vermiformis. Conclusion: Amoebic keratitis continues to rise in Iran and worldwide. To date, various genera of free-living amoebae such as Vermamoeba could be the causative agent of keratitis. Soft contact lens wearers are the most affected patients in the country, thus awareness of high-risk people for preventing free-living amoebae related keratitis is of utmost importance. PMID:26811719

  20. Recent Outbreaks of Atypical Contact Lens-Related Keratitis: What Have We Learned?

    PubMed Central

    Tu, Elmer Y.; Joslin, Charlotte E

    2010-01-01

    Purpose To examine the public health implications of two recent outbreaks of atypical contact lens-related infectious keratitis. Design A perspective based on the literature and authors’ experience. Results The contact lens-related fusarium and acanthamoeba keratitis outbreaks were both detected by dramatic rises seen in tertiary care centers in Singapore and the US, respectively. Case control studies in both outbreaks each were able to identify a strong association with the use of different contact lens disinfection solutions. Their respective recalls resulted in a steep decline of fusarium keratitis, but not of acanthamoeba keratitis. Early investigations into each solution association implicate components not directly related to their primary disinfectant, but the true pathogenesis remains unknown. However, the number of Acanthamoeba cases individually attributed to each of almost all available disinfection systems exceeds previously understood total US incidence, suggesting other risk factors. Current standards do not require demonstration of anti-acanthamoebal activity. Yet, despite the inclusion of fusarium in mandatory testing for solutions, current pre-market testing was not predictive of the outbreak. Conclusions The two recent outbreaks of atypical contact-lens related keratitis have reinforced the value of tertiary care eye care centers in detecting early rises in rare infections and the power of adaptable, well-designed epidemiologic investigations. While fusarium keratitis has significantly declined with the recall of Renu with Moistureloc, the persistence of Acanthamoeba keratitis begs fundamental changes in contact lens hygiene practices, inclusion of acanthamoeba as a test organism and contact lens disinfectant test regimens for all contact lens–related pathogens which are verifiably reflective of end user contact lens wear complications. PMID:21036209

  1. Identification of 18S ribosomal DNA genotype of Acanthamoeba from hot spring recreation areas in the central range, Taiwan

    NASA Astrophysics Data System (ADS)

    Hsu, Bing-Mu; Ma, Po-Hua; Liou, Tai-Sheng; Chen, Jung-Sheng; Shih, Feng-Cheng

    2009-04-01

    SummaryAcanthamoeba is a free-living amoebae ubiquitous to aquatic environments. Within the genus a few species are recognized as opportunistic potential human pathogens, which cause granulomatous amoebic encephalitis (GAE) and keratitis. Infections of keratitis are frequently reported through wearing lens while swimming in the non-disinfected aquatic environment. Contaminations in hot tubs, spas and public baths are also possible. As a result, in this study, we identified Acanthamoeba based on the PCR amplification with a genus-specific primer pair and investigated the distribution of Acanthamoeba at five hot spring recreation areas in central range, Taiwan. We gathered data on factors potentially associated with the pathogen's distribution, including various sampling sites, aquatic environment, physical and microbiological water quality parameters. Spring water was collected from 55 sites and Acanthamoeba was detected in 9 (16.4%). The most frequently detected was Acanthamoeba griffini, followed by Acanthamoeba jacobsi. Legionella were detected in 18 (32.7%) of the sites sampled in this study. The species of Legionella identified included Legionella pneumophila serotype 6, serotype 1, and Legionella erythra. Overall, 9.1% of the samples contained both Acanthamoeba and Legionella. The prevalence of Acanthamoeba was contrary to the levels of microbiological indicators recommended by Taiwan CDC, and no significant differences (Mann-Whitney U test, P < 0.05) were observed between the presence/absence of Acanthamoeba and water quality parameters. Results of this survey confirm the existence of Acanthamoeba in Taiwan spring recreation areas. Acanthamoeba, the organism responsible for the majority of Acanthamoeba keratitis and can serve as vehicles for facultative pathogens, should be considered a potential threat for health associated with human activities in spring recreation areas of Taiwan.

  2. Fungal keratitis presenting as radial keratoneuritis

    PubMed Central

    Kapoor, Aditya; Jain, Rajat; Sahu, Srikant; Sangwan, Virender

    2014-01-01

    A 44-year-old man presented with severe pain, redness, watering and photophobia for 10 days in the right eye without any history of trauma. Diagnosis of herpes simplex disciform keratitis was made and he was prescribed topical steroids. The patient showed clinical worsening and presented with ring infiltrate, diffuse stromal oedema and radial keratoneuritis, a finding pathognomic of acanthamoeba keratitis. With two inconclusive corneal scrapings and the patient showing clinical worsening, an urgent therapeutic penetrating keratoplasty was carried out. Histopathological and microbiological examination of the excised corneal button revealed the presence of fungus. At 5 weeks follow-up, the patient has best-corrected visual acuity 20/40 with no recurrence of infection. PMID:24717856

  3. Several staining techniques to enhance the visibility of Acanthamoeba cysts.

    PubMed

    El-Sayed, Nagwa Mostafa; Hikal, Wafaa Mohamed

    2015-03-01

    Acanthamoeba is one of the most common free-living amoebae. It is widespread in the environment and can infect humans causing keratitis. Delayed diagnosis or misdiagnosis leads to extensive corneal inflammation and profound visual loss. Therefore, accurate and rapid diagnosis of Acanthamoeba keratitis is essential for successful treatment and good prognosis. This study was designed to use different staining techniques to facilitate the identification of Acanthamoeba cysts. Acanthamoeba cysts were isolated by cultivation of either corneal scraping specimens or tap water samples onto non-nutrient agar plates seeded with Escherichia coli. Subcultures were done from positive cultures until unique cysts were isolated. Acanthamoeba cysts were stained temporarily using iodine, eosin, methylene blue, and calcofluor white (CFW) stains and as permanent slides after processing for mounting using modified trichrome, Gimenez and Giemsa staining. These stains were compared on the basis of staining quality including clarity of morphological details, differentiation between cytoplasm and nuclei, color and contrast, and also other characteristics of the staining techniques, including ease of handling, time taken for the procedure, and cost effectiveness. The cysts of Acanthamoeba were recognized in the form of double-walled cysts: the outer wall (ectocyst) that was being differentiated from the variably stained surrounding background and the inner wall (endocyst) that was sometimes stellated, polygonal, round, or oval and visualized as separate from the spherical, sometimes irregular, outline of the ectocyst. Regarding the temporary stains, it was found that they were efficient for visualizing the morphological details of Acanthamoeba cysts. In CFW staining, Acanthamoeba cysts appeared as bluish-white or turquoise oval halos although the internal detail was not evident. On the other hand, the results of permanent-stained slides showed the most consistent stain for identification of

  4. Several staining techniques to enhance the visibility of Acanthamoeba cysts.

    PubMed

    El-Sayed, Nagwa Mostafa; Hikal, Wafaa Mohamed

    2015-03-01

    Acanthamoeba is one of the most common free-living amoebae. It is widespread in the environment and can infect humans causing keratitis. Delayed diagnosis or misdiagnosis leads to extensive corneal inflammation and profound visual loss. Therefore, accurate and rapid diagnosis of Acanthamoeba keratitis is essential for successful treatment and good prognosis. This study was designed to use different staining techniques to facilitate the identification of Acanthamoeba cysts. Acanthamoeba cysts were isolated by cultivation of either corneal scraping specimens or tap water samples onto non-nutrient agar plates seeded with Escherichia coli. Subcultures were done from positive cultures until unique cysts were isolated. Acanthamoeba cysts were stained temporarily using iodine, eosin, methylene blue, and calcofluor white (CFW) stains and as permanent slides after processing for mounting using modified trichrome, Gimenez and Giemsa staining. These stains were compared on the basis of staining quality including clarity of morphological details, differentiation between cytoplasm and nuclei, color and contrast, and also other characteristics of the staining techniques, including ease of handling, time taken for the procedure, and cost effectiveness. The cysts of Acanthamoeba were recognized in the form of double-walled cysts: the outer wall (ectocyst) that was being differentiated from the variably stained surrounding background and the inner wall (endocyst) that was sometimes stellated, polygonal, round, or oval and visualized as separate from the spherical, sometimes irregular, outline of the ectocyst. Regarding the temporary stains, it was found that they were efficient for visualizing the morphological details of Acanthamoeba cysts. In CFW staining, Acanthamoeba cysts appeared as bluish-white or turquoise oval halos although the internal detail was not evident. On the other hand, the results of permanent-stained slides showed the most consistent stain for identification of

  5. Use of multiple immunosuppressive agents in recalcitrant ACANTHAMOEBA scleritis.

    PubMed

    Igras, Estera; Murphy, Conor

    2015-01-01

    A 48-year-old woman who is a contact lens wearer presented with unilateral ACANTHAMOEBA keratitis, confirmed by PCR, which responded initially to topical polyhexamethylene biguanide (PHMB) and brolene. Three months later, despite continued treatment, she developed diffuse anterior scleritis with severe pain and marked scleral injection but without evidence of recurrence keratitis. Oral non-steroidal anti-inflammatories and oral high-dose corticosteroids were added without success. Subsequent treatment with intravenous methylprednisolone and high-dose cyclosporine led to a temporary improvement. Re-presenting with signs of recurrent scleritis and severe pain, the antitumor necrosis factor monoclonal antibody adalimumab, and later oral cyclophosphamide, were added. This led to complete quiescence of the scleritis. Unfortunately, frequent recurrences of ACANTHAMOEBA keratitis and anterior uveitis occurred on immunosuppression requiring continued treatment with PHMB, brolene and topical corticosteroids. This is the first case of severe refractory ACANTHAMOEBA scleritis requiring the concomitant use of four immunosuppressive agents to achieve continued disease control. The challenges in managing this case are discussed.

  6. Statins and Voriconazole Induce Programmed Cell Death in Acanthamoeba castellanii

    PubMed Central

    López-Arencibia, Atteneri; Sifaoui, Ines; Reyes-Batlle, María; Valladares, Basilio; Martínez-Carretero, Enrique; Piñero, José E.; Maciver, Sutherland K.; Lorenzo-Morales, Jacob

    2015-01-01

    Members of the genus Acanthamoeba are facultative pathogens of humans, causing a sight-threatening keratitis and a life-threatening encephalitis. In order to treat those infections properly, it is necessary to target the treatment not only to the trophozoite but also to the cyst. Furthermore, it may be advantageous to avoid parasite killing by necrosis, which may induce local inflammation. We must also avoid toxicity of host tissue. Many drugs which target eukaryotes are known to induce programmed cell death (PCD), but this process is poorly characterized in Acanthamoeba. Here, we study the processes of programmed cell death in Acanthamoeba, induced by several drugs, such as statins and voriconazole. We tested atorvastatin, fluvastatin, simvastatin, and voriconazole at the 50% inhibitory concentrations (IC50s) and IC90s that we have previously established. In order to evaluate this phenomenon, we investigated the DNA fragmentation, one of the main characteristics of PCD, with quantitative and qualitative techniques. Also, the changes related to phosphatidylserine exposure on the external cell membrane and cell permeability were studied. Finally, because caspases are key to PCD pathways, caspase activity was evaluated in Acanthamoeba. All the drugs assayed in this study induced PCD in Acanthamoeba. To the best of our knowledge, this is the first study where PCD induced by drugs is described quantitatively and qualitatively in Acanthamoeba. PMID:25733513

  7. Thermotolerant Acanthamoeba spp. isolated from therapeutic hot springs in Northwestern Iran.

    PubMed

    Solgi, Rahmat; Niyyati, Maryam; Haghighi, Ali; Taghipour, Niloofar; Tabaei, Seyyed Javad Seyyed; Eftekhar, Mohamad; Nazemalhosseini Mojarad, Ehsan

    2012-12-01

    This study was conducted to address the distribution of Acanthamoeba genotypes in therapeutic hot springs in Iran. Sixty water and sediment samples were collected from bicarbonate, sulphur, and sodium chloride thermal springs in the northwest. All hot springs examined are used mainly for health purposes in Iran. Acanthamoeba were identified by both morphology and PCR (polymerase chain reaction). Genotype identification was based on the sequencing of a highly variable and informative region of Diagnostic Fragment 3 (stem 29-1 of 18S rRNA gene) within Acanthamoeba-specific amplimer (ASA.S1). Twenty percent of hot springs were contaminated with thermotolerant Acanthamoeba belonging to the potentially pathogenic T4 and T3 genotypes. A high number (91.7%) of strains showed growth at 37 °C, and eight isolates showed growth at 42 °C. A single isolate (HSNW2) was detected in waters at 70 °C. The presence of thermotolerant Acanthamoeba highlights a risk factor for susceptible individuals, as Acanthamoeba-related keratitis continues to rise in Iran. Periodic surveillance of thermal waters as well as improved filtration and disinfection is recommended to prevent disease related to pathogenic Acanthamoeba. This is the first comprehensive molecular study of Acanthamoeba genotypes in hot springs in Iran and the first to report the occurrence of the T3 genotype (corresponding to Acanthamoeba griffini) in thermal water sources in this country.

  8. Morpho-Physiological and Biochemical Criteria of Acanthamoeba spp. Isolated from the Egyptian Aquatic Environment

    PubMed Central

    Al-Herrawy, A; Bahgat, M; Mohammed, A; Ashour, A; Hikal, W

    2013-01-01

    Background The free-living amoebae Acanthamoeba spp., have been recognized as etiologic agents of amoebic encephalitis, keratitis, otitis, lung lesions and other skin infections mainly in immuno-compromised individuals. In this study, morpho-physiological and biochemical characterization of Acanthamoeba strains isolated from the Egyptian aquatic environment were surveyed. Methods Some Acanthamoeba species were cultivated on non-nutrient agar. Isolated strains of Acanthamoeba were identification based on the morphology of trophic and cyst forms in addition to temperature and osmo-tolerance assays. Biochemical characterization of the isolated amoeba strains was performed using quantitative assay as well as qualitative determination of proteolytic activity in zymograph analysis. Results Potentially pathogenic Acanthamoeba species were isolated from all of the examined water sources. Colorimetric assays showed protease activity in the heat-tolerant isolates of Acanthamoeba. All pathogenic isolates of Acanthamoeba exhibited higher protease activity than did the non-pathogenic ones. The zymographic protease assays showed various banding patterns for different strains of Acanthamoeba. Conclusion The incidence and prevalence of the pathogenic Acanthamoeba species in the aquatic environment using parasitological and biochemical diagnostic tools will provide baseline data against which the risk factors associated with waterborne transmission can be identified. PMID:23914245

  9. Acanthamoeba DNA can be directly amplified from corneal scrapings.

    PubMed

    El-Sayed, Nagwa Mostafa; Younis, Mohamed Saad; Elhamshary, Azza Mohamed; Abd-Elmaboud, Amina Ibrahim; Kishik, Shereen Magdy

    2014-09-01

    This study evaluated the performance of direct amplification of Acanthamoeba-DNA bypassing DNA extraction in the diagnosis of Acanthamoeba keratitis in clinically suspected cases in comparison to direct microscopic examination and in vitro culture. Corneal scrapings were collected from 110 patients who were clinically suspected to have Acanthamoeba keratitis, 63 contact lens wearers (CLW), and 47 non-contact lens wearers (NCLW). Taken samples were subjected to direct microscopic examination, cultivation onto the non-nutrient agar plate surface seeded with Escherichia coli, and PCR amplification. The diagnostic performance of these methods was statistically compared. The results showed that Acanthamoeba infection was detected in 21 (19.1%) of clinically suspected cases (110); 17 (81%) of them were CLW and the remaining 4 (19%) positive cases were NCLW. Regarding the used diagnostic methods, it was found that direct amplification of Acanthamoeba DNA bypassing nucleic acid extraction was superior to microscopy and culture in which 21 cases (19.1%) were positive for Acanthamoeba by PCR compared to 19 positive cases by culture (17.3%) and one case (0.9%) by direct smear. The difference in detection rates between culture and direct smear was highly statistically significant (P = 0.001). On the other hand, there was no significant difference in detection rates between culture and PCR (P = 0.86). On using culture as the gold standard, PCR showed three false-positive samples that were negative by culture and one false-negative sample that was positive by culture. At the same time, direct smear showed 18 false-negative samples. The sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy of PCR were 94.7, 96.7, 85.7, 98.9, and 96.4, respectively, while those of direct smear were 5.3, 100, 100, 83.5, and 83.6, respectively. In conclusion, direct amplification of Acanthamoeba-DNA bypassing DNA extraction is a reliable

  10. Lethal Effects of Helianthemum lippii (L.) on Acanthamoeba castellanii Cysts in Vitro

    PubMed Central

    Badria, F.A.; Hetta, M.H.; Sarhan, Rania M.; Ezz El-Din, M.H.

    2014-01-01

    Acanthamoeba spp. commonly cause Acanthamoeba keratitis which is typically associated with the wear of contact lenses. Therefore, finding an economic, efficient, and safe therapy of natural origin is of outmost importance. This study examined the in vitro lethal potential of ethyl acetate and methanol extracts of Helianthemum lippii (L.) (sun roses) against Acanthamoeba castellanii cysts isolated from patients with amoebic keratitis. Both extracts proved to be potent as regard to their lethal effects on A. castellanii cysts with comparable results to chlorhexidine. The ethyl acetate was more promising with cumulative lethality. It showed a highly significant lethal percentage along the duration of treatment. The analysis of the more potent ethyl acetate extract revealed the presence of 2.96 mg/100 g of total phenolics, 0.289 mg/100 ml of total flavonoids and 37 mg/100 mg of total tannins which highlighted their phytomedicinal role. PMID:25031463

  11. In Vitro Effectiveness of Povidone-Iodine on Acanthamoeba Isolates from Human Cornea

    PubMed Central

    Gatti, Simonetta; Cevini, Claudia; Bruno, Antonella; Penso, Gabriella; Rama, Paolo; Scaglia, Massimo

    1998-01-01

    Acanthamoeba keratitis is a severe ocular infection secondary to accidental macro- or microscopic trauma of the cornea. Starting in 1985, a dramatic increase of this infection was recorded along with the spread of contact lens use. This protozoal disease is difficult to treat because of the scarcity of efficacious topical and systemic drugs. We evaluated the in vitro effectiveness of povidone-iodine (PVP-I [Betadine]), an agent with broad antibacterial and antiviral activity, compared to that of chlorhexidine (CXD), a cationic antiseptic, on Acanthamoeba isolates from patients with amebic keratitis. The results showed that PVP-I solution from 0.5 to 2.5% has a better antiamebic activity both on trophic and cystic stages of Acanthamoeba spp. than does CXD. PMID:9736540

  12. Fluorescent labeling of Acanthamoeba assessed in situ from corneal sectioned microscopy

    PubMed Central

    Marcos, Susana; Requejo-Isidro, Jose; Merayo-Lloves, Jesus; Acuña, A. Ulises; Hornillos, Valentin; Carrillo, Eugenia; Pérez-Merino, Pablo; del Olmo-Aguado, Susana; del Aguila, Carmen; Amat-Guerri, Francisco; Rivas, Luis

    2012-01-01

    Acanthamoeba keratitis is a serious pathogenic corneal disease, with challenging diagnosis. Standard diagnostic methods include corneal biopsy (involving cell culture) and in vivo reflection corneal microscopy (in which the visualization of the pathogen is challenged by the presence of multiple reflectance corneal structures). We present a new imaging method based on fluorescence sectioned microscopy for visualization of Acanthamoeba. A fluorescent marker (MT-11-BDP), composed by a fluorescent group (BODIPY) inserted in miltefosine (a therapeutic agent against Acanthamoeba), was developed. A custom-developed fluorescent structured illumination sectioned corneal microscope (excitation wavelength: 488 nm; axial/lateral resolution: 2.6 μm/0.4-0.6 μm) was used to image intact enucleated rabbit eyes, injected with a solution of stained Acanthamoeba in the stroma. Fluorescent sectioned microscopic images of intact enucleated rabbit eyes revealed stained Acanthamoeba trophozoites within the stroma, easily identified by the contrasted fluorescent emission, size and shape. Control experiments show that the fluorescent maker is not internalized by corneal cells, making the developed marker specific to the pathogen. Fluorescent sectioned microscopy shows potential for specific diagnosis of Acanthamoeba keratitis. Corneal confocal microscopy, provided with a fluorescent channel, could be largely improved in specificity and sensitivity in combination with specific fluorescent marking. PMID:23082290

  13. Quick survey for detection, identification and characterization of Acanthamoeba genotypes from some selected soil and water samples across Pakistan.

    PubMed

    Tanveer, Tania; Hameed, Abdul; Gul, Asma; Matin, Abdul

    2015-01-01

    Acanthamoeba is an opportunistic protozoan pathogen which is widely distributed in nature and plays a pivotal role in ecosystem. Acanthamoeba species may cause blinding keratitis and fatal granulomatous encephalitis involving central nervous system. In this study, we investigated the presence of Acanthamoeba in soil and water resources of Pakistan. Here, Acanthamoeba were recovered on non-nutrient agar plate lawn with E. coli and identified by morphological characteristics of the cyst. Furthermore PCR was performed with genus-specific primers followed by direct sequencing of the PCR product for molecular identification. Overall our PCR and sequencing results confirmed pathogenic genotypes including T4 and T15 from both soil and water samples. This is our first report of Acanthamoeba isolation from both soil and water resources of Pakistan which may serve as a potential treat to human health across the country.

  14. In Vitro Efficacy of Corifungin against Acanthamoeba castellanii Trophozoites and Cysts

    PubMed Central

    Debnath, Anjan; Tunac, Josefino B.; Silva-Olivares, Angélica; Galindo-Gómez, Silvia

    2014-01-01

    Painful blinding keratitis and fatal granulomatous amebic encephalitis are caused by the free-living amebae Acanthamoeba spp. Several prescription eye medications are used to treat Acanthamoeba keratitis, but the infection can be difficult to control because of recurrence of infection. For the treatment of encephalitis, no single drug was found useful, and in spite of the use of a combination of multiple drugs, the mortality rate remains high. Therefore, efficient, novel drugs are urgently needed for the treatment of amebic keratitis and granulomatous amebic encephalitis. In this study, we identified corifungin, a water-soluble polyene macrolide, as amebicidal. In vitro, it was effective against both the trophozoites and the cysts. Transmission electron microscopy of Acanthamoeba castellanii incubated with corifungin showed the presence of swollen mitochondria, electron-dense granules, degeneration of cytoplasm architecture, and loss of nuclear chromatin structure. These changes were followed by lysis of amebae. Corifungin also induced the encystment process of A. castellanii. There were alterations in the cyst cell wall followed by lysis of the cysts. Corifungin is a promising therapeutic option for keratitis and granulomatous amebic encephalitis. PMID:24366747

  15. Artemether Exhibits Amoebicidal Activity against Acanthamoeba castellanii through Inhibition of the Serine Biosynthesis Pathway

    PubMed Central

    Deng, Yihong; Ran, Wei; Man, Suqin; Li, Xueping; Gao, Hongjian; Tang, Wei

    2015-01-01

    Acanthamoeba sp. parasites are the causative agents of Acanthamoeba keratitis, fatal granulomatous amoebic encephalitis, and cutaneous infections. However, there are currently no effective drugs for these organisms. Here, we evaluated the activity of the antimalarial agent artemether against Acanthamoeba castellanii trophozoites and identified potential targets of this agent through a proteomic approach. Artemether exhibited in vitro amoebicidal activity in a time- and dose-dependent manner and induced ultrastructural modification and cell apoptosis. The iTRAQ quantitative proteomic analysis identified 707 proteins that were differentially expressed after artemether treatment. We focused on phosphoglycerate dehydrogenase and phosphoserine aminotransferase in the serine biosynthesis pathway because of their importance to the growth and proliferation of protozoan and cancer cells. The expression of these proteins in Acanthamoeba was validated using quantitative real-time PCR and Western blotting after artemether treatment. The changes in the expression levels of phosphoserine aminotransferase were consistent with those of phosphoglycerate dehydrogenase. Therefore, the downregulation of phosphoserine aminotransferase may be due to the downregulation of phosphoglycerate dehydrogenase. Furthermore, exogenous serine might antagonize the activity of artemether against Acanthamoeba trophozoites. These results indicate that the serine biosynthesis pathway is important to amoeba survival and that targeting these enzymes would improve the treatment of Acanthamoeba infections. Artemether may be used as a phosphoglycerate dehydrogenase inhibitor to control or block Acanthamoeba infections. PMID:26014935

  16. Isolation and Genotyping of Acanthamoeba spp. as Neglected Parasites in North of Iran

    PubMed Central

    Shokri, Azar; Sarvi, Shahabeddin; Daryani, Ahmad; Sharif, Mehdi

    2016-01-01

    Acanthamoeba, a free-living amoeba, is widely distributed in the environment, water sources, soil, dust, and air. It can cause keratitis in contact lens wearers with poor hygiene and also fatal granulomatous amebic encephalitis (GAE) in immunocompromised hosts. The aim of this study was to gain some insights into the distribution and genotypes of the potentially pathogenic species of Acanthamoeba present in water sources in north of Iran. Total 43 Acanthamoeba species were isolated from 77 water samples taken from different water sources within the Mazandaran province in Northern Iran (Sari city and suburbs). Isolates were identified based on cyst and trophozoite morphological characteristics as well genetics. PCR fragments corresponding to the small-subunit 18S rRNA gene were sequenced for 20 of 43 positive isolates. The results revealed that 83.3% of sequenced isolates belonged to the T4 genotype and the rest belonged to the T2 genotype. Our results indicated that Acanthamoeba is widely distributed in Sari city. As the incidence in Iran of amoebic keratitis has increased in recent years, the exact estimation of the prevalence of this amoeba and its predominant genotype may play a crucial role in prevention of the disease. Sari city has several rivers, seashores, and natural recreational amenities, which attract visitors during the year. This is the first report of Acanthamoeba genotypes from water sources in Sari city, Mazandaran province of Iran, and the results suggest that more attention is needed to protect the visiting population and immunocompromised individuals. PMID:27658596

  17. Isolation and Genotyping of Acanthamoeba spp. as Neglected Parasites in North of Iran.

    PubMed

    Shokri, Azar; Sarvi, Shahabeddin; Daryani, Ahmad; Sharif, Mehdi

    2016-08-01

    Acanthamoeba, a free-living amoeba, is widely distributed in the environment, water sources, soil, dust, and air. It can cause keratitis in contact lens wearers with poor hygiene and also fatal granulomatous amebic encephalitis (GAE) in immunocompromised hosts. The aim of this study was to gain some insights into the distribution and genotypes of the potentially pathogenic species of Acanthamoeba present in water sources in north of Iran. Total 43 Acanthamoeba species were isolated from 77 water samples taken from different water sources within the Mazandaran province in Northern Iran (Sari city and suburbs). Isolates were identified based on cyst and trophozoite morphological characteristics as well genetics. PCR fragments corresponding to the small-subunit 18S rRNA gene were sequenced for 20 of 43 positive isolates. The results revealed that 83.3% of sequenced isolates belonged to the T4 genotype and the rest belonged to the T2 genotype. Our results indicated that Acanthamoeba is widely distributed in Sari city. As the incidence in Iran of amoebic keratitis has increased in recent years, the exact estimation of the prevalence of this amoeba and its predominant genotype may play a crucial role in prevention of the disease. Sari city has several rivers, seashores, and natural recreational amenities, which attract visitors during the year. This is the first report of Acanthamoeba genotypes from water sources in Sari city, Mazandaran province of Iran, and the results suggest that more attention is needed to protect the visiting population and immunocompromised individuals. PMID:27658596

  18. [Acanthamoeba spp. as opportunistic pathogens parasites].

    PubMed

    Castrillón, Juan C; Orozco, Lina P

    2013-04-01

    Among free-living amoeba in nature, species of the genus Acanthamoeba have been associated with human disease. These amoeba are among the most abundant protozoa in nature due to its cosmopolitan distribution and are able to survive in a wide variety of habitats because its low demand for food and in harsh environments by forming structures known as cysts. However, ecological changes and incursion of its different habitats have made this organism can invade a host and live as parasites within him. That's why this type of protozoa are known as amphizoic organism, because human can be constituted as its host, causing infections in the central nervous system, disseminated infections in skin and lungs, and keratitis. Thus, 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. This review summarizes what is known of this kind of free-living amoeba, focusing on the biology, ecology, pathogenesis, diagnosis, treatment and human defense mechanism against infection by the amoeba.

  19. [Acanthamoeba spp. as opportunistic pathogens parasites].

    PubMed

    Castrillón, Juan C; Orozco, Lina P

    2013-04-01

    Among free-living amoeba in nature, species of the genus Acanthamoeba have been associated with human disease. These amoeba are among the most abundant protozoa in nature due to its cosmopolitan distribution and are able to survive in a wide variety of habitats because its low demand for food and in harsh environments by forming structures known as cysts. However, ecological changes and incursion of its different habitats have made this organism can invade a host and live as parasites within him. That's why this type of protozoa are known as amphizoic organism, because human can be constituted as its host, causing infections in the central nervous system, disseminated infections in skin and lungs, and keratitis. Thus, 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. This review summarizes what is known of this kind of free-living amoeba, focusing on the biology, ecology, pathogenesis, diagnosis, treatment and human defense mechanism against infection by the amoeba. PMID:23677153

  20. Microbial keratitis in Hong Kong: relationship to climate, environment and contact-lens disinfection.

    PubMed

    Houang, E; Lam, D; Fan, D; Seal, D

    2001-01-01

    Microbial keratitis has been studied in Hong Kong as a representative sub-tropical climate of south China. An 18-month investigation in 1997/98 of 223 cases of ulcerative keratitis (presumed microbial) was conducted in the 2 million population of Shatin and Kowloon at the Prince of Wales and Hong Kong Eye Hospitals respectively with comprehensive microbiology. A case-control study was pursued at the same time between 45 contact-lens wearers (CLW) developing microbial keratitis and 135 lens-wearing volunteers matched for age, sex, educational status and visual acuity. Home water supplies were sampled for Acanthamoeba. Previous ocular surface disease and trauma (preventable by wearing goggles for grinding) were common predisposing causes while cosmetic wear of contact lenses was responsible for 26% of cases overall. Pseudomonas aeruginosa was the commonest bacterium isolated, from both CLW and non-CLW, with infection being acquired within the community. These 28 pseudomonads remained fully sensitive to the third-generation cephalosporins, aminoglycosides and quinolone antibiotics, which is very encouraging. Fungi were isolated, predominantly Fusarium sp., but less commonly than expected. A fungal/bacterial ratio was obtained of 1/17, while in comparison, the expected ratio for a tropical climate ranges from 1/5 (Singapore) to 1/2 (South India). Acanthamoeba was the second commonest microbe isolated from keratitis of CLW. The domestic water environment of 8% of homes of both patients and controls wearing contact lenses was colonized with Acanthamoeba. Lack of hygiene, use of tap water for storing lenses, failure to air-dry lens-storage cases or use of one-step hydrogen peroxide disinfectant were identified as risk factors for keratitis in CLW. The study results commend use of multipurpose solutions by CLW in Hong Kong to achieve the lowest expected rates of infection.

  1. [Peripheral ulcerative keratitis].

    PubMed

    Stamate, Alina-cristina; Avram, Corina Ioana; Malciolu, R; Oprea, S; Zemba, M

    2014-01-01

    Ulcerative keratitis is frequently associated with collagen vascular diseases and presents a predilection for peripheral corneal localization, due to the distinct morphologic and immunologic features of the limbal conjunctiva, which provides access for the circulating immune complexes to the peripheral cornea via the capillary network. Deposition of immune complexes in the terminal ends of limbal vessels initiates an immune-mediated vasculitis process, with inflammatory cells and mediators involvement by alteration of the vascular permeability. Peripheral ulcerative keratitis generally correlates with exacerbations of the background autoimmune systemic disease. Associated sceritis, specially the necrotizing form, is usually observed in severe cases, which may evolve in corneal perforation and loss of vision. Although the first-line of treatment in acute phases is represented by systemic administration of corticosteroids, immunosuppressive and cytotoxic agents are necessary for the treatment of peripheral ulcerative keratitis associated with systemic diseases.

  2. Tarantula hair keratitis.

    PubMed

    Mangat, Simran Singh; Newman, Bill

    2012-10-26

    We describe a 12-year-old boy in England with keratitis secondary to tarantula hairs embedded within the stroma of his cornea. Every attempt must be made to isolate these hairs at the first visit as they have a barbed nature and have a propensity to propagate through ocular tissues. A chronic keratitis requiring long-term steroid use may result if hairs persist in the cornea. Children who keep tarantulas as pets should be instructed on safe handling to prevent the tarantula from adopting defence mechanisms and shedding their hairs. PMID:23242405

  3. Acanthamoeba spp. in domestic tap water in houses of contact lens wearers in the metropolitan area of Mexico City.

    PubMed

    Bonilla-Lemus, Patricia; Ramírez-Bautista, Gerardo A; Zamora-Muñoz, Claudia; Ibarra-Montes, María Del Rocío; Ramírez-Flores, Elizabeth; Hernández-Martínez, María Dolores

    2010-09-01

    A survey was carried out in the metropolitan area of Mexico City to determine the presence of Acanthamoeba in the tap water of houses of contact lens wearers. Water samples were taken from the mains water entry, bathroom sinks and storage containers (roof tanks, cisterns) of 27 houses; and from the solution contained in the contact lens cases. Samples were filtered and cultured onto NNE medium. The isolates were identified based on their morphological features and pathogenicity. Total and fecal coliforms, water temperature, pH, dissolved oxygen and residual free-chlorine were measured by standard methods. Forty five isolates of Acanthamoeba from 200 water samples were obtained. The highest number of amoebae was isolated from cisterns and roof tanks. Most Acanthamoeba isolates were non-pathogenic, however, their presence in tap water is a potential hazard since some species can cause Acanthamoeba keratitis and granulomatous amoebic encephalitis. PMID:19995560

  4. Acanthamoeba genotypes T2, T4, and T11 in soil sources from El Hierro island, Canary Islands, Spain.

    PubMed

    Reyes-Batlle, María; Zamora-Herrera, Jonadab; Vargas-Mesa, Alejandro; Valerón-Tejera, Marco Antonio; Wagner, Carolina; Martín-Navarro, Carmen Ma; López-Arencibia, Atteneri; Sifaoui, Ines; Martínez-Carretero, Enrique; Valladares, Basilio; Piñero, José E; Lorenzo-Morales, Jacob

    2016-08-01

    The genus Acanthamoeba includes pathogenic strains which are causative agents of keratitis and encephalitis that often may end fatal in humans and other animals. In the present study, forty soil samples were collected in the island of El Hierro, Canary Islands, Spain, and checked for the presence of Acanthamoeba. Samples were cultivated onto 2 % non-nutrient agar plates seeded with a layer of heat killed Escherichia coli. Amplification by PCR and sequencing of the DF3 region of the 18S rDNA of Acanthamoeba was carried out in order to confirm morphological identification of the amoebae. Furthermore, Acanthamoeba spp. was isolated from 47.5 % of soil samples. Moreover, genotypes T2, T4, and T11 were identified in these samples. To the best of our knowledge, this is the first study to establish genotypes T2, T4, and T11 in soil sources from El Hierro island.

  5. Acanthamoeba spp. in domestic tap water in houses of contact lens wearers in the metropolitan area of Mexico City.

    PubMed

    Bonilla-Lemus, Patricia; Ramírez-Bautista, Gerardo A; Zamora-Muñoz, Claudia; Ibarra-Montes, María Del Rocío; Ramírez-Flores, Elizabeth; Hernández-Martínez, María Dolores

    2010-09-01

    A survey was carried out in the metropolitan area of Mexico City to determine the presence of Acanthamoeba in the tap water of houses of contact lens wearers. Water samples were taken from the mains water entry, bathroom sinks and storage containers (roof tanks, cisterns) of 27 houses; and from the solution contained in the contact lens cases. Samples were filtered and cultured onto NNE medium. The isolates were identified based on their morphological features and pathogenicity. Total and fecal coliforms, water temperature, pH, dissolved oxygen and residual free-chlorine were measured by standard methods. Forty five isolates of Acanthamoeba from 200 water samples were obtained. The highest number of amoebae was isolated from cisterns and roof tanks. Most Acanthamoeba isolates were non-pathogenic, however, their presence in tap water is a potential hazard since some species can cause Acanthamoeba keratitis and granulomatous amoebic encephalitis.

  6. Comparative analyses of different genetic markers for the detection of Acanthamoeba spp. isolates.

    PubMed

    Derda, Monika; Wojtkowiak-Giera, Agnieszka; Hadaś, Edward

    2014-09-01

    Acanthamoeba are widespread free-living amoebae which may cause granulomatous amoebic encephalitis (GAE), keratitis, skin ulcerations and disseminated tissue infection. An important diagnostic and prognostic factor for the treatment of infection is a quick and correct diagnosis of amoebae strains. The aim of our study was to develop a rapid method for detection and identification of pathogenic Acanthamoeba spp. strains from diagnostic material collected from water. In this study we analysed five amplification-based genetic markers (Aca 16S, Ac6/210, GP, JDP, Nelson) used for identification of pathogenic Acanthamoeba spp. strains isolated in water sources in Poland, Iceland and Sweden. Our results demonstrated the presence of pathogenic Acanthamoeba strains in tap water. PCR assay appeared to be a more rapid and sensitive method to detect the presence of amoebae than the limited conventional techniques. Based on our observations, we can confirm that the use of four out of five genetic markers (Aca 16S, Ac 6/210, JDP, GP, Nelson) may be helpful in identification of Acanthamoeba spp. strains, but only one Aca 16S primer pair is a highly specific marker that distinguishes between pathogenic strains of Acanthamoeba and other free-living amoeba families.

  7. Immunity to Acanthamoeba.

    PubMed

    Ferrante, A

    1991-01-01

    Human serum contains antibodies, mainly of the IgM and IgG isotypes, to pathogenic species of Acanthamoeba. This, as well as the capacity of these amebas to activate complement via the alternative pathway, may be a first-line defense against acanthamoeba infections in humans. Both antibody and complement appear to be important in promoting recognition of these amebas by phagocytic cells such as neutrophils. However, killing of amebas by neutrophils is dependent on lymphokine/monokine priming of the neutrophil. This priming augments the respiratory-burst activity and release of lysosomal enzymes of neutrophils in their response to the ameba. The products of the oxygen-dependent respiratory burst appear to be of prime importance in the killing of this free-living ameba. Antibodies also may prevent tissue invasion by Acanthamoeba by inhibiting its adherence, phagocytic activity, and migration and by neutralizing cytopathogenic amebic agents. Studies on experimental Acanthamoeba infections in mice showed marked species and strain specificity with regard to induction of protection with amebic antigens. Immune compromise or, alternatively, invasion at unique body sites in healthy individuals may form the basis for human infection with Acanthamoeba. PMID:2047675

  8. Identification of Unusual Phospholipid Fatty Acyl Compositions of Acanthamoeba castellanii

    PubMed Central

    Palusinska-Szysz, Marta; Kania, Magdalena; Turska-Szewczuk, Anna; Danikiewicz, Witold; Russa, Ryszard; Fuchs, Beate

    2014-01-01

    Acanthamoeba are opportunistic protozoan pathogens that may lead to sight-threatening keratitis and fatal granulomatous encephalitis. The successful prognosis requires early diagnosis and differentiation of pathogenic Acanthamoeba followed by aggressive treatment regimen. The plasma membrane of Acanthamoeba consists of 25% phospholipids (PL). The presence of C20 and, recently reported, 28- and 30-carbon fatty acyl residues is characteristic of amoeba PL. A detailed knowledge about this unusual PL composition could help to differentiate Acanthamoeba from other parasites, e.g. bacteria and develop more efficient treatment strategies. Therefore, the detailed PL composition of Acanthamoeba castellanii was investigated by 31P nuclear magnetic resonance spectroscopy, thin-layer chromatography, gas chromatography, high performance liquid chromatography and liquid chromatography-mass spectrometry. Normal and reversed phase liquid chromatography coupled with mass spectrometric detection was used for detailed characterization of the fatty acyl composition of each detected PL. The most abundant fatty acyl residues in each PL class were octadecanoyl (18∶0), octadecenoyl (18∶1 Δ9) and hexadecanoyl (16∶0). However, some selected PLs contained also very long fatty acyl chains: the presence of 28- and 30-carbon fatty acyl residues was confirmed in phosphatidylethanolamine (PE), phosphatidylserine, phosphatidic acid and cardiolipin. The majority of these fatty acyl residues were also identified in PE that resulted in the following composition: 28∶1/20∶2, 30∶2/18∶1, 28∶0/20∶2, 30∶2/20∶4 and 30∶3/20∶3. The PL of amoebae are significantly different in comparison to other cells: we describe here for the first time unusual, very long chain fatty acids with Δ5-unsaturation (30∶35,21,24) and 30∶221,24 localized exclusively in specific phospholipid classes of A. castellanii protozoa that could serve as specific biomarkers for the presence of these

  9. Evaluation of the immunodiagnostic potential of a recombinant surface protein domain from Acanthamoeba castellanii.

    PubMed

    Sánchez, Alemao G Carpinteyro; Virginio, Veridiana Gomes; Maschio, Vinicius José; Ferreira, Henrique Bunselmeyer; Rott, Marilise Brittes

    2016-10-01

    Acanthamoeba spp. are free-living protists widely distributed in environment, able to cause keratitis, encephalitis and skin lesions in humans and animals. Acanthamoeba spp. exist in two forms: an infective trophozoite and a dormant cyst. Several factors contribute to the pathogenesis of Acanthamoeba spp. The parasite adhesion to the host cell is the primary step for infection and is mediated by a mannose binding-protein, expressed in the surface and considered the main pathogenicity factor in Acanthamoeba spp. So far, there was no evidence of another surface protein of Acanthamoeba spp. relevant for host invasion or infection by these organisms. The aims of this study were to identify and characterize an Acanthamoeba castellanii surface protein and to evaluate its diagnostic potential. In silico predictions of surface proteins allowed to identify the A. castellanii calreticulin as a possible surface antigen. The coding sequence of a predicted extracellular domain of A. castellanii calreticulin was cloned by in vivo homologous recombination and the recombinant polypeptide (AcCRT29-130) was produced. Its immunodiagnostic potential was assessed in a recombinant antigen-based ELISA with sera from experimentally infected rats that developed keratitis and encephalitis, and sera from patients with encephalitis. The AcCRT29-130 was significantly more recognized by sera from encephalitis infected rats in comparison with the non-infected controls. Human sera from encephalitis patients, however presented no significant response. These results showed the AcCRT29-130 potential for A. castellanii infection immunodiagnosis in animals, with further studies being required for assessment of its use for human infections.

  10. Viability and morphological changes of Acanthamoeba spp. cysts after treatment with Effective microorganisms (EM).

    PubMed

    Sampaotong, Tanitta; Lek-Uthai, Usa; Roongruangchai, Jantima; Roongruangchai, Kosol

    2016-06-01

    Acanthamoeba is a free-living opportunistic protozoan parasite that is found in diverse environments. It can cause keratitis, mostly related to inappropriate use of contact lenses, as well as life threatening diseases including encephalitis, disseminated sinusitis, and skin ulcers. This study investigated morphological changes and fine structures of the cyst form of Acanthamoeba spp. after treatment with effective microorganisms (EM™) using light and scanning electron microscopies. Acanthamoeba cysts treated with 1:2, 1:4, 1:6, and undiluted EM™ showed higher percentages of non-viable cysts than those treated with 1:8, 1:10, 1:100, 1:200, and 1:400 EM™ and at 5 days post-treatment developed from cystic stage to trophozoite stage. Acanthamoeba cysts treated at concentrations of 1:2, 1:4, 1:6, and undiluted EM™ exhibited cytoplasmic clumping and shrinkage of amoeba cells away from cyst walls. The effective EM™ concentration lethal to Acanthamoeba spp. cyst could provide information to monitor the environmental control system. PMID:27413306

  11. Isolation and genotyping of acanthamoeba strains from soil sources from Jamaica, West Indies.

    PubMed

    Todd, Cheridah D; Reyes-Batlle, María; Martín-Navarro, Carmen Ma; Dorta-Gorrín, Alexis; López-Arencibia, Atteneri; Martínez-Carretero, Enrique; Piñero, José E; Valladares, Basilio; Lindo, John F; Lorenzo-Morales, Jacob

    2015-01-01

    Acanthamoeba spp. are opportunistic pathogens that are ubiquitous in nature. Many species of this genus are responsible for a fatal encephalitis and keratitis in humans and other animals. Seventy-two soil samples were collected from the parishes across Jamaica and assessed for the presence of Acanthamoeba spp. Cultivation was carried out on non-nutrient agar plates seeded with heat killed Escherichia coli. PCR and sequencing of the DF3 region were carried out in order to genotype the isolated strains of Acanthamoeba. Thermotolerance and osmotolerance assays were utilized to investigate the pathogenic potential of the Acanthamoeba isolates. Acanthamoeba spp. was isolated from 63.9% of soil samples. Sequencing of the DF3 region of the 18S rDNA resulted in the identification of genotypes T4, T5, and T11. T4 genotype was most frequently isolated. Most isolates were thermotolerant or both thermotolerant and osmotolerant, indicating that they may present the potential to cause disease in humans and other animals.

  12. Pathogenic and opportunistic free-living amoebae: Acanthamoeba spp., Balamuthia mandrillaris, Naegleria fowleri, and Sappinia diploidea.

    PubMed

    Visvesvara, Govinda S; Moura, Hercules; Schuster, Frederick L

    2007-06-01

    Among the many genera of free-living amoebae that exist in nature, members of only four genera have an association with human disease: Acanthamoeba spp., Balamuthia mandrillaris, Naegleria fowleri and Sappinia diploidea. Acanthamoeba spp. and B. mandrillaris are opportunistic pathogens causing infections of the central nervous system, lungs, sinuses and skin, mostly in immunocompromised humans. Balamuthia is also associated with disease in immunocompetent children, and Acanthamoeba spp. cause a sight-threatening infection, Acanthamoeba keratitis, mostly in contact-lens wearers. Of more than 30 species of Naegleria, only one species, N. fowleri, causes an acute and fulminating meningoencephalitis in immunocompetent children and young adults. In addition to human infections, Acanthamoeba, Balamuthia and Naegleria can cause central nervous system infections in animals. Because only one human case of encephalitis caused by Sappinia diploidea is known, generalizations about the organism as an agent of disease are premature. In this review we summarize what is known of these free-living amoebae, focusing on their biology, ecology, types of disease and diagnostic methods. We also discuss the clinical profiles, mechanisms of pathogenesis, pathophysiology, immunology, antimicrobial sensitivity and molecular characteristics of these amoebae.

  13. Anti-Acanthamoeba activity of contact lens solutions

    PubMed Central

    Niszl, I.; Markus, M.

    1998-01-01

    AIMS—This study was undertaken to investigate the effects of contact lens disinfecting solutions on strains of Acanthamoeba from the United Kingdom and southern Africa and to compare the results with those of other researchers. No information was previously available for southern African isolates.
METHODS—11 contact lens solutions were tested on cysts of 10 strains of Acanthamoeba.
RESULTS—Not all solutions used in the study were effective, with some for hard and gas permeable contact lenses being more satisfactory than those for soft contact lenses. The most effective of the gas permeable and hard contact lens solutions tested was Transoak (0.01% (wt/vol) benzalkonium chloride), which killed cysts of all strains within 4 hours of exposure. Oxysept 1 (31 mg hydrogen peroxide/ml) was the best soft contact lens solution tested. It eliminated cysts of certain strains within 4 hours, whereas cysts of other strains were only inactivated within either 8 or 72 hours.
CONCLUSIONS—Manufacturers should be aware of the killing time for Acanthamoeba by contact lens solutions and should provide appropriate guidelines for the use thereof. The killing time for cysts of the African and UK isolates studied is, in general, similar. Therefore, it must in the present state of knowledge be assumed that usage guidelines suggested in the UK are also appropriate for travellers to South Africa and for local residents in South Africa.

 Keywords: contact lenses; Acanthamoeba; keratitis PMID:9893594

  14. Campylobacter-Acanthamoeba interactions.

    PubMed

    Vieira, Ana; Seddon, Alan M; Karlyshev, Andrey V

    2015-05-01

    Campylobacter jejuni is a foodborne pathogen recognized as the major cause of human bacterial enteritis. Undercooked poultry products and contaminated water are considered as the most important sources of infection. Some studies suggest transmission and survival of this bacterial pathogen may be assisted by the free-living protozoa Acanthamoeba. The latter is known to play the role of a host for various pathogenic bacteria, protecting them from harsh environmental conditions. Importantly, there is a similarity between the mechanisms of bacterial survival within amoebae and macrophages, making the former a convenient tool for the investigation of the survival of pathogenic bacteria in the environment. However, the molecular mechanisms involved in the interaction between Campylobacter and Acanthamoeba are not well understood. Whilst some studies suggest the ability of C. jejuni to survive within the protozoa, the other reports support an extracellular mode of survival only. In this review, we focus on the studies investigating the interaction between Campylobacter and Acanthamoeba, address some reasons for the contradictory results, and discuss possible implications of these results for epidemiology. Additionally, as the molecular mechanisms involved remain unknown, we also suggest possible factors that may be involved in this process. Deciphering the molecular mechanisms of pathogen-protozoa interaction will assist in a better understanding of Campylobacter lifestyle and in the development of novel antibacterial drugs.

  15. Inactivation of Acanthamoeba spp. and Other Ocular Pathogens by Application of Cold Atmospheric Gas Plasma.

    PubMed

    Heaselgrave, Wayne; Shama, Gilbert; Andrew, Peter W; Kong, Michael G

    2016-05-15

    Currently there are estimated to be approximately 3.7 million contact lens wearers in the United Kingdom and 39.2 million in North America. Contact lens wear is a major risk factor for developing an infection of the cornea known as keratitis due to poor lens hygiene practices. While there is an international standard for testing disinfection methods against bacteria and fungi (ISO 14729), no such guidelines exist for the protozoan Acanthamoeba, which causes a potentially blinding keratitis most commonly seen in contact lens wearers, and as a result, many commercially available disinfecting solutions show incomplete disinfection after 6 and 24 h of exposure. Challenge test assays based on international standard ISO 14729 were used to determine the antimicrobial activity of cold atmospheric gas plasma (CAP) against Pseudomonas aeruginosa, Candida albicans, and trophozoites and cysts of Acanthamoeba polyphaga and Acanthamoeba castellanii P. aeruginosa and C. albicans were completely inactivated in 0.5 min and 2 min, respectively, and trophozoites of A. polyphaga and A. castellanii were completely inactivated in 1 min and 2 min, respectively. Furthermore, for the highly resistant cyst stage of both species, complete inactivation was achieved after 4 min of exposure to CAP. This study demonstrates that the CAP technology is highly effective against bacterial, fungal, and protozoan pathogens. The further development of this technology has enormous potential, as this approach is able to deliver the complete inactivation of ocular pathogens in minutes, in contrast to commercial multipurpose disinfecting solutions that require a minimum of 6 h.

  16. Prevalence of pathogenic Acanthamoeba (Protozoa:Amoebidae) in the atmosphere of the city of San Luis Potosi, Mexico.

    PubMed

    Rodriguez-Zaragoza, S; Magana-Becerra, A

    1997-01-01

    Several species of pathogenic Acanthamoeba cause infections to humans, but amoebic keratitis is more frequently found than any other due to the increasing number of contact lens wearers in the world. Cysts and trophozoites of these amebas are airborne and may pollute water from the air. We investigated the proportion of pathogenic Acanthamoeba from the atmosphere of the city of San Luis Potosi. Samples were taken by the impinger method, every month during one year. We isolated 23 strains of Acanthamoeba, 61% of them were non-pathogenic, 31% were non-pathogenic with invasive capacity and 8% were pathogenic to mice. Almost 40% of these strains represent danger of infections to humans. The isolations were more abundant during the dry season in the south (urban) and west (suburban) stations, which means that the sanitary conditions around stands may enhance the proportion of pathogenic strains in the surroundings.

  17. Comparison of Mycotic Keratitis with Nonmycotic Keratitis: An Epidemiological Study

    PubMed Central

    Khater, Mohammad M.; Shehab, Nehal S.; El-Badry, Anwar S.

    2014-01-01

    Purpose. This work aims to study the problems encountered with and the different epidemiological features of patients with fungal keratitis. Patients and Methods. All cases with keratitis attending the Outpatient Clinic of Ophthalmology Department at Tanta University Hospital during three years from the first of January 2011 to the end of December 2013 were selected and carefully examined and cases with mycotic keratitis were further examined and investigated. Results. From 66303 attendants during this period with different complaints, there were 361 cases (0.54%) with mycotic keratitis and 473 cases (0.71%) of nonmycotic origin. Mycotic keratitis is common between 40 and 60 years, more in farmers (64%), families with large number and large crowding index, rural than urban residence, and patients with outdoor water sources and insanitary sewage disposal. Positive fungal cultures were obtained in 84.5% and were negative in 15.5% of cases in spite of their typical clinical findings for diagnosis and their improvement with antifungal therapy. Conclusion. Mycotic keratitis is more frequent in farmers, rural areas, outdoor water supply, insanitary sewage disposal, and patients preceded with organic trauma. Atypical clinical findings were found in some cases and not all cases improved with specific antifungal therapy. PMID:25548657

  18. Inhibition of 3-Hydroxy-3-Methylglutaryl–Coenzyme A Reductase and Application of Statins as a Novel Effective Therapeutic Approach against Acanthamoeba Infections

    PubMed Central

    Lorenzo-Morales, Jacob; Machin, Rubén P.; López-Arencibia, Atteneri; García-Castellano, José Manuel; de Fuentes, Isabel; Loftus, Brendan; Maciver, Sutherland K.; Valladares, Basilio; Piñero, José E.

    2013-01-01

    Acanthamoeba is an opportunistic pathogen in humans, whose infections most commonly manifest as Acanthamoeba keratitis or, more rarely, granulomatous amoebic encephalitis. Although there are many therapeutic options for the treatment of Acanthamoeba, they are generally lengthy and/or have limited efficacy. Therefore, there is a requirement for the identification, validation, and development of novel therapeutic targets against these pathogens. Recently, RNA interference (RNAi) has been widely used for these validation purposes and has proven to be a powerful tool for Acanthamoeba therapeutics. Ergosterol is one of the major sterols in the membrane of Acanthamoeba. 3-Hydroxy-3-methylglutaryl–coenzyme A (HMG-CoA) reductase is an enzyme that catalyzes the conversion of HMG-CoA to mevalonate, one of the precursors for the production of cholesterol in humans and ergosterol in plants, fungi, and protozoa. Statins are compounds which inhibit this enzyme and so are promising as chemotherapeutics. In order to validate whether this enzyme could be an interesting therapeutic target in Acanthamoeba, small interfering RNAs (siRNAs) against HMG-CoA were developed and used to evaluate the effects induced by the inhibition of Acanthamoeba HMG-CoA. It was found that HMG-CoA is a potential drug target in these pathogenic free-living amoebae, and various statins were evaluated in vitro against three clinical strains of Acanthamoeba by using a colorimetric assay, showing important activities against the tested strains. We conclude that the targeting of HMG-CoA and Acanthamoeba treatment using statins is a novel powerful treatment option against Acanthamoeba species in human disease. PMID:23114753

  19. Microbial keratitis in Gujarat, Western India: findings from 200 cases

    PubMed Central

    Kumar, Anil; Pandya, Snehal; Kavathia, Ghanshyam; Antala, Sejul; Madan, Molly; Javdekar, Tanuja

    2011-01-01

    Introduction The objective of this study was to study the epidemiological characteristics and the microbiological profile of patients suspected with microbial keratitis in Gujarat. Methods Corneal scraping was collected from 200 consecutive cases of suspected microbial keratitis and was subjected to direct examination and culture. Results Of the 200 ulcers 55% were culture positive, 26.5% were bacterial ulcers of which 47% were due to Staphylococcus spp. Pure fungal growth was seen in 22% while 6% were mixed ulcers. Fusarium spp. (30%) was the most common fungus followed by Aspergillus spp. (21%). Only one case of Acanthamoeba keratitis was encountered. Patients were mainly from rural areas (61.5%) with male preponderance (61.5%). Corneal injury was seen in 78.5% cases of which 53% had injury with vegetative matter. Prior treatment was seen in 58% of which 5% had been treated by village healers. Nineteen patients (9.5%) also used some kind of traditional topical treatment. Increased incidence was seen from August to December. Five case of fugal ulcers lead to perforation of which three were due to Fusarium spp. whereas perforation was seen in only two cases of bacterial ulcers due to Pseudomonas aeruginosa. Conclusion Staphylococcus and Fusarium spp. were the most common etiological agents in our region. Predominant outdoor agricultural activity is the principal causative factor for corneal injury. Corneal ulcers complicated due to treatment by village healers are another important concern. The information regarding regional etiology will help empirical management as many eye clinics do not have microbiological facilities. PMID:22384294

  20. Isolation of Acanthamoeba Genotype T4 from a Non-Contact Lens Wearer from the Philippines

    PubMed Central

    Buerano, Corazon C.; Trinidad, Abigail D.; Fajardo, Lindsay Sydney N.; Cua, Irwin Y.; Baclig, Michael O.; Natividad, Filipinas F.

    2014-01-01

    We report the case of a 76-year old Filipino male who presented with pain, redness, and blurring of vision of the right eye. Corneal scraping was done and sent to the St. Luke’s Research and Biotechnology Group for detection and identification of the infectious agent. Morphological detection was performed by allowing the organism from the scraping to grow in 1.5% non-nutrient agar plate with heat-killed E. coli. Trophozoites with acanthopodia and double-walled cysts characteristic of Acanthamoeba were observed within the first and second week of observations, respectively. Molecular identification of the amoebae at the genus level based on the presence of Acanthamoeba-specific amplimer S1, ASA.S1 confirmed the morphological identification. Genotyping through sequence revealed that the organism belonged to T4, which is the genotype commonly present in the eye of keratitis patients. PMID:25589879

  1. Isolation and identification of Acanthamoeba spp. from thermal swimming pools and spas in Southern Brazil.

    PubMed

    Fabres, Laura Fuhrich; Rosa Dos Santos, Sayonara Peixoto; Benitez, Lisianne Brittes; Rott, Marilise Brittes

    2016-03-01

    Free-living amoebae (FLA) are widely distributed in soil and water. A few number of them are implicated in human disease: Acanthamoeba spp., Naegleria fowleri, Balamuthia mandrillaris and Sappinia diploidea. Species of Acanthamoeba can cause keratitis and brain infections. In this study, 72 water samples were taken from both hot tubs and thermal swimming pools in the city of Porto Alegre, RS, Brazil, to determine the presence of Acanthamoeba in the water as well as perform the phenotypic and genotypic characterization of the isolates. The identification of the isolates was based on the cysts morphology and PCR amplification using genus-specific oligonucleotides. When the isolates were submitted to PCR reaction only 8 were confirmed as belonging to the genus Acanthamoeba. The sequences analysis when compared to the sequences in the GenBank, showed genotype distribution in group T3 (12,5%), T5 (12,5%), T4 (25%) and T15 (50%). The results of this study confirmed the presence of potentially pathogenic isolates of free living amoebae in hot swimming pool and spas which can present risks to human health. PMID:27078644

  2. Molecular Characterization of Pathogenic Acanthamoeba Isolated from Drinking and Recreational water in East Azerbaijan, Northwest Iran.

    PubMed

    Behniafar, Hamed; Niyyati, Maryam; Lasjerdi, Zohreh

    2015-01-01

    Acanthamoeba- related infections, such as amoebic keratitis and granulomatous amoebic encephalitis, can develop in high-risk population through contaminated water sources. Thus, surveying water resources, particularly those available for human use, is of the utmost importance. In the present study, 67 water samples were collected from water resources in East Azerbaijan, a province in northwestern Iran. Samples were cultured on enriched non-nutrient agar plates, and sequencing-based approaches were used for genotyping. The pathogenic potential of the isolates was determined using thermo- and osmo-tolerance tests. Acanthamoeba were detected in 17 (25.4%) of the 67 collected samples. Sequencing analysis revealed that the isolates belonged to the T3 (23.52%), mixed T3/T4 (5.88%), T4 (58.82%), T5 (5.88%), and T13 (5.88%) genotypes. Through thermo- and osmo-tolerance tests, 88.23% of isolates were resistant to 37 °C, 40 °C temperature, and 0.5 M and 1 M osmolarity; thus, these isolates had the potential for pathogenicity. These findings point toa serious public health concern in the studied region. This study is the first to report Acanthamoeba isolated from drinking and recreational water sources in East Azerbaijan and Acanthamoeba T13 isolated from tap water in Iran. PMID:26157334

  3. Molecular Characterization of Pathogenic Acanthamoeba Isolated from Drinking and Recreational water in East Azerbaijan, Northwest Iran

    PubMed Central

    Behniafar, Hamed; Niyyati, Maryam; Lasjerdi, Zohreh

    2015-01-01

    Acanthamoeba- related infections, such as amoebic keratitis and granulomatous amoebic encephalitis, can develop in high-risk population through contaminated water sources. Thus, surveying water resources, particularly those available for human use, is of the utmost importance. In the present study, 67 water samples were collected from water resources in East Azerbaijan, a province in northwestern Iran. Samples were cultured on enriched non-nutrient agar plates, and sequencing-based approaches were used for genotyping. The pathogenic potential of the isolates was determined using thermo- and osmo-tolerance tests. Acanthamoeba were detected in 17 (25.4%) of the 67 collected samples. Sequencing analysis revealed that the isolates belonged to the T3 (23.52%), mixed T3/T4 (5.88%), T4 (58.82%), T5 (5.88%), and T13 (5.88%) genotypes. Through thermo- and osmo-tolerance tests, 88.23% of isolates were resistant to 37 °C, 40 °C temperature, and 0.5 M and 1 M osmolarity; thus, these isolates had the potential for pathogenicity. These findings point toa serious public health concern in the studied region. This study is the first to report Acanthamoeba isolated from drinking and recreational water sources in East Azerbaijan and Acanthamoeba T13 isolated from tap water in Iran. PMID:26157334

  4. Isolation and identification of Acanthamoeba spp. from thermal swimming pools and spas in Southern Brazil.

    PubMed

    Fabres, Laura Fuhrich; Rosa Dos Santos, Sayonara Peixoto; Benitez, Lisianne Brittes; Rott, Marilise Brittes

    2016-03-01

    Free-living amoebae (FLA) are widely distributed in soil and water. A few number of them are implicated in human disease: Acanthamoeba spp., Naegleria fowleri, Balamuthia mandrillaris and Sappinia diploidea. Species of Acanthamoeba can cause keratitis and brain infections. In this study, 72 water samples were taken from both hot tubs and thermal swimming pools in the city of Porto Alegre, RS, Brazil, to determine the presence of Acanthamoeba in the water as well as perform the phenotypic and genotypic characterization of the isolates. The identification of the isolates was based on the cysts morphology and PCR amplification using genus-specific oligonucleotides. When the isolates were submitted to PCR reaction only 8 were confirmed as belonging to the genus Acanthamoeba. The sequences analysis when compared to the sequences in the GenBank, showed genotype distribution in group T3 (12,5%), T5 (12,5%), T4 (25%) and T15 (50%). The results of this study confirmed the presence of potentially pathogenic isolates of free living amoebae in hot swimming pool and spas which can present risks to human health.

  5. Amebicidal activity of plant extracts from Southeast Asia on Acanthamoeba spp.

    PubMed

    Chu, D M; Miles, H; Toney, D; Ngyuen, C; Marciano-Cabral, F

    1998-09-01

    The effect of 100 polar and 100 nonpolar plant extract materials obtained from Southeast Asia were evaluated for amebicidal activity in vitro against three species of Acanthamoeba. A. culbertsoni, A. castellanii, and A. polyphaga, the causative agents of granulomatous amebic encephalitis and amebic keratitis, were studied in vitro to determine whether the plant extracts exhibited amebicidal activity or induced encystment of the amebae. Of the 200 plant extracts tested, extracts obtained from three plants (Ipomoea sp., Kaempferia galanga, and Cananga odorata) were amebicidal for all three species of Acanthamoeba and a fourth extract prepared from Gastrochilus panduratum was lytic for A. polyphaga and growth-inhibitory for A. castellanii and A. culbertsoni. Three plant extracts induced encystment of all three species of Acanthamoeba. Select plant extracts were tested as well for tumoricidal activity against B103 neuroblastoma cells. Some plant extracts that exhibited tumoricidal activity for B103 cells were not amebicidal for Acanthamoeba spp. Additionally, the polar and nonpolar extracts that exhibited amebicidal activity were also tested for activity against primary murine peritoneal macrophage cultures. Plant extracts that demonstrated tumoricidal or amebicidal activity were not lytic for normal macrophage cultures. PMID:9766904

  6. Autophagy protein 8 mediating autophagosome in encysting Acanthamoeba.

    PubMed

    Moon, Eun-Kyung; Chung, Dong-Il; Hong, Yeon-Chul; Kong, Hyun-Hee

    2009-11-01

    Autophagy is an evolutionally conserved protein degradation pathway in eukaryotes. It plays essential roles during starvation, cellular differentiation, cell death, and aging by eliminating unwanted or unnecessary organelles and recycling the components for reuse. ATG8, a member of a novel ubiquitin-like protein family, is an essential component of the autophagic machinery. The present study identified and characterized autophagy protein 8 in Acanthamoeba castellanii an amphizoic amoeba causing granulomatous amoebic encephalitis and amoebic keratitis in humans. Real-time polymerase chain reaction demonstrated that the A. castellanii Atg8 (AcAtg8) gene encoding a 118 amino acid protein was highly expressed during encystation. Fluorescence microscopic analysis following transient transfection of enhanced green fluorescent protein-AcAtg8 revealed small or large vacuolar fluorescent structures in an encysting amoeba. The Atg8 fluorescent structures on the membrane were identified as autophagosomes by co-localization analysis with LysoTracker. Chemically synthesized small interfering RNA against AcAtg8 reduced the encystation efficiency and inhibited autophagosome formation in Acanthamoeba. PMID:19560492

  7. Proteomic profiling of the infective trophozoite stage of Acanthamoeba polyphaga.

    PubMed

    Caumo, Karin Silva; Monteiro, Karina Mariante; Ott, Thiely Rodrigues; Maschio, Vinicius José; Wagner, Glauber; Ferreira, Henrique Bunselmeyer; Rott, Marilise Brittes

    2014-12-01

    Acanthamoeba polyphaga is a free-living protozoan pathogen, whose infective trophozoite form is capable of causing a blinding keratitis and fatal granulomatous encephalitis in humans. The damage caused by A. polyphaga trophozoites in human corneal or brain infections is the result of several different pathogenic mechanisms that have not yet been elucidated at the molecular level. We performed a comprehensive analysis of the proteins expressed by A. polyphaga trophozoites, based on complementary 2-DE MS/MS and gel-free LC-MS/MS approaches. Overall, 202 non-redundant proteins were identified. An A. polyphaga proteomic map in the pH range 3-10 was produced, with protein identification for 184 of 370 resolved spots, corresponding to 142 proteins. Additionally, 94 proteins were identified by gel-free LC-MS/MS. Functional classification revealed several proteins with potential importance for pathogen survival and infection of mammalian hosts, including surface proteins and proteins related to defense mechanisms. Our study provided the first comprehensive proteomic survey of the trophozoite infective stage of an Acanthamoeba species, and established foundations for prospective, comparative and functional studies of proteins involved in mechanisms of survival, development, and pathogenicity in A. polyphaga and other pathogenic amoebae.

  8. Detection of Acanthamoeba on the ocular surface in a Spanish population using the Schirmer strip test: pathogenic potential, molecular classification and evaluation of the sensitivity to chlorhexidine and voriconazole of the isolated Acanthamoeba strains.

    PubMed

    Rocha-Cabrera, Pedro; Reyes-Batlle, María; Martín-Navarro, Carmen María; Dorta-Gorrín, Alexis; López-Arencibia, Atteneri; Sifaoui, Ines; Martínez-Carretero, Enrique; Piñero, José E; Martín-Barrera, Fernando; Valladares, Basilio; Lorenzo-Morales, Jacob

    2015-08-01

    Pathogenic strains of Acanthamoeba are causative agents of a sight-threatening infection of the cornea known as Acanthamoeba keratitis, which is often associated with the misuse of contact lenses. However, there is still a question remaining to be answered, which is whether these micro-organisms are present on the ocular surface of healthy individuals. Therefore, the aim of this study was to determine the presence of Acanthamoeba on the ocular surface in healthy patients and also in those with other ocular surface infections. Sterile Schirmer test strips were used to collect samples from a group of patients who attended an ophthalmology consultation at the Hospital del Norte, Icod de los Vinos, Tenerife, Canary Islands. Most of the patients (46 individuals, 79.31  %) presented ocular surface pathologies such as blepharitis or conjunctivitis; the rest did not present any pathology. None of the patients included in the study wore contact lenses. The collected samples were cultured in 2  % non-nutrient agar plates and positive plates were then cultured in axenic conditions for further analyses. Molecular analysis classified all isolated strains as belonging to Acanthamoeba genotype tbl4, and osmotolerance and thermotolerance assays revealed that all strains were potentially pathogenic. Furthermore, all strains were assayed for sensitivity against voriconazole and chlorhexidine. Assays showed that both drugs were active against the tested strains. In conclusion, the Schirmer strip test is proposed as an effective tool for the detection of Acanthamoeba on the ocular surface.

  9. Detection of Acanthamoeba on the ocular surface in a Spanish population using the Schirmer strip test: pathogenic potential, molecular classification and evaluation of the sensitivity to chlorhexidine and voriconazole of the isolated Acanthamoeba strains.

    PubMed

    Rocha-Cabrera, Pedro; Reyes-Batlle, María; Martín-Navarro, Carmen María; Dorta-Gorrín, Alexis; López-Arencibia, Atteneri; Sifaoui, Ines; Martínez-Carretero, Enrique; Piñero, José E; Martín-Barrera, Fernando; Valladares, Basilio; Lorenzo-Morales, Jacob

    2015-08-01

    Pathogenic strains of Acanthamoeba are causative agents of a sight-threatening infection of the cornea known as Acanthamoeba keratitis, which is often associated with the misuse of contact lenses. However, there is still a question remaining to be answered, which is whether these micro-organisms are present on the ocular surface of healthy individuals. Therefore, the aim of this study was to determine the presence of Acanthamoeba on the ocular surface in healthy patients and also in those with other ocular surface infections. Sterile Schirmer test strips were used to collect samples from a group of patients who attended an ophthalmology consultation at the Hospital del Norte, Icod de los Vinos, Tenerife, Canary Islands. Most of the patients (46 individuals, 79.31  %) presented ocular surface pathologies such as blepharitis or conjunctivitis; the rest did not present any pathology. None of the patients included in the study wore contact lenses. The collected samples were cultured in 2  % non-nutrient agar plates and positive plates were then cultured in axenic conditions for further analyses. Molecular analysis classified all isolated strains as belonging to Acanthamoeba genotype tbl4, and osmotolerance and thermotolerance assays revealed that all strains were potentially pathogenic. Furthermore, all strains were assayed for sensitivity against voriconazole and chlorhexidine. Assays showed that both drugs were active against the tested strains. In conclusion, the Schirmer strip test is proposed as an effective tool for the detection of Acanthamoeba on the ocular surface. PMID:26293786

  10. Immune-mediated canine and feline keratitis.

    PubMed

    Andrew, Stacy E

    2008-03-01

    Although the normal cornea is devoid of vasculature and lymphatics, there are still several immune-mediated corneal conditions that can occur in dogs and cats. An overview of corneal immunology is presented. Diseases of dogs, including chronic superficial keratitis, superficial punctate keratitis, and canine adenovirus endotheliitis, as well as feline diseases, including eosinophilic keratitis and herpesvirus-related conditions, are discussed.

  11. High occurrence of Acanthamoeba genotype T4 in soil sources from Bolívar State, Venezuela.

    PubMed

    Wagner, Carolina; Reyes-Batlle, María; Hernán, Aurora; Rojas, Elsy; Pérez, Gladymar; López-Arencibia, Atteneri; Sifaoui, Ines; Martínez-Carretero, Enrique; Piñero, José E; Valladares, Basilio; Lorenzo-Morales, Jacob

    2016-09-01

    Pathogenic strains of Acanthamoeba are causative agents of keratitis and encephalitis that often may end fatal in humans and other animals. In the present study, twenty-seven soil samples were collected in the Bolivar State in Venezuela and checked for the presence of Acanthamoeba. Samples were cultivated onto 2% non-nutrient agar plates seeded with a layer of heat killed E. coli. Amplification by PCR and sequencing of the DF3 region of the 18S rDNA of Acanthamoeba was carried out in order to confirm morphological identification of the amoebae. Furthermore, Acanthamoeba spp. was isolated from 51.8% of soil samples. Sequencing of the DF3 region of the 18S rDNA resulted in the identification of genotype T4 in all samples. To the best of our knowledge, this is the first report of genotype T4 in soil sources from Venezuela. Further studies should be carried out in this State and in the country in order to determine the current occurrence of Acanthamoeba in Venezuelan environments. PMID:27447209

  12. Synthesis, characterization and amoebicidal potential of locally synthesized TiO2 nanoparticles against pathogenic Acanthamoeba trophozoites in vitro.

    PubMed

    Imran, Muhammad; Muazzam, Ambreen Gul; Habib, Amir; Matin, Abdul

    2016-06-01

    Acanthamoeba is an opportunistic protozoan pathogen that plays a pivotal role in the ecosystem. It may cause blinding keratitis and fatal encephalitis involving the central nervous system. Here we synthesized pure and Zn doped TiO2 nanoparticles (~10-30nm) via sol-gel and sol-hydrothermal methods and demonstrated its impact on the biological characteristics of pathogenic Acanthamoeba castellanii. Our results revealed that pure and Zn doped TiO2 nanoparticles synthesized by sol-hydrothermal methods (ranging 5, 10, 25 and 50μg/ml) exhibited amoebicidal effects i.e., >60% of trophozoites executed under normal light at maximum dose (50μg/ml) within 1h incubation. In contrast pure/doped TiO2 obtained via sol gel method showed ~40% amoeba damage. Furthermore, amoebae growth assay demonstrated that Zn doped TiO2 also inhibited Acanthamoeba numbers up to 7days in dose dependent manner. It was interesting to note that all the tested TiO2 nanoparticles have shown maximum amoebicidal effects at pH7 which is quite relevant to amoebic growth favorable conditions. Our results confirmed that TiO2 has inhibitory effects on Acanthamoeba growth and viability. Overall, we reported the amoebicidal and amoebic growth inhibition potential of pure and Zn doped TiO2 nanoparticles against Acanthamoeba due to attached OH(-) groups, reduced size and decreased band gap of sol hydrothermally synthesized TiO2 nanoparticles. PMID:27054875

  13. Characterization of a New Pathogenic Acanthamoeba Species, A. byersi n. sp., Isolated from a Human with Fatal Amoebic Encephalitis

    PubMed Central

    Qvarnstrom, Yvonne; Nerad, Thomas A.; Visvesvara, Govinda S.

    2015-01-01

    Acanthamoeba spp. are free-living amoebae that are ubiquitous in natural environments. They can cause cutaneous, nasopharyngeal and disseminated infection, leading to granulomatous amebic encephalitis (GAE) in immunocompromised individuals. In addition, they can cause amoebic keratitis in contact lens wearers. Acanthamoeba GAE is almost always fatal because of difficulty and delay in diagnosis and lack of optimal antimicrobial therapy. Here we report the description of an unusual strain isolated from skin and brain of a GAE patient. The amoebae displayed large trophozoites and star-shaped cysts, characteristics for acanthamoebas belonging to morphology Group 1. However, its unique morphology and growth characteristics differentiated this new strain from other Group 1 species. DNA sequence analysis, secondary structure prediction and phylogenetic analysis of the 18S rRNA gene confirmed that this new strain belonged to Group 1 but that it was distinct from the other sequence types within that group. Thus, we hereby propose the establishment of a new species, Acanthamoeba byersi n. sp. as well as a new sequence type, T18, for this new strain. To our knowledge, this is the first report of a Group 1 Acanthamoeba that is indisputably pathogenic in humans. PMID:23879685

  14. Cytomegalovirus keratitis after penetrating keratoplasty.

    PubMed

    Wehrly, S R; Manning, F J; Proia, A D; Burchette, J L; Foulks, G N

    1995-11-01

    We report the development of cytomegalovirus (CMV) keratitis in the penetrating keratoplasty of a 59-year-old human immunodeficiency virus-negative woman after uncomplicated corneal transplantation. Immunosuppression with topical cyclosporine A 2% in corn oil and topical prednisolone acetate 1% suspension was used postoperatively. The 15-month postoperative course was complicated by multiple episodes of endothelial rejection, medically controlled elevated intraocular pressure, polymicrobial bacterial (coagulase-negative staphlococcus and alpha-hemolytic streptococcus) keratitis, and endothelial plaque formation with associated hypopyon and epithelial defect. The graft failed and penetrating keratoplasty was repeated. Cytomegalovirus infection of superficial keratocytes in a region of scarring was identified in histological sections stained with hematoxylin and eosin and confirmed using mouse monoclonal anti-cytomegalovirus antibodies. Excision of the diseased corneal button with no additional treatment appears to have been curative. Low-grade keratitis was the only manifestation of the CMV infection, and it has not recurred 6 months postoperatively.

  15. Ochrobactrum anthropi Keratitis with Focal Descemet's Membrane Detachment and Intracorneal Hypopyon

    PubMed Central

    Venkateswaran, Nandini; Wozniak, Rachel A. F.

    2016-01-01

    Purpose. To describe a unique case of O. anthropi keratitis associated with a rare manifestation of Descemet's membrane detachment and intracorneal hypopyon and to discuss challenges in diagnosis and management. Methods. Best-corrected visual acuity was measured with Snellen letters. Corneal scrapings were performed and aerobic, viral, herpetic, acid-fast bacilli, Acanthamoeba, and fungal stains and cultures were obtained. Following evisceration, tissue was evaluated for histologic features and again stained for bacteria, mycobacteria, Acanthamoeba, fungi, and viral particles. Results. Initial presentation to our institute was notable for a corneal ulcer, focal Descemet's membrane detachment, and intracorneal hypopyon. Speciation of initial corneal scrapes revealed Ochrobactrum anthropi and initial management included fortified tobramycin. Despite medical therapy, the patient developed a corneal perforation and required subsequent evisceration. Conclusion. O. anthropi is an emerging ocular pathogen that has not been previously reported in cases of keratitis. As this pathogen becomes increasingly recognized as a source of ocular infections, it is important to identify and treat aggressively to avoid vision-threatening disease. PMID:27777806

  16. Isolation and molecular characterization of potentially pathogenic Acanthamoeba genotypes from diverse water resources including household drinking water from Khyber Pakhtunkhwa, Pakistan.

    PubMed

    Tanveer, Tania; Hameed, Abdul; Muazzam, Ambreen Gul; Jung, Suk-Yul; Gul, Asma; Matin, Abdul

    2013-08-01

    Acanthamoeba, an opportunistic protozoan pathogen, is ubiquitous in nature, and therefore plays a predatory role and helps control microbial communities in the ecosystem. These Acanthamoeba species are recognized as opportunistic human pathogens that may cause blinding keratitis and rare but fatal granulomatous encephalitis. To date, there is not a single report demonstrating Acanthamoeba isolation and identification from environmental sources in Pakistan, and that is the aim of this study. Acanthamoeba were identified by morphological characteristics of their cysts on non-nutrient agar plates seeded with Escherichia coli. Additionally, the polymerase chain reaction (PCR) was performed with genus-specific primers followed by direct sequencing of the PCR product for molecular identification. Furthermore, our PCR and sequencing results confirmed seven different pathogenic and nonpathogenic genotypes, including T2-T10, T4, T5, T7, T15, T16, and T17. To the best of our knowledge, we have identified and isolated Acanthamoeba sp., for the first time, from water resources of Khyber Pakhtunkhwa, Pakistan. There is an urgent need to address (1) the pathogenic potential of the identified genotypes and (2) explore other environmental sources from the country to examine the water quality and the current status of Acanthamoeba species in Pakistan, which may be a potential threat for public health across the country.

  17. Acanthamoeba castellanii STAT protein.

    PubMed

    Kicinska, Anna; Leluk, Jacek; Jarmuszkiewicz, Wieslawa

    2014-01-01

    STAT (signal transducers and activators of transcription) proteins are one of the important mediators of phosphotyrosine-regulated signaling in metazoan cells. We described the presence of STAT protein in a unicellular, free-living amoebae with a simple life cycle, Acanthamoeba castellanii. A. castellanii is the only, studied to date, Amoebozoan that does not belong to Mycetozoa but possesses STATs. A sequence of the A. castellanii STAT protein includes domains similar to those of the Dictyostelium STAT proteins: a coiled coil (characteristic for Dictyostelium STAT coiled coil), a STAT DNA-binding domain and a Src-homology domain. The search for protein sequences homologous to A. castellanii STAT revealed 17 additional sequences from lower eukaryotes. Interestingly, all of these sequences come from Amoebozoa organisms that belong to either Mycetozoa (slime molds) or Centramoebida. We showed that there are four separated clades within the slime mold STAT proteins. The A. castellanii STAT protein branches next to a group of STATc proteins from Mycetozoa. We also demonstrate that Amoebozoa form a distinct monophyletic lineage within the STAT protein world that is well separated from the other groups. PMID:25338074

  18. Acanthamoeba everywhere: high diversity of Acanthamoeba in soils.

    PubMed

    Geisen, Stefan; Fiore-Donno, Anna Maria; Walochnik, Julia; Bonkowski, Michael

    2014-09-01

    Acanthamoeba is a very abundant genus of soil protists with fundamental importance in nutrient cycling, but several strains can also act as human pathogens. The systematics of the genus is still unclear: currently 18 small-subunit (SSU or 18S) ribosomal RNA sequence types (T1-T18) are recognized, which sometimes contain several different morphotypes; on the other hand, some morphological identical strains belong to different sequence types, sometimes appearing in paraphyletic positions. In this study, we cultivated 65 Acanthamoeba clones from soil samples collected under grassland at three separate locations in the Netherlands, in Sardinia and at high altitude mountains in Tibet. We obtained 24 distinct partial sequences, which predominantly grouped within sequence type T4 followed by T2, T13, T16 and "OX-1" (in the T2/T6 clade). Our sequences were 98-99% similar, but none was identical to already known Acanthamoeba sequences. The community composition of Acanthamoeba strains differed between locations, T4 being the dominant sequence type in Sardinia and Tibet, but represented only half of the clones from soils in the Netherlands. The other half of clones from the Dutch soils was made up by T2, T16 and "OX-1", while T13 was only found in Sardinia and Tibet. None of the sequences was identical between localities. Several T4 clones from all three localities and all T13 clones grew at 37 °C while one T4 clone was highly cytopathogenic. PMID:24951165

  19. Acanthamoeba everywhere: high diversity of Acanthamoeba in soils.

    PubMed

    Geisen, Stefan; Fiore-Donno, Anna Maria; Walochnik, Julia; Bonkowski, Michael

    2014-09-01

    Acanthamoeba is a very abundant genus of soil protists with fundamental importance in nutrient cycling, but several strains can also act as human pathogens. The systematics of the genus is still unclear: currently 18 small-subunit (SSU or 18S) ribosomal RNA sequence types (T1-T18) are recognized, which sometimes contain several different morphotypes; on the other hand, some morphological identical strains belong to different sequence types, sometimes appearing in paraphyletic positions. In this study, we cultivated 65 Acanthamoeba clones from soil samples collected under grassland at three separate locations in the Netherlands, in Sardinia and at high altitude mountains in Tibet. We obtained 24 distinct partial sequences, which predominantly grouped within sequence type T4 followed by T2, T13, T16 and "OX-1" (in the T2/T6 clade). Our sequences were 98-99% similar, but none was identical to already known Acanthamoeba sequences. The community composition of Acanthamoeba strains differed between locations, T4 being the dominant sequence type in Sardinia and Tibet, but represented only half of the clones from soils in the Netherlands. The other half of clones from the Dutch soils was made up by T2, T16 and "OX-1", while T13 was only found in Sardinia and Tibet. None of the sequences was identical between localities. Several T4 clones from all three localities and all T13 clones grew at 37 °C while one T4 clone was highly cytopathogenic.

  20. Non-viral microbial keratitis in children

    PubMed Central

    Al-Otaibi, Abdullah G.

    2011-01-01

    Microbial (non-viral) keratitis is a serious vision-threatening condition. The management of microbial keratitis in children is particularly complicated by the children’s inability to cooperate during examinations and the lack of information prior to presentation. Predisposing factors vary according to geographical location and age. Corneal trauma is the leading cause for microbial keratitis in children, followed by systemic and ocular disease. Etiologic agents are most frequently Gram-positive and Gram-negative bacteria commonly found in contact lens-related microbial keratitis. Mycotic keratitis is a major risk factor in tropical weather conditions, particularly when associated with agricultural trauma. Early diagnosis, intensive drug treatment, and timely planned surgical intervention may effectively improve the outcome of pediatric microbial keratitis. PMID:23960991

  1. Amebicidal activity of the essential oils of Lippia spp. (Verbenaceae) against Acanthamoeba polyphaga trophozoites.

    PubMed

    Santos, Israel Gomes de Amorim; Scher, Ricardo; Rott, Marilise Brittes; Menezes, Leociley Rocha; Costa, Emmanoel Vilaça; Cavalcanti, Sócrates Cabral de Holanda; Blank, Arie Fitzgerald; Aguiar, Jaciana dos Santos; da Silva, Teresinha Gonçalves; Dolabella, Silvio Santana

    2016-02-01

    Amoebic keratitis and granulomatous amoebic encephalitis are caused by some strains of free-living amoebae of the genus Acanthamoeba. In the case of keratitis, one of the greatest problems is the disease recurrence due to the resistance of parasites, especially the cystic forms, to the drugs that are currently used. Some essential oils of plants have been used as potential active agents against this protist. Thus, the aim of this study was to determine the amebicidal activity of essential oils from plants of the genus Lippia against Acanthamoeba polyphaga trophozoites. To that end, 8 × 10(4) trophozoites were exposed for 24 h to increasing concentrations of essential oils from Lippia sidoides, Lippia gracilis, Lippia alba, and Lippia pedunculosa and to their major compounds rotundifolone, carvone, and carvacrol. Nearly all concentrations of oils and compounds showed amebicidal activity. The IC50 values for L. sidoides, L. gracilis L. alba, and L. pedunculosa were found to be 18.19, 10.08, 31.79, and 71.47 μg/mL, respectively. Rotundifolone, carvacrol, and carvone were determined as the major compounds showing IC50 of 18.98, 24.74, and 43.62 μg/mL, respectively. With the exception of oil from L. alba, the other oils evaluated showed low cytotoxicity in the NCI-H292 cell line. Given these results, the oils investigated here are promising sources of compounds for the development of complementary therapy against amoebic keratitis and granulomatous amoebic encephalitis and can also be incorporated into cleaning solutions to increase their amebicidal efficiency.

  2. Photodynamic inactivation of Acanthamoeba polyphaga with curcuminoids: an in vitro study

    NASA Astrophysics Data System (ADS)

    Corrêa, Thaila Q.; Geralde, Mariana C.; Carvalho, Mariana T.; Bagnato, Vanderlei S.; Kurachi, Cristina; de Souza, Clovis W. O.

    2016-03-01

    Acanthamoeba polyphaga are free-living amoebae that can be considered potentially pathogenic organisms by cause serious human infections, including keratitis and granulomatous amoebic encephalitis that usually results in death. Photodynamic inactivation (PDI) has been used for the biological control of microorganisms and can be promise in the control of Acanthamoeba infections. This study evaluated the in vitro effectiveness of PDI in A. polyphaga using curcuminoids salt as photosensitizer (PS) besides observing morphological changes caused by this PS in this organism, in confocal microscopy. A. polyphaga trophozoites were grown at 37°C in PYG medium for 48 to 72 hours. After, the trophozoites were incubated with PS solution during one hour and the samples were irradiated using light-emitting diodes at 460 nm at light doses 30 and 50 J/cm2. The results revealed reduction of 27.7%, 61.4% and 82.5% at 30 J/cm2 and 75.2%, 85.0% and 95.9% at 50 J/cm2, respectively, at curcuminoid salt concentrations of 500, 1000 and 1500 μg/mL. Through fluorescence images, it was possible to visualize the curcuminoid salt's uptake by the trophozoites. The PS showed toxicity to amoebae, in the dark, but the irradiation in PDI contributed to amoebae death effect. These data suggest that PDI may be an application of therapeutic intervention against Acanthamoeba infections, since it was effective in the inactivation of these amoebae.

  3. Evaluation of Acanthamoeba myosin-IC as a potential therapeutic target.

    PubMed

    Martín-Navarro, Carmen M; Lorenzo-Morales, Jacob; López-Arencibia, Atteneri; Reyes-Batlle, María; Piñero, José E; Valladares, Basilio; Maciver, Sutherland K

    2014-01-01

    Members of the genus Acanthamoeba are facultative pathogens of humans, causing a sight-threatening keratitis and a fatal encephalitis. We have targeted myosin-IC by using small interfering RNA (siRNA) silencing as a therapeutic approach, since it is known that the function of this protein is vital for the amoeba. In this work, specific siRNAs against the Acanthamoeba myosin-IC gene were developed. Treated and control amoebae were cultured in growth and encystment media to evaluate the induced effects after myosin-IC gene knockdown, as we have anticipated that cyst formation may be impaired. The effects of myosin-IC gene silencing were inhibition of cyst formation, inhibition of completion of cytokinesis, inhibition of osmoregulation under osmotic stress conditions, and death of the amoebae. The finding that myosin-IC silencing caused incompletion of cytokinesis is in agreement with earlier suggestions that the protein plays a role in cell locomotion, which is necessary to pull daughter cells apart after mitosis in a process known as "traction-mediated cytokinesis". We conclude that myosin-IC is a very promising potential drug target for the development of much-needed antiamoebal drugs and that it should be further exploited for Acanthamoeba therapy. PMID:24468784

  4. Incidence and molecular diversity of Acanthamoeba species isolated from public baths in Hungary.

    PubMed

    Kiss, Csaba; Barna, Zsófia; Vargha, Márta; Török, Júlia Katalin

    2014-07-01

    Hungary has a large number of thermal baths and spa facilities which attract hundreds of thousands of tourists annually. Until recently, however, the free-living amoebae were not of public health concern. Genotyping of Acanthamoeba species, potential agents of keratitis and granulomatous encephalitis, was carried out in 20 Hungarian public baths for the first time to assess the incidence and molecular diversity of the genus in the country. Our results show that 6.7% of the samples were positive for Acanthamoeba. Of these positive samples, 6.5 and 7% was from sterilized and unsterilized pools, respectively. The 18S rRNA gene investigation of the nine Acanthamoeba strains found reveals that seven belong to the hazardous T4 genotype. The remaining two samples were of the T15 type. All the strains kept growing at 36 °C. Our results underline the need to develop a control system for free-living amoebae and supervise the disinfection of Hungarian public baths.

  5. Evaluation of Acanthamoeba Myosin-IC as a Potential Therapeutic Target

    PubMed Central

    Lorenzo-Morales, Jacob; López-Arencibia, Atteneri; Reyes-Batlle, María; Piñero, José E.; Valladares, Basilio; Maciver, Sutherland K.

    2014-01-01

    Members of the genus Acanthamoeba are facultative pathogens of humans, causing a sight-threatening keratitis and a fatal encephalitis. We have targeted myosin-IC by using small interfering RNA (siRNA) silencing as a therapeutic approach, since it is known that the function of this protein is vital for the amoeba. In this work, specific siRNAs against the Acanthamoeba myosin-IC gene were developed. Treated and control amoebae were cultured in growth and encystment media to evaluate the induced effects after myosin-IC gene knockdown, as we have anticipated that cyst formation may be impaired. The effects of myosin-IC gene silencing were inhibition of cyst formation, inhibition of completion of cytokinesis, inhibition of osmoregulation under osmotic stress conditions, and death of the amoebae. The finding that myosin-IC silencing caused incompletion of cytokinesis is in agreement with earlier suggestions that the protein plays a role in cell locomotion, which is necessary to pull daughter cells apart after mitosis in a process known as “traction-mediated cytokinesis”. We conclude that myosin-IC is a very promising potential drug target for the development of much-needed antiamoebal drugs and that it should be further exploited for Acanthamoeba therapy. PMID:24468784

  6. Diagnosis and treatment outcome of mycotic keratitis at a tertiary eye care center in eastern india

    PubMed Central

    2011-01-01

    Background Mycotic keratitis is an important cause of corneal blindness world over including India. Geographical location and climate are known to influence the profile of fungal diseases. While there are several reports on mycotic keratitis from southern India, comprehensive clinico-microbiological reports from eastern India are few. The reported prevalence of mycotic keratitis are 36.7%,36.3%,25.6%,7.3% in southern, western, north- eastern and northern India respectively. This study reports the epidemiological characteristics, microbiological diagnosis and treatment outcome of mycotic keratitis at a tertiary eye care center in eastern India. Methods A retrospective review of medical and microbiology records was done for all patients with laboratory proven fungal keratitis. Results Between July 2006 and December 2009, 997 patients were clinically diagnosed as microbial keratitis. While no organisms were found in 25.4% (253/997) corneal samples, 23.4% (233/997) were bacterial, 26.4% (264/997) were fungal (45 cases mixed with bacteria), 1.4% (14/997) were Acanthamoeba with or without bacteria and 23.4% (233/997) were microsporidial with or without bacteria. Two hundred fifteen of 264 (81.4%, 215/264) samples grew fungus in culture while 49 corneal scrapings were positive for fungal elements only in direct microscopy. Clinical diagnosis of fungal keratitis was made in 186 of 264 (70.5%) cases. The microscopic detection of fungal elements was achieved by 10% potassium hydroxide with 0.1% calcoflour white stain in 94.8%(238/251) cases. Aspergillus species (27.9%, 60/215) and Fusarium species (23.2%, 50/215) were the major fungal isolates. Concomitant bacterial infection was seen in 45 (17.1%, 45/264) cases of mycotic keratitis. Clinical outcome of healed scar was achieved in 94 (35.6%, 94/264) cases. Fifty two patients (19.7%, 52/264) required therapeutic PK, 9 (3.4%, 9/264) went for evisceration, 18.9% (50/264) received glue application with bandage contact lens (BCL

  7. Scleritis and Peripheral Ulcerative Keratitis

    PubMed Central

    Galor, Anat; Thorne, Jennifer E.

    2008-01-01

    Scleritis and peripheral ulcerative keratitis (PUK) can present as isolated conditions or as part of a systemic inflammatory or infectious disorder. Both are serious ocular conditions that can result in vision loss and therefore require early diagnosis and treatment. Nearly two-thirds of patients with non-infectious scleritis require systemic glucocorticoid therapy, and one fourth need a glucocorticoid-sparing agent, as well. Essentially all patients with non-infectious PUK require systemic glucocorticoids. Detailed clinical history, thorough physical examination, and thoughtful laboratory evaluations are all important in the exclusion of underlying disorders and extraocular involvement. PMID:18037120

  8. Corneal ulcers and infections

    MedlinePlus

    Bacterial keratitis; Fungal keratitis; Acanthamoeba keratitis; Herpes simplex keratitis ... occur in people with a suppressed immune system. Herpes simplex keratitis is a serious viral infection. It ...

  9. Corneal cross linking and infectious keratitis: a systematic review with a meta-analysis of reported cases

    PubMed Central

    2013-01-01

    Background Collagen cross linking (CXL) of the cornea has been developed recently as a new treatment for multidrug-resistant infectious keratitis. The aim of this study is to summarize the previously published data and evaluate the effectiveness of this treatment. Results The search identified 12 articles. The number of eyes was 104. The infectious keratitis was associated with bacteria in 58 eyes (57f%): Gram-positive bacteria in 44 (43%; 4 of which were infected with Mycobacterium (3.6%)) and Gram-negative bacteria in 14 eyes (13%), fungus in 13 eyes (12%), and Acanthamoeba in 7 eyes (7%). In 26 eyes (25%), the microbiological culture was negative or not performed. The mean time of re-epithelization after CXL was 20.7±28.1 days (minimum of 3, maximum of 145). Sixteen eyes underwent deep or lamellar keratoplasty. The pooled analysis suggested that CXL has a favorable effect on the block of corneal melting in 85% (95%; CI 0.77, 0.91) of eyes. Conclusion Although randomized controlled trials are needed, the available evidence supports the use of CXL in the treatment of infectious keratitis. PMID:23718849

  10. Alternaria keratitis after deep anterior lamellar keratoplasty.

    PubMed

    Naik, Mekhla; Mohd Shahbaaz; Sheth, Jay; Sunderamoorthy, S K

    2014-01-01

    To describe a case of Alternaria keratitis in a 30-year-old male patient who presented with bilateral vascularised central corneal opacity and underwent deep anterior lamellar keratoplasty (DALK) in the left eye. Patient was treated for recurrent epithelial defect with a bandage contact lens in the follow-up visits after DALK. Subsequently, patient presented with pigmented fungal keratitis, which on culture examination of the corneal scrapping demonstrated Alternaria species. Patient had to undergo a repeat DALK as the keratitis did not resolve with medical therapy alone. Patient did not have a recurrence for 11 months following the regraft. This case report highlights the importance of considering the Alternaria species as a possibile cause of non-resolving fungal keratitis after DALK.

  11. Some secrets are revealed: parasitic keratitis amoebae as vectors of the scarcely described pandoraviruses to humans.

    PubMed

    Scheid, Patrick; Balczun, Carsten; Schaub, Günter A

    2014-10-01

    In this article, the results of a long effort to derive valuable phylogenetic data about an extraordinary spore-like infectious particle (endocytobiont) within host amoebae (Acanthamoeba sp.) recently isolated from the contact lens and the inflamed eye of a patient with keratitis are presented. The development of these endocytobionts has already been demonstrated with electron microscopic photo sequences, leading to a relevant model of its development presented here. The molecular biological investigation following the discovery of two other Pandoravirus species within aquatic sediments in 2013 led to the taxonomic affiliation of our endocytobiont with the genus Pandoravirus. A range of endocytobionts (intracellular biofilms) have been found in recent years, among which are several viruses which obligatorily proliferate within free-living amoebae. In human medicine, foreign objects which are placed in or on humans cause problems with microorganisms in biofilms. Contact lenses are especially important, because they are known as a source of a rapid formation of biofilm. These were the first Pandoraviruses described, and because this is additionally the first documented association with humans, we have clearly demonstrated how easily such (viral) endocytobionts can be transferred to humans. This case counts as an example of parasites acting as vectors of phylogenetically different microorganisms especially when living sympatric within their biocoenosis of biofilms. As the third part of the "Pandoravirus trilogy", it finally reveals the phylogenetic nature of these "extraordinary endocytobionts" within Acanthamoebae.

  12. Some secrets are revealed: parasitic keratitis amoebae as vectors of the scarcely described pandoraviruses to humans.

    PubMed

    Scheid, Patrick; Balczun, Carsten; Schaub, Günter A

    2014-10-01

    In this article, the results of a long effort to derive valuable phylogenetic data about an extraordinary spore-like infectious particle (endocytobiont) within host amoebae (Acanthamoeba sp.) recently isolated from the contact lens and the inflamed eye of a patient with keratitis are presented. The development of these endocytobionts has already been demonstrated with electron microscopic photo sequences, leading to a relevant model of its development presented here. The molecular biological investigation following the discovery of two other Pandoravirus species within aquatic sediments in 2013 led to the taxonomic affiliation of our endocytobiont with the genus Pandoravirus. A range of endocytobionts (intracellular biofilms) have been found in recent years, among which are several viruses which obligatorily proliferate within free-living amoebae. In human medicine, foreign objects which are placed in or on humans cause problems with microorganisms in biofilms. Contact lenses are especially important, because they are known as a source of a rapid formation of biofilm. These were the first Pandoraviruses described, and because this is additionally the first documented association with humans, we have clearly demonstrated how easily such (viral) endocytobionts can be transferred to humans. This case counts as an example of parasites acting as vectors of phylogenetically different microorganisms especially when living sympatric within their biocoenosis of biofilms. As the third part of the "Pandoravirus trilogy", it finally reveals the phylogenetic nature of these "extraordinary endocytobionts" within Acanthamoebae. PMID:25033816

  13. [Contact lens-related keratitis].

    PubMed

    Steiber, Zita; Berta, András; Módis, László

    2013-11-10

    Nowadays, keratitis, corneal infection due to wearing contact lens means an increasingly serious problem. Neglected cases may lead to corneal damage that can cause blindness in cases of otherwise healthy eyes. Early diagnosis based on the clinical picture and the typical patient history is an important way of prevention. Prophylaxis is substantial to avoid bacterial and viral infection that is highly essential in this group of diseases. Teaching contact lens wearers the proper contact lens care, storage, sterility, and hygiene regulations is of great importance. In case of corneal inflammation early accurate diagnosis supported by microbiological culture from contact lenses, storage boxes or cornea is very useful. Thereafter, targeted drug therapy or in therapy-resistant cases surgical treatment may even be necessary in order to sustain suitable visual acuity.

  14. Cellulose degradation: a therapeutic strategy in the improved treatment of Acanthamoeba infections.

    PubMed

    Lakhundi, Sahreena; Siddiqui, Ruqaiyyah; Khan, Naveed Ahmed

    2015-01-01

    Acanthamoeba is an opportunistic free-living amoeba that can cause blinding keratitis and fatal brain infection. Early diagnosis, followed by aggressive treatment is a pre-requisite in the successful treatment but even then the prognosis remains poor. A major drawback during the course of treatment is the ability of the amoeba to enclose itself within a shell (a process known as encystment), making it resistant to chemotherapeutic agents. As the cyst wall is partly made of cellulose, thus cellulose degradation offers a potential therapeutic strategy in the effective targeting of trophozoite encased within the cyst walls. Here, we present a comprehensive report on the structure of cellulose and cellulases, as well as known cellulose degradation mechanisms with an eye to target the Acanthamoeba cyst wall. The disruption of the cyst wall will make amoeba (concealed within) susceptible to chemotherapeutic agents, and at the very least inhibition of the excystment process will impede infection recurrence, as we bring these promising drug targets into focus so that they can be explored to their fullest.

  15. Molecular characterization of Acanthamoeba strains isolated from domestic dogs in Tenerife, Canary Islands, Spain.

    PubMed

    Valladares, María; Reyes-Batlle, María; Martín-Navarro, Carmen M; López-Arencibia, Atteneri; Dorta-Gorrín, Alexis; Wagner, Carolina; Martínez-Carretero, Enrique; Piñero, José E; Valladares, Basilio; Lorenzo-Morales, Jacob

    2015-06-01

    The present study describes two cases of Acanthamoeba infections (keratitis and ascites/peritonitis) in small breed domestic dogs in Tenerife, Canary Islands, Spain. In both cases, amoebic trophozoites were observed under the inverted microscope and isolated from the infected tissues and/or fluids, without detecting the presence of other viral, fungal or bacterial pathogens. Amoebae were isolated using 2 % non-nutrient agar plates and axenified for further biochemical and molecular analyses. Osmotolerance and thermotolerance assays revealed that both isolates were able to grow up to 37 °C and 1 M of mannitol and were thus considered as potentially pathogenic. Moreover, the strains were classified as highly cytotoxic as they cause more than 75 % of toxicity when incubated with two eukaryotic cell lines. In order to classify the strains at the molecular level, the diagnostic fragment 3 (DF3) region of the 18S rDNA of Acanthamoeba was amplified and sequenced, revealing that both isolates belonged to genotype T4. In both cases, owners of the animals did not allow any further studies or follow-up and therefore the current status of these animals is unknown. Furthermore, the isolation of these pathogenic amoebae should raise awareness with the veterinary community locally and worldwide.

  16. Recent advances in diagnosis and management of Mycotic Keratitis

    PubMed Central

    Maharana, Prafulla K; Sharma, Namrata; Nagpal, Ritu; Jhanji, Vishal; Das, Sujata; Vajpayee, Rasik B

    2016-01-01

    Mycotic keratitis is a major cause of corneal blindness, especially in tropical and subtropical countries. The prognosis is markedly worse compared to bacterial keratitis. Delayed diagnosis and scarcity of effective antifungal agents are the major factors for poor outcome. Over the last decade, considerable progress has been made to rapidly diagnose cases with mycotic keratitis and increase the efficacy of treatment. This review article discusses the recent advances in diagnosis and management of mycotic keratitis with a brief discussion on rare and emerging organisms. A MEDLINE search was carried out for articles in English language, with the keywords, mycotic keratitis, fungal keratitis, emerging or atypical fungal pathogens in mycotic keratitis, investigations in mycotic keratitis, polymerase chain reaction in mycotic keratitis, confocal microscopy, treatment of mycotic keratitis, newer therapy for mycotic keratitis. All relevant articles were included in this review. Considering the limited studies available on newer diagnostic and therapeutic modalities in mycotic keratitis, case series as well as case reports were also included if felt important. PMID:27380973

  17. MOLECULAR AND PHYSIOLOGICAL EVALUATION OF SUBTROPICAL ENVIRONMENTAL ISOLATES OF ACANTHAMOEBA KERATITIS. (R828830)

    EPA Science Inventory

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  18. Rethinking contact lens associated keratitis.

    PubMed

    Efron, Nathan; Morgan, Philip B

    2006-09-01

    This review presents a critical analysis of the literature relating to the use of binomial and polynomial classification schemes for categorising corneal infiltrative events (CIEs) associated with contact lens wear and the epidemiology of such events. The results of the Manchester Keratitis Study-a 12-month, prospective, hospital-based epidemiological study of contact lens wearer suffering from CIEs-are used as a tool to challenge and test traditional thinking in relation to contact lens associated keratitis. An innovative aspect of this study is the use of a novel clinical severity matrix to systematically score the severity of CIEs based on 10 key signs and symptoms. The ambiguities inherent in using binomial classification schemes (such as, microbial versus sterile, ulcerative versus non-ulcerative etcetera) are highlighted. The failure of a polynomial scheme-due to extensive classification overlap between proposed sub-types of CIEs-is demonstrated using a Venn diagram. A cartographic analysis reveals that infiltrates tend to occur in the superior cornea of patients wearing extended wear silicone hydrogel lenses, in the central cornea of patients wearing daily wear hydrogel daily disposable lenses and in the peripheral cornea of patients wearing daily wear hydrogel (excluding daily disposable) lenses. Infiltrates that occur more towards the limbus are less severe. The incidence of CIEs is higher when contact lenses are worn overnight. Logistic analysis reveals that the risk of developing a severe CIE is five times greater with conventional hydrogel extended wear versus silicone hydrogel extended wear. The male gender, smoking, a healthy eye and body, and the late Winter months are associated with an increased risk of developing CIEs. The rate of significant visual loss as a result of developing a CIE is low. Two key conclusions are drawn from this work, which represent a radical rethinking of this potentially sight-threatening condition. CIEs should be

  19. Unexpected postmortem diagnosis of acanthamoeba meningoencephalitis in an immunocompetent child

    PubMed Central

    Binesh, Fariba; Karimi, Mehran; Navabii, Hossein

    2011-01-01

    Meningoencephalitis caused by Acanthamoeba spp. is a rare opportunistic infection, difficult to diagnose and treat, which causes death in almost all cases. Here, the authors report a 5-year-old Iranian immunocompetent girl who died of fulminant acanthamoeba meningoencephalitis. To the authors’ knowledge, this is the first case of acanthamoeba meningoencephalitis in Iran. PMID:22679147

  20. Targeting Herpetic Keratitis by Gene Therapy

    PubMed Central

    Elbadawy, Hossein Mostafa; Gailledrat, Marine; Desseaux, Carole; Ponzin, Diego; Ferrari, Stefano

    2012-01-01

    Ocular gene therapy is rapidly becoming a reality. By November 2012, approximately 28 clinical trials were approved to assess novel gene therapy agents. Viral infections such as herpetic keratitis caused by herpes simplex virus 1 (HSV-1) can cause serious complications that may lead to blindness. Recurrence of the disease is likely and cornea transplantation, therefore, might not be the ideal therapeutic solution. This paper will focus on the current situation of ocular gene therapy research against herpetic keratitis, including the use of viral and nonviral vectors, routes of delivery of therapeutic genes, new techniques, and key research strategies. Whereas the correction of inherited diseases was the initial goal of the field of gene therapy, here we discuss transgene expression, gene replacement, silencing, or clipping. Gene therapy of herpetic keratitis previously reported in the literature is screened emphasizing candidate gene therapy targets. Commonly adopted strategies are discussed to assess the relative advantages of the protective therapy using antiviral drugs and the common gene therapy against long-term HSV-1 ocular infections signs, inflammation and neovascularization. Successful gene therapy can provide innovative physiological and pharmaceutical solutions against herpetic keratitis. PMID:23326647

  1. Congenital insensitivity to pain with neuroparalytic keratitis.

    PubMed

    Biedner, B; Dagan, M; Gedalia, A; David, R

    1990-08-01

    Congenital insensitivity to pain is a well-defined entity in the group of sensory deficiency syndromes. To the best of our knowledge, unilateral neuroparalytic keratitis associated with congenital insensitivity to pain has not been reported. We report such a case to alert clinicians to this potentially blinding problem.

  2. Update: Fusarium Keratitis - United States, 2005 - 2006

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This report describes the results of a Fusarium keratitis outbreak investigation being conducted by the United States Centers for Disease Control and Prevention. The epidemiological data indicate that the 2005-2006 outbreaks of corneal infections within the United States are linked to the use of on...

  3. Lacrimal Duct Occlusion Is Associated with Infectious Keratitis

    PubMed Central

    Li, Guigang; Guo, Jingmin; Liu, Rong; Hu, Weikun; Xu, Lingjuan; Wang, Juan; Cai, Subo; Zhang, Hong; Zhu, Yingting

    2016-01-01

    Background: To explore the prevalence of lacrimal duct obstruction in patients with infectious keratitis, and the necessity of lacrimal duct dredge in the treatment of human infectious keratitis. Methodology/Principle Findings: The design is prospective, non-control case series. Thirty-one eyes from twenty-eight continuous patients with infectious keratitis were included in this study. The presence/absence of lacrimal duct obstruction was determined by the lacrimal duct irrigation test. The diagnosis of infectious keratitis was made based on clinical manifestations, cornea scraping microscopic examination and bacterial/fungus culture. Diagnosis of viral keratitis was set up based on the recurrent history, deep neovascularization and typical outlook of the cornea scar. The treatment of keratitis included drugs, eye drops or surgery, while treatment of chronic dacryocystitis was lacrimal duct dredging with supporting tube implantation surgery. In the thirty-one eyes with infectious keratitis, fifteen suffered from fungal keratitis (48%), two bacterial keratitis (6%), and fourteen viral keratitis (45%). Eleven eyes (35%) from ten patients with infectious keratitis also suffered from lacrimal duct obstruction. In those cases, six eyes also suffered from lower canalicular obstruction, three nasolacrimal duct obstruction and chronic dacryocystitis, one a combination of upper and lower canalicular obstruction, one upper canalicular obstruction. After local and systemic applications of anti-bacterial, anti-viral, anti-fungal and anti-inflammatory drugs, twenty-eight eyes (90%) recovered within three weeks, while the ulceration of three patients required the lacrimal duct dredging and supporting tube implantation surgery for the healing. Conclusions: Herein, we first report that the prevalence of infectious keratitis is closely correlated to the occurrence of lacrimal duct obstruction. When both confirmed, simultaneous treatment of keratitis and lacrimal duct obstruction

  4. Effect of non-steroidal anti-inflammatory drugs on biological properties of Acanthamoeba castellanii belonging to the T4 genotype.

    PubMed

    Siddiqui, Ruqaiyyah; Lakhundi, Sahreena; Iqbal, Junaid; Khan, Naveed Ahmed

    2016-09-01

    Non-steroidal anti-inflammatory drug, Diclofenac, targeting COX have shown promise in the treatment of Acanthamoeba keratitis, but the underlying mechanisms remain unknown. Using various NSAIDs, Diclofenac sodium, Indomethacin, and Acetaminophen, here we determined the effects of NSAIDs on the biological properties of Acanthamoeba castellanii belonging to the T4 genotype. Using amoebicidal assays, the results revealed that Diclofenac sodium, and Indomethacin affected growth of A. castellanii. In contrast, none of the compounds tested had any effect on the viability of A. castellanii. Importantly, all NSAIDs tested abolished A. castellanii encystation. This is a significant finding as the ability of amoebae to transform into the dormant cyst form presents a significant challenge in the successful treatment of infection. The NSAIDs inhibit production of cyclo-oxegenase, which regulates the synthesis of prostaglandins suggesting that cyclooxygenases (COX-1 and COX-2) and prostaglandins play significant role(s) in Acanthamoeba biology. As NSAIDs are routinely used in the clinical practice, these findings may help design improved preventative strategies and/or of therapeutic value to improve prognosis, when used in combination with other anti-amoebic drugs. PMID:27381503

  5. Current Thoughts in Fungal Keratitis: Diagnosis and Treatment

    PubMed Central

    Ansari, Zubair; Miller, Darlene; Galor, Anat

    2013-01-01

    Fungal keratitis remains a challenging and often elusive diagnosis in geographic regions where it is endemic. Marred by delays in diagnosis, the sequelae of corneal fungal infections, though preventable, can be irreversible. Recent studies and advances in the arena have broadened the approach and treatment to mycotic keratitis. This review will discuss current diagnostic modalities of fungal keratitis and will particularly focus on treatment regimens. It will also explore future therapeutic models and critique the potential benefit of each. PMID:24040467

  6. Unilateral Punctate Keratitis Secondary to Wallenberg Syndrome

    PubMed Central

    Boto, Ana; Del Hierro, Almudena; Capote, Maria; Noval, Susana; Garcia, Amanda; Santiago, Susana

    2014-01-01

    We studied three patients who developed left unilateral punctate keratitis after suffering left-sided Wallenberg Syndrome. A complex evolution occurred in two of them. In all cases, neurophysiological studies showed damage in the trigeminal sensory component at the bulbar level. Corneal involvement secondary to Wallenberg syndrome is a rare cause of unilateral superficial punctate keratitis. The loss of corneal sensitivity caused by trigeminal neuropathy leads to epithelial erosions that are frequently unobserved by the patient, resulting in a high risk of corneal-ulcer development with the possibility of superinfection. Neurophysiological studies can help to locate the anatomical level of damage at the ophthalmic branch of the trigeminal nerve, confirming the suspected etiology of stroke, and demonstrating that prior vascular involvement coincides with the location of trigeminal nerve damage. In some of these patients, oculofacial pain is a distinctive feature. PMID:24882965

  7. Evaluation of the activity of new cationic carbosilane dendrimers on trophozoites and cysts of Acanthamoeba polyphaga.

    PubMed

    Heredero-Bermejo, Irene; Copa-Patiño, Jose Luis; Soliveri, Juan; Fuentes-Paniagua, Elena; de la Mata, Francisco Javier; Gomez, Rafael; Perez-Serrano, Jorge

    2015-02-01

    Dendrimers are repetitively branched molecules with a broad spectrum of applications, mainly for their antimicrobial properties and as nanocarriers for other molecules. Recently, our research group have synthesized and studied their activity against Acanthamoeba sp., causative agent of a severe ocular disease in humans: Acanthamoeba keratitis. New cationic carbosilane dendrimers were tested against the protozoa forms at different concentrations and for different incubation times. Trophozoite viability was determined by manual counting and cyst viability by observing excystment in microplates with fresh culture medium. Cytotoxicity was checked on HeLa cells using the microculture tetrazolium assay. Alterations were observed by optical microscopy and by flow cytometry staining with propidium iodide. Six out of the 18 dendrimers tested were non-cytotoxic and effective against the trophozoite form, having one of them (dendrimer 14 with an IC50 of 2.4 + 0.1 mg/L) a similar activity to chlorhexidine digluconate (IC50 1.7 + 0.1 mg/L). This dendrimer has a polyphenoxo core and a sulphur atom close to the six -NH3+ terminal groups. On the other hand, only two dendrimers showed some effect against cysts (dendrimers 14 and 17). However, their minimum cysticidal concentrations were cytotoxic and less effective than the control drug. The alterations on the amoeba morphology produced by the treatment with dendrimers were size reduction, increased complexity, loss of acanthopodia and cell membrane disruption. In conclusion, these results suggest that some dendrimers may be studied in animal models to test their effect and that new dendrimers with similar features should be synthesized. PMID:25358240

  8. First aid for complications of infectious keratitis

    PubMed Central

    2008-01-01

    Infectious keratitis is a fairly common entity in India. However while paying attention to the primary entity, the associated events may be overlooked. Enhanced pain usually suggests a worsening of the condition or development of associated problems like secondary glaucoma. However, contrary to logic, a sudden decrease in pain is also liley to suggest a worsening, e.g. perforation of the corneal ulcer. Various such problems with their management are outlined. PMID:18417823

  9. Contact lens care solution killing efficacy against Acanthamoeba castellanii by in vitro testing and live-imaging.

    PubMed

    Kolar, Satya Sree N; Manarang, Joseph C; Burns, Alan R; Miller, William L; McDermott, Alison M; Bergmanson, Jan P G

    2015-12-01

    In the past decade there has been an increased incidence of Acanthamoeba keratitis, particularly in contact lens wearers. The aim of this study was to utilize in vitro killing assays and to establish a novel, time-lapse, live-cell imaging methodology to demonstrate the efficacy of contact lens care solutions in eradicating Acanthamoeba castellanii (A. castellanii) trophozoites and cysts. Standard qualitative and quantitative in vitro assays were performed along with novel time-lapse imaging coupled with fluorescent dye staining that signals cell death. Quantitative data obtained demonstrated that 3% non-ophthalmic hydrogen peroxide demonstrated the highest percent killing at 87.4% corresponding to a 4.4 log kill. The other contact lens care solutions which showed a 72.9 to 29.2% killing which was consistent with 4.3-2.8 log reduction in trophozoite viability. Both analytical approaches revealed that polyquaternium/PHMB-based was the least efficacious in terms of trophicidal activity. The cysticidal activity of the solutions was much less than activity against trophozoites and frequently was not detected. Live-imaging provided a novel visual endpoint for characterizing the trophocidal activity of the care solutions. All solutions caused rapid rounding or pseudocyst formation of the trophozoites, reduced motility and the appearance of different morphotypes. Polyquaternium/alexidine-based and peroxide-based lens care system induced the most visible damage indicated by significant accumulation of debris from ruptured cells. Polyquaternium/PHMB-based was the least effective showing rounding of the cells but minimal death. These observations are in keeping with care solution biocides having prominent activity at the plasma membrane of Acanthamoeba.

  10. Contact lens care solution killing efficacy against Acanthamoeba castellanii by in vitro testing and live-imaging.

    PubMed

    Kolar, Satya Sree N; Manarang, Joseph C; Burns, Alan R; Miller, William L; McDermott, Alison M; Bergmanson, Jan P G

    2015-12-01

    In the past decade there has been an increased incidence of Acanthamoeba keratitis, particularly in contact lens wearers. The aim of this study was to utilize in vitro killing assays and to establish a novel, time-lapse, live-cell imaging methodology to demonstrate the efficacy of contact lens care solutions in eradicating Acanthamoeba castellanii (A. castellanii) trophozoites and cysts. Standard qualitative and quantitative in vitro assays were performed along with novel time-lapse imaging coupled with fluorescent dye staining that signals cell death. Quantitative data obtained demonstrated that 3% non-ophthalmic hydrogen peroxide demonstrated the highest percent killing at 87.4% corresponding to a 4.4 log kill. The other contact lens care solutions which showed a 72.9 to 29.2% killing which was consistent with 4.3-2.8 log reduction in trophozoite viability. Both analytical approaches revealed that polyquaternium/PHMB-based was the least efficacious in terms of trophicidal activity. The cysticidal activity of the solutions was much less than activity against trophozoites and frequently was not detected. Live-imaging provided a novel visual endpoint for characterizing the trophocidal activity of the care solutions. All solutions caused rapid rounding or pseudocyst formation of the trophozoites, reduced motility and the appearance of different morphotypes. Polyquaternium/alexidine-based and peroxide-based lens care system induced the most visible damage indicated by significant accumulation of debris from ruptured cells. Polyquaternium/PHMB-based was the least effective showing rounding of the cells but minimal death. These observations are in keeping with care solution biocides having prominent activity at the plasma membrane of Acanthamoeba. PMID:26208952

  11. Pattern recognition receptors in microbial keratitis.

    PubMed

    Taube, M-A; del Mar Cendra, M; Elsahn, A; Christodoulides, M; Hossain, P

    2015-11-01

    Microbial keratitis is a significant cause of global visual impairment and blindness. Corneal infection can be caused by a wide variety of pathogens, each of which exhibits a range of mechanisms by which the immune system is activated. The complexity of the immune response to corneal infection is only now beginning to be elucidated. Crucial to the cornea's defences are the pattern-recognition receptors: Toll-like and Nod-like receptors and the subsequent activation of inflammatory pathways. These inflammatory pathways include the inflammasome and can lead to significant tissue destruction and corneal damage, with the potential for resultant blindness. Understanding the immune mechanisms behind this tissue destruction may enable improved identification of therapeutic targets to aid development of more specific therapies for reducing corneal damage in infectious keratitis. This review summarises current knowledge of pattern-recognition receptors and their downstream pathways in response to the major keratitis-causing organisms and alludes to potential therapeutic approaches that could alleviate corneal blindness.

  12. Photodynamic inactivation of pathogens causing infectious keratitis

    NASA Astrophysics Data System (ADS)

    Simon, Carole; Wolf, G.; Walther, M.; Winkler, K.; Finke, M.; Hüttenberger, D.; Bischoff, Markus; Seitz, B.; Cullum, J.; Foth, H.-J.

    2014-03-01

    The increasing prevalence of antibiotic resistance requires new approaches also for the treatment of infectious keratitis. Photodynamic Inactivation (PDI) using the photosensitizer (PS) Chlorin e6 (Ce6) was investigated as an alternative to antibiotic treatment. An in-vitro cornea model was established using porcine eyes. The uptake of Ce6 by bacteria and the diffusion of the PS in the individual layers of corneal tissue were investigated by fluorescence. After removal of the cornea's epithelium Ce6-concentrations < 1 mM were sufficient to reach a penetration depth of 500 μm. Liquid cultures of microorganisms were irradiated using a specially constructed illumination chamber made of Spectralon(R) (reflectance: 99 %), which was equipped with high power light emitting diodes (λ = 670 nm). Clinical isolates of Staphylococcus aureus (SA) and Pseudomonas aeruginosa (PA) from keratitis patients were tested in liquid culture against different concentrations of Ce6 (1 - 512 μM) using 10 minutes irradiation (E = 18 J/cm2 ). This demonstrated that a complete inactivation of the pathogen strains were feasible whereby SA was slightly more susceptible than PA. 3909 mutants of the Keio collection of Escherichia coli (E.coli) were screened for potential resistance factors. The sensitive mutants can be grouped into three categories: transport mutants, mutants in lipopolysaccharide synthesis and mutants in the bacterial SOS-response. In conclusion PDI is seen as a promising therapy concept for infectious keratitis.

  13. Is there evidence of sexual reproduction (meiosis) in Acanthamoeba?

    PubMed Central

    Khan, Naveed A.; Siddiqui, Ruqaiyyah

    2015-01-01

    Evolution of independently breeding species into males and females (gametes) has remained a puzzle. Given the significant advantages of sexual reproduction over asexual reproduction as a long-term species survival strategy; here, we pose the question whether there is some form of meiosis in Acanthamoeba species, which represents our ancient lineage. The recently available Acanthamoeba genome revealed several genes implicated in meiosis in sexual eukaryotes such as Spo11, Mre11, Rad50, Rad51, Rad52, Mnd1, Dmc1, Msh, and Mlh, suggesting that Acanthamoeba is capable of some form of meiosis, inferring the presence of sexual reproduction in Acanthamoeba, and that meiosis evolved early in eukaryotic evolution. PMID:25800982

  14. Is there evidence of sexual reproduction (meiosis) in Acanthamoeba?

    PubMed

    Khan, Naveed A; Siddiqui, Ruqaiyyah

    2015-06-01

    Evolution of independently breeding species into males and females (gametes) has remained a puzzle. Given the significant advantages of sexual reproduction over asexual reproduction as a long-term species survival strategy; here, we pose the question whether there is some form of meiosis in Acanthamoeba species, which represents our ancient lineage. The recently available Acanthamoeba genome revealed several genes implicated in meiosis in sexual eukaryotes such as Spo11, Mre11, Rad50, Rad51, Rad52, Mnd1, Dmc1, Msh, and Mlh, suggesting that Acanthamoeba is capable of some form of meiosis, inferring the presence of sexual reproduction in Acanthamoeba, and that meiosis evolved early in eukaryotic evolution.

  15. Amebic meningoencephalitis caused by Acanthamoeba castellani in a dog.

    PubMed

    Pearce, J R; Powell, H S; Chandler, F W; Visvesvara, G S

    1985-11-01

    A natural infection of Acanthamoeba castellani, a free-living ameba, was determined to be the cause of acute, hemorrhagic, necrotizing amebic meningoencephalitis in a dog. This case is unique because previous reports of infection by the Acanthamoeba spp in dogs have not indicated its presence in the brain. Naturally developing meningoencephalitis by Acanthamoeba spp in the dog may have a pathogenesis similar to that of human beings. The ameba in this case also was observed in the lungs and kidneys, which are believed to be the primary sites of lesions in human beings that develop amebic meningoencephalitis from Acanthamoeba spp. PMID:3932273

  16. Identification of Paenibacillus as a Symbiont in Acanthamoeba.

    PubMed

    Maschio, Vinicius José; Corção, Gertrudes; Bücker, Francielle; Caumo, Karin; Rott, Marilise Brittes

    2015-09-01

    Amoebae of the genus Acanthamoeba occur worldwide and in addition to being pathogens, are important vehicles for microorganisms with clinical and environmental importance. This study aimed to evaluate the profiling of endosymbionts in 12 isolates of Acanthamoeba using V3 region of 16S rDNA denaturing gradient gel electrophoresis (DGGE) and sequencing. The DGGE enabled us to characterize the endosymbionts diversity in isolates of Acanthamoeba, and to identify Paenibacillus sp., an emerging pathogen, as an amoebic endosymbiont. The results of this study demonstrated that Acanthamoeba is capable of transporting a large number of endosymbionts. This is the first study that reports, the presence of Paenibacillus sp. as amebic symbiont.

  17. Area 51: How do Acanthamoeba invade the central nervous system?

    PubMed

    Siddiqui, Ruqaiyyah; Emes, Richard; Elsheikha, Hany; Khan, Naveed Ahmed

    2011-05-01

    Acanthamoeba granulomatous encephalitis generally develops as a result of haematogenous spread, but it is unclear how circulating amoebae enter the central nervous system (CNS) and cause inflammation. At present, the mechanisms which Acanthamoeba use to invade this incredibly well-protected area of the CNS and produce infection are not well understood. In this paper, we propose two key virulence factors: mannose-binding protein and extracellular serine proteases as key players in Acanthamoeba traversal of the blood-brain barrier leading to neuronal injury. Both molecules should provide excellent opportunities as potential targets in the rational development of therapeutic interventions against Acanthamoeba encephalitis.

  18. The role of Src kinase in the biology and pathogenesis of Acanthamoeba castellanii

    PubMed Central

    2012-01-01

    Background Acanthamoeba species are the causative agents of fatal granulomatous encephalitis in humans. Haematogenous spread is thought to be a primary step, followed by blood–brain barrier penetration, in the transmission of Acanthmaoeba into the central nervous system, but the associated molecular mechanisms remain unclear. Here, we evaluated the role of Src, a non-receptor protein tyrosine kinase in the biology and pathogenesis of Acanthamoeba. Methods Amoebistatic and amoebicidal assays were performed by incubating amoeba in the presence of Src kinase-selective inhibitor, PP2 (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine) and its inactive analog, PP3 (4-amino-7-phenylpyrazolo[3,4-d]pyrimidine). Using this inhibitor, the role of Src kinase in A. castellanii interactions with Escherichia coli was determined. Zymographic assays were performed to study effects of Src kinase on extracellular proteolytic activities of A. castellanii. The human brain microvascular endothelial cells were used to determine the effects of Src kinase on A. castellanii adhesion to and cytotoxicity of host cells. Results Inhibition of Src kinase using a specific inhibitor, PP2 (4-amino-5-(4 chlorophenyl)-7-(t-butyl)pyrazolo [3,4-d] pyrimidine) but not its inactive analog, PP3 (4-amino-7-phenylpyrazolo[3,4-d] pyrimidine), had detrimental effects on the growth of A. castellanii (keratitis isolate, belonging to the T4 genotype). Interestingly, inhibition of Src kinase hampered the phagocytic ability of A. castellanii, as measured by the uptake of non-invasive bacteria, but, on the contrary, invasion by pathogenic bacteria was enhanced. Zymographic assays revealed that inhibition of Src kinases reduced extracellular protease activities of A. castellanii. Src kinase inhibition had no significant effect on A. castellanii binding to and cytotoxicity of primary human brain microvascular endothelial cells, which constitute the blood–brain barrier. Conclusions For the first

  19. Acanthamoeba Encephalitis in Patient with Systemic Lupus, India

    PubMed Central

    Shirwadkar, Charudatt G.; Samant, Rohini; Sankhe, Milind; Deshpande, Ramesh; Yagi, Shigeo; Schuster, Frederick L.; Sriram, Rama

    2006-01-01

    We report a fatal case of encephalitis caused by Acanthamoeba in a 24-year-old woman from India with systemic lupus erythematosus. Diagnosis was made by identification of amebas in brain sections by immunofluorescence analysis and confirmed by demonstrating Acanthamoeba mitochondrial 16S rRNA gene DNA in brain tissue sections. PMID:16707057

  20. Occurrence of Potentially Pathogenic Bacterial-Endosymbionts in Acanthamoeba Spp.

    PubMed Central

    NIYYATI, Maryam; MAFI, Mahyar; HAGHIGHI, Ali; HAKEMI VALA, Mojdeh

    2015-01-01

    Background: Acanthamoeba- bacteria interactions enable pathogenic bacteria to tolerate harsh conditions and lead to transmission to the susceptible host. The present study was aimed to address the presence of bacterial endosymbionts of Acanthamoeba isolated from recreational water sources of Tehran, Iran. To the best of our knowledge this is the first study regarding occurrence of bacteria in environmental Acanthamoeba spp. in Iran. Methods: A total of 75 samples of recreational water sources were collected. Samples were cultured on non- nutrient agar 1.5% plates. Positive Acanthamoeba spp. were axenically grown. DNA extraction and PCR reaction was performed using JDP1-2 primers. All positive samples of Acanthamoeba were examined for the presence of endosymbionts using staining and molecular methods. The PCR products were then sequenced in order to determine the genotypes of Acanthamoeba and bacteria genera. Results: Out of 75 samples, 16 (21.3%) plates were positive for Acanthamoeba according to the morphological criteria. Molecular analysis revealed that Acanthamoeba belonged to T4 and T5 genotypes. Five isolates (35.7%) were positive for bacterial endosymbionts using staining method and PCR test. Sequencing of PCR products confirmed the presence of Pseudomonas aeruginosa and Agrobacterium tumefasiens. Conclusion: The presence of Acanthamoeba bearing pathogenic endosymbionts in water sources leads us to public health issues including improved sanitation and decontamination measures in recreational water sources in order to prevent amoebae-related infection. To the best of our knowledge this is the first report regarding the isolation of A. tumefasiens from Acanthamoeba in Iran and worldwide. PMID:26246815

  1. Clinical and epidemiological characteristics of infectious keratitis in Paraguay.

    PubMed

    Nentwich, Martin M; Bordón, M; di Martino, D Sánchez; Campuzano, A Ruiz; Torres, W Martínez; Laspina, F; Lichi, S; Samudio, M; Farina, N; Sanabria, Rosa R; de Kaspar, Herminia Mino

    2015-06-01

    To describe the clinical and epidemiological characteristics of patients with severe infectious keratitis in Asunción, Paraguay between April 2009 and September 2011. All patients with the clinical diagnosis of severe keratitis (ulcer ≥2 mm in size and/or central location) were included. Empiric treatment consisted of topical antibiotics and antimycotics; in cases of advanced keratitis, fortified antibiotics were used. After microbiological analysis, treatment was changed if indicated. In total 48 patients (62.5 % males, 25 % farmers) were included in the analysis. A central ulcer was found in 81.3 % (n = 39). The median delay between onset of symptoms and time of first presentation at our institution was 7 days (range 1-30 days). Fungal keratitis was diagnosed in 64.5 % (n = 31) of patients, of which Fusarium sp. (n = 17) was the most common. Twenty-one patients (43.8 %) reported previous trauma to the eye. The globe could be preserved in all cases. While topical therapy only was sufficient in most patients, a conjunctival flap was necessary in six patients suffering from fungal keratitis. The high rate of fungal keratitis in this series is remarkable, and microbiological analysis provided valuable information for the appropriate treatment. In this setting, one has to be highly suspicious of fungal causes of infectious keratitis.

  2. Keratitis-Ichthyosis-Deafness syndrome: A rare congenital disorder

    PubMed Central

    Shanker, Vinay; Gupta, Mudita; Prashar, Aditi

    2012-01-01

    Keratitis-Icthyosis-Deafness syndrome is a rare congenital disorder characterized by keratitis, ichthyosis, and deafness. We report a 13 year old female child who presented with diffuse alopecia of the scalp and body. There was erythrokeratoderma of face and discrete hyperkeratotic hyperpigmented papulo plaque lesions on the body. Patient also had reticulate hyperkeratosis of palms and soles. There was history of recurrent episodes of folliculitis over the scalp and body. There was no evidence of any malignancy. Eye involvement in the form of bilateral vascularising keratitis was present. There was bilateral mixed hearing loss. PMID:23130264

  3. [Confocal microscopy for the diagnostics of fungal keratitis].

    PubMed

    Daas, L; Viestenz, A; Bischoff, M; Hasenfus, A; Seitz, B

    2016-09-01

    Fungal keratitis is a rare but very serious eye disease in industrial nations with a frequency of 1-5 % of all forms of keratitis from microbial causes. We present two patients with keratitis of primary unknown cause. Using confocal microscopy fungal filaments could be identified that partially showed a parallel configuration (like "railway tracks"). Thus, the correct diagnosis can often be made and suitable therapy can be non-invasively initiated even before the results of in vitro cultivation (fungal culture), polymerase chain reaction (PCR) and histological investigations are available.

  4. Multidrug-resistant Fusarium keratitis: diagnosis and treatment considerations.

    PubMed

    Sara, Sergio; Sharpe, Kendall; Morris, Sharon

    2016-01-01

    Mycotic keratitis is an ocular infective process derived from any fungal species capable of corneal invasion. Despite its rarity in developed countries, its challenging and elusive diagnosis may result in keratoplasty or enucleation following failed medical management. Filamentous fungi such as Fusarium are often implicated in mycotic keratitis. Bearing greater morbidity than its bacterial counterpart, mycotic keratitis requires early clinical suspicion and initiation of antifungal therapy to prevent devastating consequences. We describe a case of multidrug-resistant mycotic keratitis in a 46-year-old man who continued to decline despite maximal therapy and therapeutic keratoplasty. Finally, enucleation was performed as a means of source control preventing dissemination of a likely untreatable fungal infection into the orbit. Multidrug-resistant Fusarium is rare, and may progress to endophthalmitis. We discuss potential management options which may enhance diagnosis and outcome in this condition. PMID:27489066

  5. Geographic Disparities in the Etiology of Bacterial and Fungal Keratitis in the United States of America.

    PubMed

    Estopinal, Christopher B; Ewald, Mark D

    2016-01-01

    Infectious keratitis is a serious cause of vision loss. Proper treatment of infectious keratitis requires antimicrobials that target the organism responsible for a patient's ulcer. The frequency of infection by a given organism varies by location. We examined the literature to determine geographic disparities in the etiology of bacterial and fungal keratitis in the United States of America. Bacterial keratitis makes up a greater proportion of cases in northern locations, and fungal keratitis increases in prevalence in southern locations. Gram-negative organisms make up a greater proportion of bacterial keratitis in southern locations when compared to northern locations.

  6. Diversity of Microbial Species Implicated in Keratitis: A Review

    PubMed Central

    Karsten, Elisabeth; Watson, Stephanie Lousie; Foster, Leslie John Ray

    2012-01-01

    Background: Microbial keratitis is an infectious disease of the cornea characterised by inflammation and is considered an ophthalmic emergency requiring immediate attention. While a variety of pathogenic microbes associated with microbial keratitis have been identified, a comprehensive review identifying the diversity of species has not been completed. Methods: A search of peer-reviewed publications including case reports and research articles reporting microorganims implicated in keratitis was conducted. Search engines including PubMed, Scopus and Web of Science with years ranging from 1950-2012 were used. Results: 232 different species from 142 genera, representing 80 families were found to be implicated in microbial keratitis. Fungi exhibited the largest diversity with 144 species from 92 genera. In comparison, 77 species of bacteria from 42 genera, 12 species of protozoa from 4 genera and 4 types of virus were identified as the infectious agents. A comparison of their aetiologies shows reports of similarities between genera. Conclusions: The diversity of microbial species implicated in keratitis has not previously been reported and is considerably greater than suggested by incidence studies. Effective treatment is heavily reliant upon correct identification of the responsible microorganisms. Species identification, the risk factors associated with, and pathogenesis of microbial keratitis will allow the development of improved therapies. This review provides a resource for clinicians and researchers to assist in identification and readily source treatment information. PMID:23248737

  7. Gene discovery in the Acanthamoeba castellanii genome

    SciTech Connect

    Anderson, Iain J.; Watkins, Russell F.; Samuelson, John; Spencer,David F.; Majoros, William H.; Gray, Michael W.; Loftus, Brendan J.

    2005-08-01

    Acanthamoeba castellanii is a free-living amoeba found in soil, freshwater, and marine environments and an important predator of bacteria. Acanthamoeba castellanii is also an opportunistic pathogen of clinical interest, responsible for several distinct diseases in humans. In order to provide a genomic platform for the study of this ubiquitous and important protist, we generated a sequence survey of approximately 0.5 x coverage of the genome. The data predict that A. castellanii exhibits a greater biosynthetic capacity than the free-living Dictyostelium discoideum and the parasite Entamoeba histolytica, providing an explanation for the ability of A. castellanii to inhabit adversity of environments. Alginate lyase may provide access to bacteria within biofilms by breaking down the biofilm matrix, and polyhydroxybutyrate depolymerase may facilitate utilization of the bacterial storage compound polyhydroxybutyrate as a food source. Enzymes for the synthesis and breakdown of cellulose were identified, and they likely participate in encystation and excystation as in D. discoideum. Trehalose-6-phosphate synthase is present, suggesting that trehalose plays a role in stress adaptation. Detection and response to a number of stress conditions is likely accomplished with a large set of signal transduction histidine kinases and a set of putative receptorserine/threonine kinases similar to those found in E. histolytica. Serine, cysteine and metalloproteases were identified, some of which are likely involved in pathogenicity.

  8. Acanthamoeba encephalitis: A Case Report and Review of Therapy

    PubMed Central

    Zamora, A.; Henderson, H.; Swiatlo, E.

    2014-01-01

    Background: Acanthamoeba is a rare cause of encephalitis yet is associated with high mortality. Treatment protocols vary greatly and generally include combination therapy across a wide spectrum of antiinfective classes. Case Description: A 63-year-old male who underwent renal transplantation presented 6 months after transplantation with depressed level of consciousness. Imaging of the head with computerized tomography showed an enhancing lesion suspicious for brain abscess. Biopsy of the lesion showed Acanthamoeba cysts. The patient was treated with sulfadiazine, fluconazole, flucytosine, azithromycin, and miltefosine but without success. We review recently published cases of Acanthamoeba encephalitis with an emphasis on treatment protocols and outcomes. Conclusion: Free-living protozoans such as Acanthamoeba are ubiquitous in the environment and should be suspected in immunosuppressed persons who present with central nervous system findings and brain abscess. Biopsy is critical to establish the etiology so that appropriate combination therapy can be deployed. PMID:24991471

  9. Crescent Bodies of Parachlamydia acanthamoeba and Its Life Cycle within Acanthamoeba polyphaga: an Electron Micrograph Study

    PubMed Central

    Greub, Gilbert; Raoult, Didier

    2002-01-01

    Parachlamydiaceae are endosymbionts of free-living amoeba first identified in 1997. Two developmental stages, elementary and reticulate bodies, were observed; however, their localization and proportions according to culture condition and duration remain unknown. The life cycle of Parachlamydia acanthamoeba within Acanthamoeba polyphaga was studied by transmission electron microscopy of 8-, 36-, and 144-h coculture. Morphometry and quantification were performed using SAMBA software. The elementary body, the predominant stage within the amoebae, was located mainly within their vacuoles. The multiplication of Parachlamydia bacteria by binary fission of reticulate bodies was independently associated with culture in PYG broth (odds ratio [OR] = 4.4; 95% confidence interval [CI], 1.55 to 12.46) and with the presence of reticulate bodies within the amoebae (OR = 2.10; 95% CI, 1.53 to 2.89). A third developmental stage was observed, the crescent body. Its presence outside and inside the amoebae was associated mainly with prolonged incubation time (OR = 3.98; 95% CI, 1.49 to 10.68, and OR = 5.98; 95% CI, 1.75 to 20.4, respectively). Elementary and crescent bodies were released into the extracellular medium within vesicles or after amoebal lysis. For both, phagocytosis was their mode of entry. This electron micrograph study revealed another infective developmental stage, the crescent body, and provided quantitative analysis of the life cycle of P. acanthamoeba within A. polyphaga. PMID:12039769

  10. Pathogenic Acanthamoeba strains from water sources in Jamaica, West Indies.

    PubMed

    Lorenzo-Morales, J; Lindo, J F; Martinez, E; Calder, D; Figueruelo, E; Valladares, B; Ortega-Rivas, A

    2005-12-01

    In 2004, samples of tap water and of river and sea water associated with human activities were collected in Jamaica, West Indies, and checked for free-living Acanthamoeba. The morphologies of the cysts and trophozoites observed and the results of PCR-based amplifications with a genus-specific primer pair were used to identify the Acanthamoeba isolates. The potential of each isolate as a human pathogen was then evaluated using thermotolerance and osmotolerance assays and two PCR-based assays for Acanthamoeba pathogenesis. Acanthamoeba were identified in 36.1%, 26.4% and 49.6% of the tap-, river- and sea-water samples collected, respectively. Pathogenic potential was shown by 60.0% of the Acanthamoeba strains isolated from tap water, 68.4% of the strains from river water, and 40.4% of the seawater strains. Sequencing of ribosomal DNA revealed the T1, T2, T4, T5, T7, T9 and T11 genotypes. Isolates of the T4 genotype were collected from tap, rain and sea water and, as expected, exhibited the most pathogenic traits; most were osmotolerant, thermotolerant and expressing extracellular serine protease. This is the first study of the occurrence and distribution of Acanthamoeba in water in the West Indies, and the results confirm the presence of potentially pathogenic strains in Jamaica.

  11. Fatal Acanthamoeba Encephalitis in a Patient With a Total Artificial Heart (Syncardia) Device

    PubMed Central

    Tan, Susanna K.; Gajurel, Kiran; Tung, Christie; Albers, Gregory; Deresinski, Stan; Montoya, Jose G.; Sheikh, Ahmad Y.; Banerjee, Dipanjan; Ha, Richard

    2014-01-01

    Acanthamoeba encephalitis is an uncommon but often fatal infection complication. Here we report the first case of Acanthamoeba encephalitis in a patient with a Total Artificial Heart device. PMID:25734127

  12. Acanthamoeba castellanii of the T4 genotype is a potential environmental host for Enterobacter aerogenes and Aeromonas hydrophila

    PubMed Central

    2013-01-01

    Background Acanthamoeba can interact with a wide range of microorganisms such as viruses, algae, yeasts, protists and bacteria including Legionella pneumophila, Pseudomonas aeruginosa, Vibrio cholerae, Helicobacter pylori, Listeria monocytogenes, Mycobacterium spp., and Escherichia coli. In this capacity, Acanthamoeba has been suggested as a vector in the transmission of bacterial pathogens to the susceptible hosts. Methods Here, we used a keratitis isolate of A. castellanii of the T4 genotype and studied its interactions with two bacterial genera which have not been tested before, Enterobacter aerogenes, and Aeromonas hydrophila, as well as E. coli. Assays were performed to determine bacterial association with and invasion of A. castellanii. Additionally, bacterial survival intracellular of A. castellanii trophozoites as well as cysts was determined. Results All three bacterial isolates tested, associated, invaded, and survived inside A. castellanii trophozoites as well as A. castellanii cysts. However, E. aerogenes and E. coli exhibited significantly reduced association with and invasion of A. castellanii as compared with A. hydrophila (P < 0.01 using paired T-test, one tail distribution). In the long term survival assays, all three bacterial isolates tested remained viable inside A. castellanii trophozoites, while amoeba remained intact; however A. hydrophila exhibited higher survival inside amoebae (14.54 ± 3.3 bacteria:amoeba ratio) compared with E. aerogenes (3.96 ± 0.7 bacteria:amoeba ratio) and E. coli (5.85 ± 1.1 bacteria:amoeba ratio). A. hydrophila, E. coli, and E. aerogenes remained viable during the encystment process and exhibited higher levels of recovery from mature cysts (14.13 ± 0.89 A. hydrophila:amoeba ratio, 10.13 ± 1.17 E. aerogenes:amoeba ratio, and 11.95 ± 0.7 E. coli:amoeba ratio). Conclusions A. hydrophila and E. aerogenes also joined the ranks of other bacteria that could benefit from A. castellanii

  13. identification of Pseudomonas spp. as amoeba-resistant microorganisms in isolates of Acanthamoeba.

    PubMed

    José Maschio, Vinicius; Corção, Gertrudes; Rott, Marilise Brittes

    2015-01-01

    Acanthamoeba is a "Trojan horse" of the microbial world. The aim of this study was to identify the presence of Pseudomonas as an amoeba-resistant microorganism in 12 isolates of Acanthamoeba. All isolates showed the genus Pseudomonas spp. as amoeba-resistant microorganisms. Thus, one can see that the Acanthamoeba isolates studied are hosts of Pseudomonas.

  14. IDENTIFICATION OF Pseudomonas spp. AS AMOEBA-RESISTANT MICROORGANISMS IN ISOLATES OF Acanthamoeba

    PubMed Central

    Maschio, Vinicius José; Corção, Gertrudes; Rott, Marilise Brittes

    2015-01-01

    Acanthamoeba is a “Trojan horse” of the microbial world. The aim of this study was to identify the presence of Pseudomonas as an amoeba-resistant microorganism in 12 isolates of Acanthamoeba. All isolates showed the genus Pseudomonas spp. as amoeba-resistant microorganisms. Thus, one can see that the Acanthamoeba isolates studied are hosts of Pseudomonas. PMID:25651331

  15. Actinomyces bowdenii ulcerative keratitis in a dog.

    PubMed

    Sherman, Amanda; Daniels, Joshua B; Wilkie, David A; Lutz, Elizabeth

    2013-09-01

    A 5-year-old spayed female diabetic mixed-breed dog underwent phacoemulsification and intraocular lens implantation to correct bilateral hypermature cataracts. Two months postsurgery, the patient presented with ulcerative keratitis and multifocal stromal abscessation OD, which was controlled, but never resolved, with topical fluoroquinolone therapy. The patient re-presented 2 months later with a new, raised, white gritty corneal opacity associated with hyperemia, chemosis, and blepharospasm OD. Cytology of the right cornea revealed filamentous bacteria, suggestive of Actinomyces spp. Actinomyces bowdenii was subsequently isolated in pure culture and identified via 16s rDNA sequencing. Actinomyces bowdenii has never before been described as a cause of ocular infection. An immunosuppressed corneal environment likely contributed to this opportunistic Actinomycosis. The infection was not controlled with fluoroquinolone therapy, and the isolate, in vitro, was resistant to three fluoroquinolones (ciprofloxacin, ofloxacin, and levofloxacin), which also has not been previously reported for this species of Actinomyces. A superficial keratectomy with conjunctival graft was employed to successfully manage the infection.

  16. Topical Corticosteroids in the Management of Bacterial Keratitis.

    PubMed

    Tuli, Sonal S

    2013-12-01

    Bacterial keratitis can cause significant morbidity from ulceration of the cornea and the resultant scarring. The use of steroids to decrease these complications is controversial with arguments for and against their use. The SCUT (Steroids for Corneal Ulcers Trial) was initiated in 2006 to definitively determine whether steroids in bacterial keratitis were beneficial or harmful. While the SCUT showed no benefit or harm overall, subgroup analyses showed that larger, more central ulcers with very poor initial visual acuity may benefit. On the other hand, Nocardia ulcers that were treated with steroids had worse outcomes. The study did have some limitations as the patient population was not typical for bacterial keratitis in the United States, and there were some criticisms of the therapeutic approach so the question is still not definitively answered.

  17. Mycotic keratitis: profile of Fusarium species and their mycotoxins.

    PubMed

    Naiker, S; Odhav, B

    2004-02-01

    In this study, Fusarium species isolated from 29 patients with mycotic keratitis were identified and tested for their ability to produce mycotoxins. Members of the F. solani species complex (Fs complex) were the predominant species isolated, followed by F. verticillioides, F. dimerum, members of the F. oxysporum species complex Fo complex), F. incarnatum, F. chlamydosporum and F. lateritium. Of these, 76% of the Fusarium isolates produced fusaric acid, moniliformin or fumonisin B1. Many of the fusaria isolated are common aetiological agents of mycotic keratitis infections. However, F. incarnatum, F. chlamydosporum and F. lateritium have previously not been found in this infection. These findings indicate that a greater variety of fusarial species are becoming associated with mycotic keratitis infections. This paper further demonstrates the mycotoxin-producing ability of these clinical isolates and assesses cellular cytotoxicity.

  18. Corneal cross-linking in 9 horses with ulcerative keratitis

    PubMed Central

    2013-01-01

    Background Corneal ulcers are one of the most common eye problems in the horse and can cause varying degrees of visual impairment. Secondary infection and protease activity causing melting of the corneal stroma are always concerns in patients with corneal ulcers. Corneal collagen cross-linking (CXL), induced by illumination of the corneal stroma with ultraviolet light (UVA) after instillation of riboflavin (vitamin B2) eye drops, introduces crosslinks which stabilize melting corneas, and has been used to successfully treat infectious ulcerative keratitis in human patients. Therefore we decided to study if CXL can be performed in sedated, standing horses with ulcerative keratitis with or without stromal melting. Results Nine horses, aged 1 month to 16 years (median 5 years) were treated with a combination of CXL and medical therapy. Two horses were diagnosed with mycotic, 5 with bacterial and 2 with aseptic ulcerative keratitis. A modified Dresden-protocol for CXL could readily be performed in all 9 horses after sedation. Stromal melting, diagnosed in 4 horses, stopped within 24 h. Eight of nine eyes became fluorescein negative in 13.5 days (median time; range 4–26 days) days after CXL. One horse developed a bacterial conjunctivitis the day after CXL, which was successfully treated with topical antibiotics. One horse with fungal ulcerative keratitis and severe uveitis was enucleated 4 days after treatment due to panophthalmitis. Conclusions CXL can be performed in standing, sedated horses. We did not observe any deleterious effects attributed to riboflavin or UVA irradiation per se during the follow-up, neither in horses with infectious nor aseptic ulcerative keratitis. These data support that CXL can be performed in the standing horse, but further studies are required to compare CXL to conventional medical treatment in equine keratitis and to optimize the CXL protocol in this species. PMID:23803176

  19. Protein profiles and immunoreactivities of Acanthamoeba morphological groups and genotypes.

    PubMed

    Pumidonming, Wilawan; Koehsler, Martina; Leitsch, David; Walochnik, Julia

    2014-11-01

    Acanthamoeba is a free-living protozoan found in a wide variety of habitats. A classification of Acanthamoeba into currently eighteen genotypes (T1-T18) has been established, however, data on differences between genotypes on the protein level are scarce. The aim of this study was to compare protein and immunoreactivity profiles of Acanthamoeba genotypes. Thirteen strains, both clinical and non-clinical, from genotypes T4, T5, T6, T7, T9, T11 and T12, representing three morphological groups, were investigated for their protein profiles and IgG, IgM and IgA immunoreactivities. It was shown that protein and immunoreactivity profiles of Acanthamoeba genotypes T4, T5, T6, T7, T9, T11 and T12 are clearly distinct from each other, but the banding patterns correlate to the morphological groups. Normal human sera revealed anti-Acanthamoeba antibodies against isolates of all investigated genotypes, interestingly, however only very weak IgM and virtually no IgA immunoreactivity with T7 and T9, both representing morphological group I. The strongest IgG, IgM and IgA immunoreactivities were observed for genotypes T4, T5 and T6. Differences of both, protein and immunological patterns, between cytopathic and non-cytopathic strains, particularly within genotype T4, were not at the level of banding patterns, but rather in expression levels.

  20. Genetic diversity of Acanthamoeba isolates from ocean sediments

    PubMed Central

    Liu, Hua; Ha, Young-Ran; Lee, Sung-Tae; Hong, Yean-Chul; Kong, Hyun-Hee

    2006-01-01

    Genetic diversity of 18 Acanthamoeba isolates from ocean sediments was evaluated by comparing mitochondrial (mt) DNA RFLP, 18S rDNA sequences and by examining their cytopathic effects on human corneal epithelial cells versus reference strains. All isolates belonged to morphologic group II. Total of 16 restriction phenotypes of mtDNA from 18 isolates demonstrated the genetic diversity of Acanthamoeba in ocean sediments. Phylogenetic analysis using 18s rDNA sequences revealed that the 18 isolates were distinct from morphological groups I and III. Fifteen isolates showed close relatedness with 17 clinical isolates and A. castellanii Castellani and formed a lineage equivalent to T4 genotype of Byers' group. Two reference strains from ocean sediment, A. hatchetti BH-2 and A. griffini S-7 clustered unequivocally with these 15 isolates. Diversity among isolates was also evident from their cytopathic effects on human corneal cells. This is the first time describing Acanthamoeba diversity in ocean sediments in Korea. PMID:16809959

  1. Interaction between Vibrio mimicus and Acanthamoeba castellanii

    PubMed Central

    Abd, Hadi; Valeru, Soni Priya; Sami, Susan Marouf; Saeed, Amir; Raychaudhuri, Saumya; Sandström, Gunnar

    2010-01-01

    Vibrio mimicus is a Gram-negative bacterium, which causes gastroenteritis and is closely related to Vibrio cholerae. The environmental reservoir of this bacterium is far from defined. Acanthamoeba as well as Vibrio species are found in diverse aquatic environments. The present study was aimed to investigate the ability of A. castellanii to host V. mimicus, the role of bacterial protease on interaction with A. castellanii and to disclose the ability of cysts to protect intracellular V. mimicus. Co-cultivation, viable counts, gentamicin assay, electron microscopy and statistical analysis showed that co-cultivation of wild type and luxO mutant of V. mimicus strains with A. castellanii did not inhibit growth of the amoeba. On the other hand co-cultivation enhanced growth and survival of V. mimicus strains. Vibrio mimicus showed intracellular behaviour because bacteria were found to be localized in the cytoplasm of amoeba trophozoites and remain viable for 14 days. The cysts protected intracellular V. mimicus from high level of gentamicin. The intracellular growth of V. mimicus in A. castellanii suggests a role of A. castellanii as a host for V. mimicus. PMID:20454692

  2. Acanthamoeba royreba sp. n. from a human tumor cell culture.

    PubMed

    Willaert, E; Stevens, A R; Tyndall, R L

    1978-02-01

    A new species of Acanthamoeba was isolated from a culture of an established line of human choriocarcinoma cells. The identification of this strain, originally called the Oak Ridge strain, and the establishment of a new species for it were based on morphologic, serologic, and immunochemical studies. In general, the structure of the trophozoite did not differ significantly from that of other species of Acanthamoeba, except that a body which more closely resembled a centriole than material described previously as centriolar satellites was observed in trophozoites examined with the electron microscope. The dimensions of the trophozoite were the smallest among the species of Acanthamoeba. The cyst was typical of the genus, but differed from those of other species by its smaller size and the presence of numerous ostioles. Studies of the Oak Ridge strain by immunofluorescence using antisera developed against the isolate and Acanthamoeba culbertsoni, A. castellanii, A. polyphaga, A. rhysodes, A. astronyxis, and A. palestinensis revealed the antigenic uniqueness of the Oak Ridge strain. It was demonstrated by immunoelectrophoretic analyses of the soluble proteins of the Oak Ridge strain that shared approximately 1/2 of its antigenic structure with A. castellanii and A. culbertsoni. The antigenic differences of the isolate from other species of Acanthamoeba were deduced from comparison of the antigenic constitution of these species and the Oak Ridge strain with A. culbertsoni and A. castellanii. Although the strain was initially recognized by its cytopathogenicity for cultures, it did not produce acute infections in mice after intranasal inoculation of 1 X 10(4) ameba/mouse. The foregoing results constituted the basis for the establishment of the Oak Ridge strain as a new species, A. royreba sp. n., in the genus Acanthamoeba. PMID:566323

  3. Rhizopus Keratitis Associated with Poor Contact Lens Hygiene

    PubMed Central

    Warner, David B.; WrightIII, Hugh E.; Rosenbaum, Eric R.

    2016-01-01

    We report a case of Rhizopus keratitis in a young woman with poor contact lens hygiene. The mold was highly sensitive to treatment with amphotericin 0.15% drops, after a relatively prompt diagnosis. Obtaining cultures of both corneal infiltrates and presumably infected contact lenses may help to avoid a delay in proper treatment. PMID:26889155

  4. Spectrum and Sensitivity of Bacterial Keratitis Isolates in Auckland

    PubMed Central

    Swift, S.; Dean, S. J.; Ormonde, S. E.

    2016-01-01

    Background. The bacteria isolated from severe cases of keratitis and their antibiotic sensitivity are recognised to vary geographically and over time. Objectives. To identify the most commonly isolated bacteria in keratitis cases admitted over a 24-month period to a public hospital in Auckland, New Zealand, and to investigate in vitro sensitivity to antibiotics. Methods. Hospital admissions for culture-proven bacterial keratitis between January 2013 and December 2014 were identified. Laboratory records of 89 culture positive cases were retrospectively reviewed and antibiotic sensitivity patterns compared with previous studies from other NZ centres. Results. From 126 positive cultures, 35 species were identified. Staphylococcus was identified to be the most common isolate (38.2%), followed by Pseudomonas (21.3%). Over the last decade, infection due to Pseudomonas species, in the same setting, has increased (p ≤ 0.05). Aminoglycosides, cefazolin, ceftazidime, erythromycin, tetracycline, and doxycycline were 100% effective against tested isolates in vitro. Amoxicillin (41.6%), cefuroxime (33.3%), and chloramphenicol (94.7%) showed reduced efficacy against Gram-negative bacteria, whereas penicillin (51%) and ciprofloxacin (98.8%) showed reduced efficacy against Gram-positive bacteria. Conclusions. Despite a shift in the spectrum of bacterial keratitis isolates, antibiotic sensitivity patterns have generally remained stable and show comparability to results within the last decade from NZ centres. PMID:27213052

  5. Colonization of broilers by Campylobacter jejuni internalized within Acanthamoeba castellanii

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We present the first report that Campylobacter jejuni, internalized within Acanthamoeba castellanii, colonized broilers. After 1, 3, 7 and 14 days post challenge none of the broilers challenged with negative controls were colonized, but were with internalized C. jejuni. The biology of protozoa-Cam...

  6. Fatal Disseminated Acanthamoeba lenticulata Acanthamebiasis in a Heart Transplant Patient

    PubMed Central

    Combes, Alain; de Jonckheere, Johan F.; Datry, Annick; Varnous, Shaïda; Martinez, Valérie; Ptacek, Sara García; Caumes, Eric; Capron, Frédérique; Francès, Camille; Gibert, Claude; Chosidow, Olivier

    2007-01-01

    We report a fatal case of disseminated acanthamebiasis caused by Acanthamoeba lenticulata (genotype T5) in a 39-year-old heart transplant recipient. The diagnosis was based on skin histopathologic results and confirmed by isolation of the ameba from involved skin and molecular analysis of a partial 18S rRNA gene sequence (DF3). PMID:17553253

  7. Isolation of Acanthamoeba culbertsoni from a patient with meningitis.

    PubMed

    Lalitha, M K; Anandi, V; Srivastava, A; Thomas, K; Cherian, A M; Chandi, S M

    1985-04-01

    A case of amoebic meningitis, presumably primary, was encountered in the Christian Medical College Hospital, Vellore, South India, in November 1983. The patient, a 40-year-old man, had cerebrospinal fluid rhinorrhea before the meningitis developed. Acanthamoeba culbertsoni was repeatedly demonstrated in and cultured from the cerebrospinal fluid. The patient responded dramatically to a combination therapy of penicillin and chloramphenicol. PMID:3988911

  8. Acanthamoeba polyphaga mimivirus Virophage Seroconversion in Travelers Returning from Laos

    PubMed Central

    Parola, Philippe; Renvoisé, Aurélie; Botelho-Nevers, Elisabeth; La Scola, Bernard; Desnues, Christelle

    2012-01-01

    During January 2010, a husband and wife returned from Laos to France with probable parasitic disease. Increased antibodies against an Acanthamoeba polyphaga mimivirus virophage indicated seroconversion. While in Laos, they had eaten raw fish, a potential source of the virophage. This virophage, associated with giant viruses suspected to cause pneumonia, could be an emerging pathogen. PMID:22932431

  9. Degeneration and Regeneration of Subbasal Corneal Nerves after Infectious Keratitis: A Longitudinal In Vivo Confocal Microscopy Study

    PubMed Central

    Müller, Rodrigo; Abedi, Farshad; Cruzat, Andrea; Witkin, Deborah; Baniasadi, Neda; Cavalcanti, Bernardo M.; Jamali, Arsia; Chodosh, James; Dana, Reza; Pavan-Langston, Deborah; Hamrah, Pedram

    2015-01-01

    Purpose To investigate the longitudinal alterations of subbasal corneal nerves in patients with infectious keratitis (IK) during acute phase, cessation of treatment and recovery phase by in vivo confocal microscopy (IVCM). Design Prospective, longitudinal, case-control, single-center study. Subjects Fifty-six eyes of 56 patients with the diagnosis of bacterial (n=28), fungal (n=15), and Acanthamoeba (n=13) keratitis were included in the study. Thirty eyes of 30 normal volunteers constituted the control group. Methods Corneal sensation and serial IVCM of the central cornea were performed prospectively, by using the Heidelberg Retina Tomograph 3/Rostock Cornea Module (Heidelberg Engineering, Germany). IVCM images were assessed at 3 time points: at the first visit of the patient to the cornea service, at cessation of antimicrobial treatment, and up to six months after the resolution of infection. Main outcome measures Total nerve number and length, main nerve trunks, branching and corneal sensation were assessed during the follow-up period. Results Corneal nerves were significantly reduced during the acute phase in eyes with IK compared with controls across all subgroups, with total nerve length of 5.47 ± 0.69 vs. 20.59 ± 1.06 mm/mm2; p<0.0001. At the cessation of treatment, corneal nerves in patients with IK had regenerated, including total nerve length (8.49 ± 0.94; p=0.02) and nerve branch length (4.80 ± 0.37; p=0.005). During the recovery phase, after resolution of infection, corneal nerves further regenerated, including total nerve length (12.13 ± 1.97; p=0.005), main nerve trunk length (5.80 ± 1.00; p=0.01) and nerve branch length (6.33 ± 0.76; p=0.003) as compared to the acute phase, but were still significantly lower when compared to controls (p<0.05 for all parameters). Corneal degeneration and regeneration correlated with corneal sensation (r=0.47, p=0.0009). Conclusion Patients with IK, suffering from profound loss of corneal nerves during the acute

  10. Lethal Keratitis, Ichthyosis, and Deafness Syndrome Due to the A88V Connexin 26 Mutation.

    PubMed

    Esmer, Carmen; Salas-Alanis, Julio C; Fajardo-Ramirez, Oscar R; Ramírez, Brenda; Hua, Rong; Choate, Keith

    2016-01-01

    Keratitis-ichthyosis-deafness syndrome is a well-characterized disease that has been related to mutations in the GJB6 gene. Clinical features such as erythrokeratoderma, palmoplantar keratoderma, alopecia, and progressive vascularizing keratitis, among others, are well known in this entity. In this report we describe a newborn female patient diagnosed with keratitis-ichthyosis-deafness syndrome with a lethal outcome due to sepsis. The patient harbored the mutation A88V that has been previously reported in lethal cases.

  11. Lethal Keratitis, Ichthyosis, and Deafness Syndrome Due to the A88V Connexin 26 Mutation.

    PubMed

    Esmer, Carmen; Salas-Alanis, Julio C; Fajardo-Ramirez, Oscar R; Ramírez, Brenda; Hua, Rong; Choate, Keith

    2016-01-01

    Keratitis-ichthyosis-deafness syndrome is a well-characterized disease that has been related to mutations in the GJB6 gene. Clinical features such as erythrokeratoderma, palmoplantar keratoderma, alopecia, and progressive vascularizing keratitis, among others, are well known in this entity. In this report we describe a newborn female patient diagnosed with keratitis-ichthyosis-deafness syndrome with a lethal outcome due to sepsis. The patient harbored the mutation A88V that has been previously reported in lethal cases. PMID:27409001

  12. The treatment of herpes simplex virus epithelial keratitis.

    PubMed Central

    Wilhelmus, K R

    2000-01-01

    PURPOSE: Epithelial keratitis is the most common presentation of ocular infection by herpes simplex virus (HSV). Quantitative assessment of available therapy is needed to guide evidence-based ophthalmology. This study aimed to compare the efficacy of various treatments for dendritic or geographic HSV epithelial keratitis and to evaluate the role of various clinical characteristics on epithelial healing. METHODS: Following a systematic review of the literature, information from clinical trials of HSV dendritic or geographic epithelial keratitis was extracted, and the methodological quality of each study was scored. Methods of epithelial cauterization and curettage were grouped as relatively equivalent physicochemical therapy, and solution and ointment formulations of a given topical antiviral agent were combined. The proportion healed with 1 week of therapy, a scheduled follow-up day that approximated the average time of resolution with antiviral therapy, was selected as the primary outcome based on a masked evaluation of maximum treatment differences in published healing curves. The proportion healed at 14 days was recorded as supplemental information. Fixed-effects and random-effects meta-analysis models were used to obtain summary estimates by pooling results from comparative treatment trials. Hypotheses about which prognostic factors might affect epithelial healing during antiviral therapy were developed by multivariate analysis of the Herpetic Eye Disease Study dataset. RESULTS: After excluding 48 duplicate reports, 14 nonrandomized studies, 15 studies with outdated or similar treatments, and 29 trials lacking sufficient data on healing or accessibility, 76 primary reports were identified. These reports involved 4,251 patients allocated to 93 treatment comparisons of dendritic epithelial keratitis in 28 categories and 9 comparisons of geographic epithelial keratitis in 6 categories. For dendritic keratitis, idoxuridine was better than placebo at 7 days

  13. Climate change and predicted trend of fungal keratitis in Egypt.

    PubMed

    Saad-Hussein, A; El-Mofty, H M; Hassanien, M A

    2011-06-01

    Rising rates of invasive fungal infections may be linked to global climate change. A study was made of the trend of ophthalmic fungal corneal keratitis in the greater Cairo area of Egypt and its association with climate records during the same period. Data on diagnosed cases of fungal keratitis were collected from records of ophthalmic departments of Cairo University hospital and atmospheric temperature and humidity for the greater Cairo area were obtained from online records. Statistical analysis showed a significant increase in the relative frequency of keratomycosis during 1997-2007. The rise correlated significantly with rises n min,mum temperature and the maximum atmospheric humidity in the greater Cairo area over the same period (after exclusion of the effect of the maximum atmos pheric temperature). The predicted increase in keratomycosis up to the year 2030 corresponds to predicted increases in CO2 emissions and surface temperature from climate change models for Egypt.

  14. [Keratitis by Lasiodiplodia theobromae: a case report and literature review].

    PubMed

    Samudio, Margarita; Laspina, Florentina; Fariña, Norma; Franco, Alicia; Mino de Kaspar, Herminia; Giusiano, Gustavo

    2014-12-01

    We report a case of mycotic keratitis caused by Lasiodiplodia theobromae in a 60-year-old man with a history of ocular trauma with vegetable matter. Ophthalmological assessment with slit-lamp and microbiological evaluation of the corneal ulcer by conventional microbiological techniques were performed. Mycology study of the corneal scraping showed the presence of fungal filaments and the isolate was identified as Lasiodiplodia theobromae. Patient was treated with natamycin 5% and fluconazole 0.2% for 37 days. The infection was controlled but the corneal scars required a cornea transplant. This is the first case of keratitis by Lasiodiplodia theobromae in Paraguay. Difficulties in the management of these cases, which often requires surgical procedures, are discussed.

  15. Immunological Aspects of Acute and Recurrent Herpes Simplex Keratitis

    PubMed Central

    Hus, Iwona

    2014-01-01

    Herpes simplex keratitis (HSK) belongs to the major causes of visual morbidity worldwide and available methods of treatment remain unsatisfactory. Primary infection occurs usually early in life and is often asymptomatic. Chronic visual impairment and visual loss are caused by corneal scaring, thinning, and vascularization connected with recurrent HSV infections. The pathogenesis of herpetic keratitis is complex and is still not fully understood. According to the current knowledge, corneal scarring and vascularization are the result of chronic inflammatory reaction against HSV antigens. In this review we discuss the role of innate and adaptive immunities in acute and recurrent HSV ocular infection and present the potential future targets for novel therapeutical options based on immune interventions. PMID:25276842

  16. [Keratitis by Lasiodiplodia theobromae: a case report and literature review].

    PubMed

    Samudio, Margarita; Laspina, Florentina; Fariña, Norma; Franco, Alicia; Mino de Kaspar, Herminia; Giusiano, Gustavo

    2014-12-01

    We report a case of mycotic keratitis caused by Lasiodiplodia theobromae in a 60-year-old man with a history of ocular trauma with vegetable matter. Ophthalmological assessment with slit-lamp and microbiological evaluation of the corneal ulcer by conventional microbiological techniques were performed. Mycology study of the corneal scraping showed the presence of fungal filaments and the isolate was identified as Lasiodiplodia theobromae. Patient was treated with natamycin 5% and fluconazole 0.2% for 37 days. The infection was controlled but the corneal scars required a cornea transplant. This is the first case of keratitis by Lasiodiplodia theobromae in Paraguay. Difficulties in the management of these cases, which often requires surgical procedures, are discussed. PMID:25679935

  17. Magnetotherapy in the treatment of viral conjunctivitis and keratitis.

    PubMed

    Pasek, Jarosław; Pasek, Tomasz; Herba, Ewa; Misiak, Anna; Sieroń-Stołtny, Karolina; Sieroń, Aleksander

    2008-01-01

    Ocular infections are one of the most frequent causes of ailments among the patients coming to ophthalmologic offices. This article presents one of the physical medicine's methods--magnetotherapy--which uses the alternating low frequency magnetic fields in the therapy of viral conjunctivitis and keratitis in a 49-year-old female patient. Basing on the obtained results it was stated that this method broadens the treatment possibilities becoming a precious supplement and support treatment method in ophthalmology. PMID:19323071

  18. Seasonal distribution of potentially pathogenic Acanthamoeba species from drinking water reservoirs in Taiwan.

    PubMed

    Kao, Po-Min; Hsu, Bing-Mu; Hsu, Tsui-Kang; Liu, Jorn-Hon; Chang, Hsiang-Yu; Ji, Wen-Tsai; Tzeng, Kai-Jiun; Huang, Shih-Wei; Huang, Yu-Li

    2015-03-01

    In order to detect the presence/absence of Acanthamoeba along with geographical variations, water quality variations and seasonal change of Acanthamoeba in Taiwan was investigated by 18S ribosomal RNA (rRNA) gene TaqMan quantitative real-time PCR. Samples were collected quarterly at 19 drinking water reservoir sites from November 2012 to August 2013. Acanthamoeba was detected in 39.5 % (30/76) of the water sample, and the detection rate was 63.2 % (12/19) from samples collected in autumn. The average concentration of Acanthamoeba was 3.59 × 10(4) copies/L. For geographic distribution, the detection rate for Acanthamoeba at the northern region was higher than the central and southern regions in all seasons. Results of Spearman rank test revealed that heterotrophic plate count (HPC) had a negative correlation (R = -0.502), while dissolved oxygen (DO) had a positive correlation (R = 0.463) in summer. Significant differences were found only between the presence/absence of Acanthamoeba and HPC in summer (Mann-Whitney U test, P < 0.05). T2 and T4 genotypes of Acanthamoeba were identified, and T4 was the most commonly identified Acanthamoeba genotypes. The presence of Acanthamoeba in reservoirs presented a potential public health threat and should be further examined.

  19. Peripheral Ulcerative Keratitis following Laser in situ Keratomileusis

    PubMed Central

    Burkholder, Bryn M.; Kuo, Irene C.

    2016-01-01

    Purpose We report a case of a patient with a history of glomerulonephropathy, not disclosed prior to laser in situ keratomileusis (LASIK), who developed severe postoperative peripheral ulcerative keratitis (PUK) soon after surgery. Method Case report. Results Within a week of surgery, the patient, who had no blepharitis or ocular surface disease, also developed diffuse lamellar keratitis (DLK) that was not contiguous with the PUK. Microbiologic evaluation of the flap interface disclosed no organisms, and no epithelial ingrowth was found. Both PUK and DLK resolved with topical and oral steroid therapy, and the patient's induced refractive error improved over the 12 months following LASIK. Conclusions Necrotizing keratitis has been described after LASIK surgery in patients with or without autoimmune disease. However, to our knowledge, there has been no case of PUK following LASIK. As shown by our patient's clinical course and the typical association of PUK with systemic conditions, patients with a history of atypical postinfectious sequelae may require additional preoperative counseling, vigilant postoperative monitoring, and possibly additional intervention. Because patients do not always divulge medical details, especially if an extraocular site was involved or illness occurred many years prior, this case demonstrates the importance of performing a diligent history that excludes autoimmune disorders or atypical postinfectious sequelae prior to proceeding with keratorefractive intervention. PMID:26889153

  20. Antifungal Effect of Essential Oils against Fusarium Keratitis Isolates.

    PubMed

    Homa, Mónika; Fekete, Ildikó Pálma; Böszörményi, Andrea; Singh, Yendrembam Randhir Babu; Selvam, Kanesan Panneer; Shobana, Coimbatore Subramanian; Manikandan, Palanisamy; Kredics, László; Vágvölgyi, Csaba; Galgóczy, László

    2015-09-01

    The present study was carried out to investigate the antifungal effects of Cinnamomum zeylanicum, Citrus limon, Juniperus communis, Eucalyptus citriodora, Gaultheria procumbens, Melaleuca alternifolia, Origanum majorana, Salvia sclarea, and Thymus vulgaris essential oils against Fusarium species, the most common etiologic agents of filamentous fungal keratitis in South India. C. zeylanicum essential oil showed strong anti-Fusarium activity, whereas all the other tested essential oils proved to be less effective. The main component of C. zeylanicum essential oil, trans-cinnamaldehyde, was also tested and showed a similar effect as the oil. The in vitro interaction between trans-cinnamaldehyde and natamycin, the first-line therapeutic agent of Fusarium keratitis, was also investigated; an enhanced fungal growth inhibition was observed when these agents were applied in combination. Light and fluorescent microscopic observations revealed that C. zeylanicum essential oil/trans-cinnamaldehyde reduces the cellular metabolism and inhibits the conidia germination. Furthermore, necrotic events were significantly more frequent in the presence of these two compounds. According to our results, C. zeylanicum essential oil/trans-cinnamaldehyde provides a promising basis to develop a novel strategy for the treatment of Fusarium keratitis. PMID:26227503

  1. Antifungal Effect of Essential Oils against Fusarium Keratitis Isolates.

    PubMed

    Homa, Mónika; Fekete, Ildikó Pálma; Böszörményi, Andrea; Singh, Yendrembam Randhir Babu; Selvam, Kanesan Panneer; Shobana, Coimbatore Subramanian; Manikandan, Palanisamy; Kredics, László; Vágvölgyi, Csaba; Galgóczy, László

    2015-09-01

    The present study was carried out to investigate the antifungal effects of Cinnamomum zeylanicum, Citrus limon, Juniperus communis, Eucalyptus citriodora, Gaultheria procumbens, Melaleuca alternifolia, Origanum majorana, Salvia sclarea, and Thymus vulgaris essential oils against Fusarium species, the most common etiologic agents of filamentous fungal keratitis in South India. C. zeylanicum essential oil showed strong anti-Fusarium activity, whereas all the other tested essential oils proved to be less effective. The main component of C. zeylanicum essential oil, trans-cinnamaldehyde, was also tested and showed a similar effect as the oil. The in vitro interaction between trans-cinnamaldehyde and natamycin, the first-line therapeutic agent of Fusarium keratitis, was also investigated; an enhanced fungal growth inhibition was observed when these agents were applied in combination. Light and fluorescent microscopic observations revealed that C. zeylanicum essential oil/trans-cinnamaldehyde reduces the cellular metabolism and inhibits the conidia germination. Furthermore, necrotic events were significantly more frequent in the presence of these two compounds. According to our results, C. zeylanicum essential oil/trans-cinnamaldehyde provides a promising basis to develop a novel strategy for the treatment of Fusarium keratitis.

  2. Clinical and microbiological profile of infectious keratitis in children

    PubMed Central

    2013-01-01

    Background Infectious keratitis is a sight-threatening condition for children. The purpose of this study was to describe the clinical profile, risk factors and microbiological profile of infectious keratitis in children. Methods Retrospective review of clinical records of patients under 16 years of age with history of microbial keratitis seen at a tertiary referral center. Clinical characteristics, risk factors, visual and surgical outcomes as well as the microbiological profile are analyzed. Results Forty-one eyes of 41 patients. Mean age was 8.7 years. Time between the onset of symptoms and ophthalmological examination was 12.7 days. Predisposing factors were found in 78%; ocular trauma was the most common (25%). Visual acuity equal or worse than 20/200 at admission correlated positively with a poorer visual outcome, p=0.002. Positivity of cultures was 34%. Gram-positive bacteria were isolated in 78.5%; Staphylococcus epidermidis (28.6%) was the most common microorganism. Conclusions Our study emphasizes the importance of a prompt diagnosis and treatment of infectious corneal ulcers in children. Trauma and contact lenses were the main predisposing factors. Gram-positive organisms were isolated in the vast majority of cases and visual outcomes are usually poor. PMID:24131681

  3. Microbial Keratitis Profile at a University Hospital in Hong Kong

    PubMed Central

    Lai, Tracy H. T.; Young, Alvin L.

    2014-01-01

    Purpose. To evaluate the recent trends in demographics, risk factors, and microbiological profiles of microbial keratitis at a university hospital in Hong Kong. Design. Retrospective review. Methods. The medical records of 51 patients admitted to the Prince of Wales Hospital for microbial keratitis from January 2010 to June 2012 were reviewed. Demographics, risk factors, clinical features, microbiological results, and treatment were recorded. Data was analyzed and compared to our historical sampled data collected 11 years ago. Results. The mean age of patients was 41.6 ± 20.3 years. Contact lens use was the major risk factor (45%), followed by injury (12%). The culture positive rate was 59%, of which 37% were Gram-positive organisms and 53% were Gram-negative organisms. Pseudomonas aeruginosa (50%) and coagulase-negative Staphylococcus (13%) were the most commonly isolated pathogens. No resistance to fluoroquinolones was identified. Conclusions. Our study showed that contact lens wear remained the major risk factor for microbial keratitis in Hong Kong and Pseudomonas aeruginosa remained the commonest bacterium isolated. This is comparable to our historical data and other studies conducted in East Asia. PMID:27433502

  4. Lasiodiplodia theobromae keratitis: a case report and review of literature.

    PubMed

    Saha, Suman; Sengupta, Jayangshu; Banerjee, Debdulal; Khetan, Archana

    2012-10-01

    A case report and review of literature is reported of a rare case of fungal keratitis from eastern India. A 32-year-old woman with a history of vegetative trauma presented with keratitis in left eye. Microbiological examination of corneal scraping showed refractile hyphae with aseptate branching filaments and black pigmented colonies on multiple solid agar medium. Organism was identified from culture using D1/D2 region of LSU (Large Sub Unit: 28S rDNA)-based molecular technique. PCR amplified a band with a sequence that was 100 % homologous with Lasiodiplodia theobromae. The organism was susceptible to amphotericin B and voriconazole and demonstrated resistance to itraconazole and fluconazole. A therapeutic keratoplasty was performed following non-responsiveness to initial topical voriconazole (2 %) therapy. Recurrence in graft was controlled with topical voriconazole and intracameral amphotericin B. However, the graft failed at the end of 3 months. L. theobromae is a rare cause of fungal keratitis. Management of these cases is difficult, often involving surgical procedures. PMID:22544631

  5. Detection of glycoproteins in the Acanthamoeba plasma membrane

    SciTech Connect

    Paatero, G.I.L. ); Gahmberg, C.G. )

    1988-11-01

    In the present study the authors have shown that glycoproteins are present in the plasma membrane of Acanthamoeba castellanii by utilizing different radioactive labeling techniques. Plasma membrane proteins in the amoeba were iodinated by {sup 125}I-lactoperoxidase labeling and the solubilized radiolabeled glycoproteins were separated by lectin-Sepharose affinity chromatography followed by polyacrylamide gel electrophoresis. The periodate/NaB{sup 3}H{sub 4} and galactose oxidase/NaB{sup 3}H{sub 4} labeling techniques were used for labeling of surface carbohydrates in the amoeba. Several surface-labeled glycoproteins were observed in addition to a diffusely labeled region with M{sub r} of 55,000-75,000 seen on electrophoresis, which could represent glycolipids. The presence of glycoproteins in the plasma membrane of Acanthamoeba castellanii was confirmed by metabolic labeling with ({sup 35}S)methionine followed by lectin-Sepharose affinity chromatography and polyacrylamide gel electrophoresis.

  6. [A case of meningoencephalitis caused by Acanthamoeba sp. in Dakar].

    PubMed

    Ndiaye, M; Diop, A G; Dieng, Y; Seydi, M; Diouf, F S; Diop, B M; Ndiaye, I P

    2005-01-01

    Primary meningoencephalitis caused by free-living soil ameba is rare. We report the first diagnosed case of meningoencephalitis due to Acanthamoeba sp. in Senegal. The patient was a 24-year-old Senegalese woman hospitalized in the neurology department of Fann Hospital. Diagnosis was made 6 months after the onset of symptoms based mainly on headache with fever usually occurring in the evening, chills, and lumbar puncture demonstrating turbid fluid. Parasitological examination of cool cerebrospinal fluid sediment revealed the presence of free-living ameba trophozoites of the Acanthamoeba genus. Species determination by culture on 1.5% agar-agar enriched with Escherichia coli failed. The patient died one month following initiation of treatment using amphotericine B. PMID:15903081

  7. In vitro fusion of Acanthamoeba phagolysosomes. I. Demonstration and quantitation of vacuole fusion in Acanthamoeba homogenates.

    PubMed

    Oates, P J; Touster, O

    1976-02-01

    Fusion of phagolysosomes (PLs) has been demonstrated to occur in vitro. Two separate cell homogenates of the ameba Acanthamoeba sp. (Neff) were prepared, each rich in PLs labeled with distinctive particulate markers. Portions of each were incubated together in vitro and fusion occurred as evidenced by the appearance of PLs containing both types of markers. Fusion was confirmed by electron microscopy, including serial sectioning. The membranes of fused vacuoles excluded the dye eosin Y. Surviving cells in the homogenates were not responsible for the observed fusion. Fusion was obtained using either synthetic markers (polystyrene and polyvinyltoluene latex) or biological markers (autoclaved yeast cells and glutaraldehyde-fixed goat red blood cells), or a combination of both. The specificity of PL fusion in vivo appeared to be maintained in vitro. As determined by light and electron microscopy, the fusion reaction was dependent on time and temperature, and on the initial presence of membrane around both marker particles. A minimum of 10% of the vacuoles fused by 10 min of incubation at 30 degrees C, and no rupture of the vacuoles was detected during this time. After 10 min of incubation, vacuole rupture began and fusion ceased. At a constant initial vacuole concentration, the extent of PL fusion in vitro was quantitatively reproducible. This appears to be a promising system for further investigation of membrane fusion in the lysosomal system. PMID:1245550

  8. Field's stain--a rapid staining method for Acanthamoeba spp.

    PubMed

    Pirehma, M; Suresh, K; Sivanandam, S; Anuar, A K; Ramakrishnan, K; Kumar, G S

    1999-10-01

    Acanthamoeba sp. is a free-living amoeba known to cause chronic central nervous system infection or eye infection in humans. Many cases remain undetected for want of a good detection system. We report for the first time a rapid staining method to facilitate the identification of Acanthamoeba sp. using the modified Field's staining technique. A. castellanii, which was used in the present experiment, is maintained in our laboratory in mycological peptone medium (Gibco). The cultures were pooled together and smears were made on glass slides for staining purposes. Different types of stains such as Field's stain, modified Field's stain, Wright's stain, Giemsa stain, Ziehl-Neelsen stain, and trichrome stain were used to determine the best stain for the identification of this amoeba. The concentration of various stains and the duration of staining were varied to provide the best color and contrast for each stain. Acanthamoeba was also obtained from the brain of experimentally infected mice and was stained with various stains as mentioned above to determine the best stain for use in identifying the presence of this parasite in experimentally infected animals. The modified Field's stain gives a very good color contrast as compared with other stains. Furthermore, it takes only 20 s to be carried out using the least number of reagents, making it suitable for both laboratory and field use.

  9. An Acanthamoeba sp. containing two phylogenetically different bacterial endosymbionts

    PubMed Central

    Heinz, Eva; Kolarov, Irina; Kästner, Christian; Toenshoff, Elena R; Wagner, Michael; Horn, Matthias

    2007-01-01

    Acanthamoebae are ubiquitous free-living amoebae and important predators of microbial communities. They frequently contain obligate intracellular bacterial symbionts, which show a worldwide distribution. All Acanthamoeba spp. described so far harboured no or only a single specific endosymbiont phylotype, and in some cases evidence for coevolution between the symbiotic bacteria and the amoeba host has been reported. In this study we have isolated and characterized an Acanthamoeba sp. (strain OEW1) showing a stable symbiotic relationship with two morphologically different endosymbionts. 16S rRNA sequence analysis assigned these symbionts to the candidate genus Procabacter (Betaproteobacteria) and the genus Parachlamydia (Chlamydiae) respectively. Fluorescence in situ hybridization and transmission electron microscopy confirmed the affiliation of the endosymbionts and showed their co-occurrence in the amoeba host cells and their intracellular location within separate compartments enclosed by host-derived membranes. Further analysis of this stable relationship should provide novel insights into the complex interactions of intracellular multiple-partner associations. PMID:17504498

  10. Efficiency of water disinfectants against Legionella pneumophila and Acanthamoeba.

    PubMed

    Dupuy, Mathieu; Mazoua, Stéphane; Berne, Florence; Bodet, Charles; Garrec, Nathalie; Herbelin, Pascaline; Ménard-Szczebara, Florence; Oberti, Sandrine; Rodier, Marie-Hélène; Soreau, Sylvie; Wallet, France; Héchard, Yann

    2011-01-01

    Free-living amoebae might be pathogenic by themselves and be a reservoir for bacterial pathogens, such as Legionella pneumophila. Not only could amoebae protect intra-cellular Legionella but Legionella grown within amoebae could undergo physiological modifications and become more resistant and more virulent. Therefore, it is important to study the efficiency of treatments on amoebae and Legionella grown within these amoebae to improve their application and to limit their impact on the environment. With this aim, we compared various water disinfectants against trophozoites of three Acanthamoeba strains and L. pneumophila alone or in co-culture. Three oxidizing disinfectants (chlorine, monochloramine, and chlorine dioxide) were assessed. All the samples were treated with disinfectants for 1 h and the disinfectant concentration was followed to calculate disinfectant exposure (Ct). We noticed that there were significant differences of susceptibility among the Acanthamoeba strains. However no difference was observed between infected and non-infected amoebae. Also, the comparison between the three disinfectants indicates that monochloramine was efficient at the same level towards free or co-cultured L. pneumophila while chlorine and chlorine dioxide were less efficient on co-cultured L. pneumophila. It suggests that these disinfectants should have different modes of action. Finally, our results provide for the first time disinfectant exposure values for Acanthamoeba treatments that might be used as references for disinfection of water systems.

  11. Partial characterization of Acanthamoeba castellanii (T4 genotype) DNase activity.

    PubMed

    Iqbal, Junaid; Panjwani, Shamvil; Siddiqui, Ruqaiyyah; Khan, Naveed Ahmed

    2015-02-01

    The deoxyribonuclease (DNase) activities of Acanthamoeba castellanii belonging to the T4 genotype were investigated. Using zymographic assays, the DNase activities had approximate molecular masses of 25 and 35 kDa. A. castellanii DNases exhibited activity at wide-ranging temperature of up to 60 °C and at pH ranging from 4 to 9. The DNases activities were unaffected by proteinase-K treatment, divalent cations such as Ca(++), Cu(++), Mg(++), and Zn(++), or divalent cation chelating agent ethylenediaminetetraacetic acid (EDTA) or sodium dodecyl sulfate (SDS). The non-reliance on divalent cations and homology data suggests that A. castellanii DNases belong to the class of eukaryotic lysosomal DNase II but exhibit robust properties. The DNases activity in A. castellanii interfered with the genomic DNA extraction. Extraction methods involving EDTA, SDS, and proteinase-K resulted in low yield of genomic DNA. On the other hand, these methods resulted in high yield of genomic DNA from human cells suggesting the robust nature of A. castellanii DNases that are unaffected by reagents normally used in blocking eukaryotic DNases. In contrast, the use of chaotropic agent such as guanidine thiocyanate improved the yield of genomic DNA from A. castellanii cells significantly. Further purification and characterization of Acanthamoeba DNases is needed to study their non-classic distinct properties and to determine their role in the biology, cellular differentiation, cell cycle progression, and arrest of Acanthamoeba.

  12. Balamuthia mandrillaris and Acanthamoeba Amebic Encephalitis with Neurotoxoplasmosis Coinfection in a Patient with Advanced HIV Infection

    PubMed Central

    Chan, Joseph C.; Castellano-Sanchez, Amilcar; Hirzel, Alicia; Laowansiri, Panthipa; Tuda, Claudio; Visvesvara, Govinda S.; Qvarnstrom, Yvonne; Ratzan, Kenneth R.

    2012-01-01

    We describe a patient with advanced HIV infection and Balamuthia mandrillaris and Acanthamoeba amebic encephalitis with Toxoplasma gondii coinfection. A multidisciplinary effort and state-of-the-art diagnostic techniques were required for diagnosis. Our patient is the first reported case of an HIV-infected person with dual Balamuthia mandrillaris and Acanthamoeba amebic encephalitis with neurotoxoplasmosis coinfection. PMID:22170911

  13. Insights from the DNA databases: approaches to the phylogenetic structure of Acanthamoeba.

    PubMed

    Fuerst, Paul A

    2014-11-01

    Species of Acanthamoeba have been traditionally described using morphology (primarily cyst structure), or cytology of nuclear division (used by Pussard and Pons, 1977). Twenty-plus putative species were proposed based on such criteria. Morphology, however, is often plastic, dependent upon culture conditions. DNA sequences of the nuclear small subunit (18S) rRNA that can be used for the study of the phylogeny of Acanthamoeba have increased from a single sequence in 1986 to more than 1800 in 2013. Some of the patterns of the sequence data for Acanthamoeba are reviewed, and some of the insights that this data illuminates are illustrated. In particular, the data suggest the existence of 20 or more genotypic types, a number not dissimilar to the number of named species of Acanthamoeba. However, molecular studies make clear that the relationship between phylogenetic relatedness and species names as we know them for Acanthamoeba is tenuous at best.

  14. Isolation and Molecular Characterization of Acanthamoeba Strains from Dental Units in Costa Rica.

    PubMed

    Retana-Moreira, Lissette; Abrahams-Sandí, Elizabeth; Castro-Artavia, Esteban; Fernández-Sánchez, Ana; Castro-Castillo, Alfredo; Reyes-Batlle, María; Lorenzo-Morales, Jacob

    2015-01-01

    Free-living amoebae are protozoa widely distributed in nature, which can be found in a variety of environments. Four genera are recognized as causal agents of infections in humans and animals: Acanthamoeba, Naegleria, Balamuthia, and Sappinia. In this study, the presence of Acanthamoeba in dental units was determined and the isolates obtained were molecularly characterized; osmotolerance and thermotolerance assays were also performed to evaluate multiplication under these conditions, frequently associated with pathogenicity. The morphological analysis and partial sequencing of the 18S rDNA gene revealed the presence of Acanthamoeba genotype T4 in 14% of the units sampled. Osmotolerance and thermotolerance tests were positive for more than 80% of the isolates. Up to date, this is the first study that reports the detection, identification, and genotyping of Acanthamoeba isolated from dental units in Costa Rica and even in Latin-America. Further assays to determine the potential pathogenicity of these Acanthamoeba isolates are underway.

  15. Bacterial keratitis: a prospective clinical and microbiological study

    PubMed Central

    Schaefer, F.; Bruttin, O.; Zografos, L.; Guex-Crosier, Y.

    2001-01-01

    AIM—To define the clinical and microbiological profile of bacterial keratitis at the Jules Gonin Eye Hospital and to test the in vitro bacterial resistance.
METHODS—Patients presenting with bacterial keratitis were prospectively followed; clinical features (age, risk factors, visual acuity) and response to therapy were analysed. Bacteriological profile was determined and the sensitivity/resistance of isolated strains were tested towards 12 ocular antibiotics (NCCLS disc diffusion test).
RESULTS—85 consecutive patients (mean age 44.3 (SD 20.7) years) were prospectively enrolled from 1 March 1997 to 30 November 1998. The following risk factors were identified: contact lens wear, 36%; blepharitis, 21%; trauma, 20%; xerophthalmia, 15%; keratopathies, 8%; and eyelid abnormalities, 6%. The most commonly isolated bacteria were Staphylococcus epidermidis, 40%; Staphylococcus aureus, 22%; Streptococcus pneumoniae, 8%; others Streptococcus species, 5%; Pseudomonas, 9%; Moraxella and Serratia marcescens, 5% each; Bacillus, Corynebacterium, Alcaligenes xyloxidans, Morganella morganii, and Haemophilus influenza, 1% each. 1-15% of strains were resistant to fluoroquinolones, 13-22% to aminoglycosides, 37% to cefazolin, 18% to chloramphenicol, 54% to polymyxin B, 51% to fusidic acid, and 45% to bacitracin. Five of the 85 patients (5.8%) had a poor clinical outcome with a visual loss of one or more lines of visual acuity.
CONCLUSION—Fluoroquinolones appear to be the therapy of choice for bacterial keratitis, but, based upon these in vitro studies, some strains may be resistant.

 PMID:11423460

  16. Spectrum of fungal keratitis: clinicopathologic study of 44 cases

    PubMed Central

    Punia, Rajpal Singh; Kundu, Reetu; Chander, Jagdish; Arya, Sudesh Kumar; Handa, Uma; Mohan, Harsh

    2014-01-01

    AIM To determine the causative agents of fungal keratitis and study the predisposing factors over a period of ten years in a single tertiary care hospital. METHODS A retrospective analysis of fungal corneal ulcers was done from 2003-2012. Patients' clinical data were noted from the file records. Correlation of histopathological diagnosis was done with the report on fungal culture. RESULTS Mycotic keratitis was established in 44 cases by a positive fungal culture. Direct microscopic examination of potassium hydroxide (KOH) mounts revealed fungal elements in 39 cases while 40 cases showed fungus on Gram stained smears. Males (54.55%) were more commonly affected than the females (45.45%). The age ranged from 18 to 82 years. Most common age group to be involved was 41-60 years. Predisposing risk factors were seen in 34 (77.27%) cases. Most common findings on clinical examination were anterior chamber reaction and conjunctival injection seen in all the cases. Other common findings were stromal infiltration and hypopyon seen in 20 (45.45%) and 18 (40.91%) cases respectively. On histopathological examination the fungus was typed, as aspergillus in 34 cases while no definite typing was possible in 10 cases. The predominant isolate was aspergillus flavus (59.09%) followed by fusarium (15.91%). Mixed fungal and bacterial infection was seen in 3 (6.82%) cases. CONCLUSION Although culture is the gold standard for definitive diagnosis of fungal keratitis, direct microscopic examination of corneal scrapings or histomorphological evaluation of biopsies allow a rapid preliminary diagnosis. Early administration of antifungal treatment helps in preventing dreadful complications. PMID:24634875

  17. Peripheral ulcerative keratitis as a complication of acute myeloid leukaemia

    PubMed Central

    Morjaria, Rupal; Barge, Tom; Mordant, David; Elston, John

    2014-01-01

    We report a rare presentation of acute bilateral peripheral ulcerative keratitis (PUK) in a patient with a new diagnosis of untreated acute myeloid leukaemia (AML). To the best of our knowledge, this is the first report of PUK associated with untreated AML and we stress the importance of a rapid and thorough testing to exclude other diagnoses. The patient lost his vision within 10 days to counting fingers. Rapid diagnosis allowed a good visual recovery following prompt treatment with oral steroids and systemic chemotherapy treatment for the AML. PMID:25362188

  18. Surgical management of fungal endophthalmitis resulting from fungal keratitis

    PubMed Central

    Gao, Yan; Chen, Nan; Dong, Xiao-Guang; Yuan, Gong-Qiang; Yu, Bin; Xie, Li-Xin

    2016-01-01

    AIM To report the fungal organisms, clinical features, surgical treatment strategies, and outcomes of patients with culture-proven exogenous fungal endophthalmitis (EFE) secondary to keratitis, and evaluate the role of surgery in the treatment. METHODS The clinical records of 27 patients (27 eyes) with culture-proven EFE resulting from fungal keratitis treated at Shandong Eye Institute from January 2007 to January 2015 were retrospectively reviewed. Information about fungal culture results, clinical features, surgical procedures, and final visual acuity was obtained. RESULTS There were 39 positive culture results from samples of cornea, hypopyon, vitreous and lens capsule, accounting for 56%, 26%, 15% and 2.5%, respectively. Fusarium was identified in 44% (12/27) of the eyes, followed by Aspergillus in 22% (6/27). Posterior segment infection was involved in 78% (21/27) of the patients. The corneal infection was larger than 3 mm ×3 mm in 89% (24/27) of the patients, and 22% (6/27) of them had the entire cornea, and even the sclera involved. Three eyes had silicone oil tamponade, and two eyes had retinal detachment. Twenty-two eyes (81.5%) underwent penetrating keratoplasty (PKP), and over half of them (54.5%) were operated within 3d from the onset of antifungal therapy. Fourteen eyes (52%) underwent intracameral antifungal drug injection, and three of them required repeated injections. Fifteen eyes (55.6%) underwent pars plana vitrectomy (PPV). The rate of the eyes undergoing PPV as the initial surgical procedure was 60% (9/15), lower than 77% in PKP. Intravitreal injection was given in 59% of the eyes (16/27), and 75% of them required repeated injections. The final visual acuity was 20/100 or better in 37% of the eyes, and better than counting fingers in 55.6% of the eyes. Five eyes (18.5%) were eviscerated. In the two eyes with concurrent retinal detachment, one achieved retinal reattachment, and the other was eviscerated. In the three eyes with silicone oil

  19. Keratoprosthesis in pediatric keratitis-icthyosiform-deafness syndrome.

    PubMed

    Brown, Cortlyn; Rowlands, Megan; Lee, Daniel; Geffin, Joel A; Huang, John

    2016-02-01

    We report the first case of Boston Keratoprosthesis (KPro) implantation in a 7-year-old girl with keratitis-ichthyosiform-deafness syndrome and persistent, highly vascular corneal surface disease. An adult aphakic KPro with 8.5 mm backplate was implanted successfully and without operative or postoperative complications following 3 failed penetrating keratoplasties. Visual acuity improved from hand motions to 20/70 in the left eye 22 months after KPro surgery and from hand motion to hand motion with direction in the right eye. Glaucoma developed in the left eye, but intraocular pressure was successfully controlled with Ahmed valve placement and topical dorzolamide-timolol.

  20. Microsporidial keratitis in patients with hot springs exposure.

    PubMed

    Fan, Nai-Wen; Wu, Chih-Chiau; Chen, Te-Li; Yu, Wei-Kuang; Chen, Chien-Pei; Lee, Shui-Mei; Lin, Pei-Yu

    2012-02-01

    This retrospective study included 10 eyes of 9 patients diagnosed with microsporidial keratitis. All of them were known to contract this disease after taking baths in hot springs. The disease was diagnosed based on detecting microsporidia in corneal scrapings using Gram stain and the modified Kinyoun's acid-fast stain. The specimens from the last six patients were subjected to PCR and then sequencing. All of them revealed that the microorganism identified has a high similarity to Vittaforma corneae. Repeated debridement of the epithelial lesions successfully eradicated the microsporidial infection in all nine patients.

  1. Fonsecaea Pedrosoi: A Rare Etiology in Fungal Keratitis

    PubMed Central

    Sangwan, Jyoti; Lathwal, Sumit; Juyal, Deepak; Sharma, Neelam

    2013-01-01

    Fungal corneal ulcer is common in India due to tropical climate and a large agrarian population that is at risk. Fonsecaea pedrosoi is the most common agent of chromoblastomycosis, a chronic localized fungal infection of the skin and subcutaneous tissues mainly involving lower extremities. We report a rare case of corneal chromoblastomycosis caused by F.pedrosoi, which was successfully treated with topical Amphotericin B followed by a long course of oral antifungal therapy. To the best of our knowledge, this is the first case of mycotic keratitis caused by F.pedrosoi, from the foothills of Himalayas. PMID:24298496

  2. Biochemical characterization and functional studies of Acanthamoeba mannose-binding protein.

    PubMed

    Garate, Marco; Cubillos, Ibis; Marchant, Jeffrey; Panjwani, Noorjahan

    2005-09-01

    Acanthamoebae produce a painful, sight-threatening corneal infection. The adhesion of parasites to the host cells is a critical first step in the pathogenesis of infection. Subsequent to adhesion, the parasites produce a potent cytopathic effect (CPE) leading to target cell death. Recent studies showing that acanthamoebae express a mannose-binding protein (MBP) and that free alpha-mannose (alpha-Man) specifically inhibits the adhesion of parasites to host cells suggest that the MBP plays a key role in the pathogenesis of Acanthamoeba infection by mediating host-parasite interactions. However, direct evidence showing that Acanthamoeba MBP is a virulence protein has been lacking. In this study, we demonstrate that the polyclonal immunoglobulin Y (IgY) antibodies prepared against affinity-purified Acanthamoeba MBP markedly inhibit the adhesion of parasites to host cells. The antibody also inhibited the Acanthamoeba-induced CPE on host cells. In contrast, preimmune IgY did not influence either the adhesion of the parasites to host cells or the amoeba-induced CPE. Using a variety of approaches, including affinity chromatography on an alpha-Man gel, electrophoresis under native and denaturing conditions, biotinylation of cell surface proteins, and immunostaining, it was conclusively established that Acanthamoeba MBP is located on the surface membranes of the parasites. Neutral-sugar analysis and lectin binding experiments using succinylated concanavalin A, a plant lectin with high affinity for mannose, revealed that Acanthamoeba MBP is itself a mannose-containing glycoprotein. N-Glycanase treatment to remove N-linked oligosaccharides shifted the subunit molecular mass of MBP from 130 kDa to 110 kDa. Hexosamine analysis revealed that Acanthamoeba MBP lacks detectable levels of GalNAc, suggesting the absence of O-linked oligosaccharides. In summary, we have characterized Acanthamoeba MBP and have shown that it is a major virulence protein responsible for host

  3. A case of fungal keratitis and onychomycosis simultaneously infected by Trichophyton species

    PubMed Central

    2014-01-01

    Background Fungal keratitis is difficult to treat that can result in corneal blindness requiring penetrating keratoplasty and in fungal endothalmitis. We report a case of fungal keratitis and onychomycosis simultaneously infected by Trichophyton. Case presentation A 77-year old male presented with ocular pain, conjunctival injection, and severe loss of vision in his left eye. His best corrected visual acuity was hand movements in the left eye, and slit-lamp examination showed a corneal ulcer with feathery margin and hypopyon. Bacterial and fungal smear/culture showed no organism, and there was no improvement in spite of treatment with topical fortified 5% cefazolin and 2% tobramycin. Trichophyton species was identified by repeated cultures. We found onychomycosis on the patient’s foot, where the same fungal species were identified. Regimen was changed to topical itraconazole and systemic intravenous itraconazole. No clinical improvement was observed, so therapeutic penetrating keratoplasty and cryotherapy was done with continuation of antifungal therapy. The graft was clear at postoperative 1 month and no evidence of recurrence was found. Conclusion It is important to identify the pathogen of keratitis because early identification of pathogen causing keratitis provides the appropriate treatment in early phase of keratitis. It is necessary to search for other fungal skin infections such as onychomycosis and athelete’s foot considering the fungal keratitis following skin infection. In addition, fungal skin infection including onychomycosis should be treated for prevention of fungal keratitis as soon as possible. PMID:25015110

  4. The treatment of herpes simplex virus epithelial keratitis.

    PubMed Central

    Wilhelmus, K R

    2000-01-01

    PURPOSE: Epithelial keratitis is the most common presentation of ocular infection by herpes simplex virus (HSV). Quantitative assessment of available therapy is needed to guide evidence-based ophthalmology. This study aimed to compare the efficacy of various treatments for dendritic or geographic HSV epithelial keratitis and to evaluate the role of various clinical characteristics on epithelial healing. METHODS: Following a systematic review of the literature, information from clinical trials of HSV dendritic or geographic epithelial keratitis was extracted, and the methodological quality of each study was scored. Methods of epithelial cauterization and curettage were grouped as relatively equivalent physicochemical therapy, and solution and ointment formulations of a given topical antiviral agent were combined. The proportion healed with 1 week of therapy, a scheduled follow-up day that approximated the average time of resolution with antiviral therapy, was selected as the primary outcome based on a masked evaluation of maximum treatment differences in published healing curves. The proportion healed at 14 days was recorded as supplemental information. Fixed-effects and random-effects meta-analysis models were used to obtain summary estimates by pooling results from comparative treatment trials. Hypotheses about which prognostic factors might affect epithelial healing during antiviral therapy were developed by multivariate analysis of the Herpetic Eye Disease Study dataset. RESULTS: After excluding 48 duplicate reports, 14 nonrandomized studies, 15 studies with outdated or similar treatments, and 29 trials lacking sufficient data on healing or accessibility, 76 primary reports were identified. These reports involved 4,251 patients allocated to 93 treatment comparisons of dendritic epithelial keratitis in 28 categories and 9 comparisons of geographic epithelial keratitis in 6 categories. For dendritic keratitis, idoxuridine was better than placebo at 7 days

  5. Proteomic analysis of keratitis-associated Pseudomonas aeruginosa

    PubMed Central

    Sewell, Abby; Dunmire, Jeffrey; Wehmann, Michael; Rowe, Theresa

    2014-01-01

    Purpose To compare the proteomic profile of a clinical isolate of Pseudomonas aeruginosa (P. aeruginosa) obtained from an infected cornea of a contact lens wearer and the laboratory strain P. aeruginosa ATCC 10145. Methods Antibiotic sensitivity, motility, biofilm formation, and virulence tests were performed using standard methods. Whole protein lysates were analyzed with liquid chromatography/ tandem mass spectrometry (LC-MS/MS) in triplicate, and relative protein abundances were determined with spectral counting. The G test followed by a post hoc Holm-Sidak adjustment was used for the statistical analyses to determine significance in the differential expression of proteins between the two strains. Results A total of 687 proteins were detected. One-hundred thirty-three (133) proteins were significantly different between the two strains. Among these, 13 were upregulated, and 16 were downregulated in the clinical strain compared to ATCC 10145, whereas 57 were detected only in the clinical strain. The upregulated proteins are associated with virulence and pathogenicity. Conclusions Proteins detected at higher levels in the clinical strain of P. aeruginosa were proteins known to be virulence factors. These results confirm that the keratitis-associated P. aeruginosa strain is pathogenic and expresses a higher number of virulence factors compared to the laboratory strain ATCC 10145. Identification of the protein profile of the corneal strain of P. aeruginosa in this study will aid in elucidating novel intervention strategies for reducing the burden of P. aeruginosa infection in keratitis. PMID:25221424

  6. A randomised clinical trial comparing 2% econazole and 5% natamycin for the treatment of fungal keratitis

    PubMed Central

    Prajna, N V; John, R K; Nirmalan, P K; Lalitha, P; Srinivasan, M

    2003-01-01

    Aim: To compare 2% econazole and 5% natamycin in the management of fungal keratitis. Methods: A randomised clinical trial was performed using 2% econazole or 5% natamycin as the two treatment arms on patients presenting with culture positive fungal keratitis to the cornea service at Aravind Eye Care System, Madurai, India. Results: 116 patients were recruited, and 112 continued in the study. There were no significant differences between the two arms at baseline or for success (defined as a healed or healing ulcer) at final visit (p = 0.79). Conclusions: 2% Econazole appears to be as effective as 5% natamycin for the management of fungal keratitis. PMID:14507756

  7. Mechanisms associated with phagocytosis of Arcobacter butzleri by Acanthamoeba castellanii.

    PubMed

    Medina, Gustavo; Flores-Martin, Sandra; Fonseca, Belchiolina; Otth, Carola; Fernandez, Heriberto

    2014-05-01

    Acanthamoeba castellanii is a free-living amoeba widely found in environmental matrices such as soil and water. Arcobacter butzleri is an emerging potential zoonotic pathogen that can be isolated from environmental water sources, where they can establish endosymbiotic relationships with amoebas. The aim of this study was to describe the implication of mannose-binding proteins and membrane-associated receptors of glucose and galactose present in the amoebic membrane, during the attachment of Arcobacter butzleri by blocking with different saccharides. Another objective was to describe the signaling pathways involved in phagocytosis of these bacteria using specific inhibitors and analyze the implication of phagolysosome formation on the survival of Arcobacter butzleri inside the amoeba. We infer that the attachment of Arcobacter butzleri to the amoeba is a process which involves the participation of mannose-binding proteins and membrane-associated receptors of glucose and galactose present in the amoeba. We also demonstrated an active role of protozoan actin polymerization in the phagocytosis of Arcobacter butzleri and a critical involvement of PI3K and RhoA pathways. Further, we demonstrated that the tyrosine kinase-induced actin polymerization signal is essential in Acanthamoeba-mediated bacterial uptake. Through phagolysosomal formation analysis, we conclude that the survival of Arcobacter butzleri inside the amoeba could be related with the ability to remain inside vacuoles not fused with lysosomes, or with the ability to retard the fusion between these structures. All these results help the understanding of the bacterial uptake mechanisms used by Acanthamoeba castellanii and contribute to evidence of the survival mechanisms of Arcobacter butzleri.

  8. Susceptibility of Acanthamoeba castellanii to contact lens disinfecting solutions.

    PubMed

    Zanetti, S; Fiori, P L; Pinna, A; Usai, S; Carta, F; Fadda, G

    1995-07-01

    A corneal isolate of Acanthamoeba castellanii was exposed to commercial contact lens disinfecting solutions containing hydrogen peroxide, benzalkonium chloride, polyaminopropyl biguanide, polyquaternium 1, and chlorhexidine-thimerosal. The minimum trophozoite amebicidal concentration and exposure times required to kill trophozoites and cysts were determined. Solutions containing hydrogen peroxide or chlorhexidine-thimerosal were active against both trophozoites and cysts. The benzalkonium chloride-based solution was effective only against trophozoites. Solutions containing polyaminopropyl biguanide or polyquaternium 1 were completely ineffective. The need for adequate exposure times must be stressed.

  9. Susceptibility of Acanthamoeba castellanii to contact lens disinfecting solutions.

    PubMed Central

    Zanetti, S; Fiori, P L; Pinna, A; Usai, S; Carta, F; Fadda, G

    1995-01-01

    A corneal isolate of Acanthamoeba castellanii was exposed to commercial contact lens disinfecting solutions containing hydrogen peroxide, benzalkonium chloride, polyaminopropyl biguanide, polyquaternium 1, and chlorhexidine-thimerosal. The minimum trophozoite amebicidal concentration and exposure times required to kill trophozoites and cysts were determined. Solutions containing hydrogen peroxide or chlorhexidine-thimerosal were active against both trophozoites and cysts. The benzalkonium chloride-based solution was effective only against trophozoites. Solutions containing polyaminopropyl biguanide or polyquaternium 1 were completely ineffective. The need for adequate exposure times must be stressed. PMID:7492111

  10. Acanthamoeba castellanii: identification and distribution of actin cytoskeleton.

    PubMed

    González-Robles, Arturo; Castañón, Guadalupe; Hernández-Ramírez, Verónica Ivonne; Salazar-Villatoro, Lizbeth; González-Lázaro, Mónica; Omaña-Molina, Maritza; Talamás-Rohana, Patricia; Martínez-Palomo, Adolfo

    2008-07-01

    The presence of the cytoskeleton of Acanthamoeba castellanii was observed by means of cryo-electronmicroscopy and immunofluorescence techniques. This structure is formed largely by fibers and networks of actin located mainly in cytoplasmic locomotion structures as lamellipodia and as well as in various endocytic structures. In addition, the comparison between total actin content in whole extracts among different amoebae was made. The molecular weight of actin in A. castellanii was 44 kDa, and 45 kDa for Naegleria fowleri and Entamoeba histolytica.

  11. Pathogenic and nonpathogenic Acanthamoeba spp. in thermally polluted discharges and surface waters

    SciTech Connect

    de Jonckheere, J.F.

    1981-02-01

    During spring and autumn, the total number of amoebae and the number of acanthamoeba species able to grow at 37 degrees C were determined in six thermally polluted factory discharges and the surrounding surface waters. The isolated Acanthamoeba strains were studied for growth in axenic medium, cytopathic effect in Vito cell cultures, and virulence in mice. Although more amoebae were isolated in autumn, the number of Acanthamoeba species was lower than in spring, when the percent of pathogenic strains among the isolates was highest. Higher concentrations of amoebae were found in warm discharges, and more virulent strains occurred in thermal discharges than in surface waters.

  12. Acanthamoeba differentiation: a two-faced drama of Dr Jekyll and Mr Hyde.

    PubMed

    Siddiqui, Ruqaiyyah; Dudley, Ricky; Khan, Naveed Ahmed

    2012-06-01

    The ability of cyst-forming protists such as Acanthamoeba to escape death by transforming into a cyst form, that is resistant to harsh physiological, environmental and pharmacological conditions, has continued to pose a serious challenge to human and animal health. A complete understanding of the fundamental principles of genome evolution and biochemical pathways of cellular differentiation offers unprecedented opportunities to counter detrimental outcomes. Acanthamoeba can elude inhospitable conditions by forming cysts. Here we unravel the processes involved in the phenotypic switching of Acanthamoeba, which are critical in our efforts to find potential targets for chemotherapy. PMID:22309612

  13. Acanthamoeba differentiation: a two-faced drama of Dr Jekyll and Mr Hyde.

    PubMed

    Siddiqui, Ruqaiyyah; Dudley, Ricky; Khan, Naveed Ahmed

    2012-06-01

    The ability of cyst-forming protists such as Acanthamoeba to escape death by transforming into a cyst form, that is resistant to harsh physiological, environmental and pharmacological conditions, has continued to pose a serious challenge to human and animal health. A complete understanding of the fundamental principles of genome evolution and biochemical pathways of cellular differentiation offers unprecedented opportunities to counter detrimental outcomes. Acanthamoeba can elude inhospitable conditions by forming cysts. Here we unravel the processes involved in the phenotypic switching of Acanthamoeba, which are critical in our efforts to find potential targets for chemotherapy.

  14. Pathogenic and nonpathogenic Acanthamoeba spp. in thermally polluted discharges and surface waters.

    PubMed

    de Jonckheere, J F

    1981-02-01

    During spring and autumn, the total number of amoebae and the number of acanthamoeba species able to grow at 37 degrees C were determined in six thermally polluted factory discharges and the surrounding surface waters. The isolated Acanthamoeba strains were studied for growth in axenic medium, cytopathic effect in Vero cell cultures, and virulence in mice. Although more amoebae were isolated in autumn, the number of Acanthamoeba species was lower than in spring, when the percent of pathogenic strains among the isolates was highest. Higher concentrations of amoebae were found in warm discharges, and more virulent strains occurred in thermal discharges than in surface waters.

  15. Infectious keratitis with corneal perforation associated with corneal hydrops and contact lens wear in keratoconus.

    PubMed Central

    Donnenfeld, E D; Schrier, A; Perry, H D; Ingraham, H J; Lasonde, R; Epstein, A; Farber, B

    1996-01-01

    BACKGROUND: Corneal perforation is an uncommon complication associated with keratoconus. The first cases of infectious keratitis and corneal perforation associated with corneal hydrops and contact lens wear are reported in two keratoconus patients. METHODS: A retrospective chart review and histopathological examination were carried out. RESULTS: Both patients progressed to corneal perforation and emergency penetrating keratoplasty. One patient cultured Fusarium and the second patient Serratia marcesens. Both patients wore contact lenses against medical advice. CONCLUSIONS: The tear in Descement's membrane, stromal oedema, and epithelial bedewing associated with corneal hydrops results in loss of the epithelial-endothelial barrier of the cornea, creating a conduit for infectious organisms through the cornea. Acute hydrops associated with epithelial keratitis, stromal swelling, and a Descement's membrane tear may be a significant risk factor for infectious keratitis and corneal perforation. Contact lenses should not be worn during an active corneal hydrops owing to the increased risk for severe infectious keratitis and corneal perforation. Images PMID:8695560

  16. A case of fungal keratitis caused by Scopulariopsis brevicaulis: treatment with antifungal agents and penetrating keratoplasty.

    PubMed Central

    Ragge, N K; Hart, J C; Easty, D L; Tyers, A G

    1990-01-01

    A case of fungal keratitis caused by Scopulariopsis brevicaulis following a penetrating eye injury is described. Treatment with antifungal agents and keratoplasty resulted in a favourable outcome. Images PMID:2168203

  17. 46 CFR 7.170 - Alaska Peninsula, AK to Aleutian Islands, AK.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 1 2011-10-01 2011-10-01 false Alaska Peninsula, AK to Aleutian Islands, AK. 7.170... BOUNDARY LINES Alaska § 7.170 Alaska Peninsula, AK to Aleutian Islands, AK. (a) A line drawn from the southernmost extremity of Cape Kumlium to the westernmost extremity of Nakchamik Island; thence to...

  18. 46 CFR 7.165 - Kenai Peninsula, AK to Kodiak Island, AK.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 1 2010-10-01 2010-10-01 false Kenai Peninsula, AK to Kodiak Island, AK. 7.165 Section... BOUNDARY LINES Alaska § 7.165 Kenai Peninsula, AK to Kodiak Island, AK. (a) A line drawn from the southernmost extremity of Kenai Peninsula at longitude 151°44.0′ W. to East Amatuli Island Light; thence to...

  19. 46 CFR 7.170 - Alaska Peninsula, AK to Aleutian Islands, AK.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 1 2012-10-01 2012-10-01 false Alaska Peninsula, AK to Aleutian Islands, AK. 7.170... BOUNDARY LINES Alaska § 7.170 Alaska Peninsula, AK to Aleutian Islands, AK. (a) A line drawn from the southernmost extremity of Cape Kumlium to the westernmost extremity of Nakchamik Island; thence to...

  20. 46 CFR 7.165 - Kenai Peninsula, AK to Kodiak Island, AK.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 1 2014-10-01 2014-10-01 false Kenai Peninsula, AK to Kodiak Island, AK. 7.165 Section... BOUNDARY LINES Alaska § 7.165 Kenai Peninsula, AK to Kodiak Island, AK. (a) A line drawn from the southernmost extremity of Kenai Peninsula at longitude 151°44.0′ W. to East Amatuli Island Light; thence to...

  1. 46 CFR 7.170 - Alaska Peninsula, AK to Aleutian Islands, AK.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 1 2014-10-01 2014-10-01 false Alaska Peninsula, AK to Aleutian Islands, AK. 7.170... BOUNDARY LINES Alaska § 7.170 Alaska Peninsula, AK to Aleutian Islands, AK. (a) A line drawn from the southernmost extremity of Cape Kumlium to the westernmost extremity of Nakchamik Island; thence to...

  2. 46 CFR 7.170 - Alaska Peninsula, AK to Aleutian Islands, AK.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 1 2010-10-01 2010-10-01 false Alaska Peninsula, AK to Aleutian Islands, AK. 7.170... BOUNDARY LINES Alaska § 7.170 Alaska Peninsula, AK to Aleutian Islands, AK. (a) A line drawn from the southernmost extremity of Cape Kumlium to the westernmost extremity of Nakchamik Island; thence to...

  3. 46 CFR 7.165 - Kenai Peninsula, AK to Kodiak Island, AK.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 1 2012-10-01 2012-10-01 false Kenai Peninsula, AK to Kodiak Island, AK. 7.165 Section... BOUNDARY LINES Alaska § 7.165 Kenai Peninsula, AK to Kodiak Island, AK. (a) A line drawn from the southernmost extremity of Kenai Peninsula at longitude 151°44.0′ W. to East Amatuli Island Light; thence to...

  4. 46 CFR 7.165 - Kenai Peninsula, AK to Kodiak Island, AK.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 1 2011-10-01 2011-10-01 false Kenai Peninsula, AK to Kodiak Island, AK. 7.165 Section... BOUNDARY LINES Alaska § 7.165 Kenai Peninsula, AK to Kodiak Island, AK. (a) A line drawn from the southernmost extremity of Kenai Peninsula at longitude 151°44.0′ W. to East Amatuli Island Light; thence to...

  5. 46 CFR 7.170 - Alaska Peninsula, AK to Aleutian Islands, AK.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 1 2013-10-01 2013-10-01 false Alaska Peninsula, AK to Aleutian Islands, AK. 7.170... BOUNDARY LINES Alaska § 7.170 Alaska Peninsula, AK to Aleutian Islands, AK. (a) A line drawn from the southernmost extremity of Cape Kumlium to the westernmost extremity of Nakchamik Island; thence to...

  6. 46 CFR 7.165 - Kenai Peninsula, AK to Kodiak Island, AK.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 1 2013-10-01 2013-10-01 false Kenai Peninsula, AK to Kodiak Island, AK. 7.165 Section... BOUNDARY LINES Alaska § 7.165 Kenai Peninsula, AK to Kodiak Island, AK. (a) A line drawn from the southernmost extremity of Kenai Peninsula at longitude 151°44.0′ W. to East Amatuli Island Light; thence to...

  7. New long chain bases in lipophosphonoglycan of Acanthamoeba castellanii.

    PubMed

    Karaś, Magdalena A; Russa, Ryszard

    2013-06-01

    The polymer called lipophosphonoglycan (LPG) was isolated from Acanthamoeba castellanii membranes after exhaustive delipidation and butanol extraction. A novel extremely long phytosphingosine was revealed in glycoinositolphosphosphingolipid (GIPSL). All data obtained by gas-liquid chromatography coupled with MS analyses of products liberated during acid methanolysis and products of sodium metaperiodate and permanganate-periodate oxidations showed an unusual pattern of long chain bases (LCB) with branched bases (anteiso-C₂₄, anteiso-C₂₅, anteiso-C₂₆, iso-C₂₆, anteiso-C₂₇, and anteiso-C28) and normal ones (C₂₄, C₂₅, C₂₆, C₂₇). The phytosphingosines with hexa-, hepta-, and octacosanoic chains have not been detected in Acanthamoeba cells up to now. Also, the isomer configuration of long chain bases in LPG of A. castellanii was not defined in earlier reports. In the GC-MS chromatograms, the component forming a peak corresponding to anteiso-C₂₅ phytosphingosine was the most abundant and constituted more than 50 % of all LCB.

  8. [Amoebic meningoencephalitis by "Eaegleria" and "Acanthamoeba" (author's transl)].

    PubMed

    de Carneri, I

    1977-01-01

    Primary amoebic meningoencephalitis (ME) by Naegleria fowleri a free-living protozoon found in fresh, warm waters, is a well known fatal disease lasting less than one week. It affects sporadically swimmers and children playing in mud puddles. Less than 100 cases have been described. Recently a more rare, distinct amoebic meningoencephalitis due to some species of free-living Acanthamoeba was identified, lasting some weeks or more but still with a fatal evolution. In this case the amoebae do not always enter the brain directly through the cribrous membranes but cause mild, primary infections of respiratory airways: exceptionally, mainly in immunodepressed subjects, they then reach the CNS causing a secondary ME. Naegleria is fairly sensitive in vitro to some drugs, but in vivo their efficacy is dramtically lowered for pharmacokinetic reasons. Acanthamoeba is in every respect less sensitive. Prophylaxis is almost impossible to achieve. Some diagnostic procedures are described and the importance of their use in diagnosis of the so called aseptic purulent ME is stressed. PMID:616244

  9. Diagnosis of Fusarium keratitis in an animal model using the polymerase chain reaction

    PubMed Central

    Alexandrakis, G.; Jalali, S.; Gloor, P.

    1998-01-01

    AIMS/BACKGROUND—The purpose of this study was apply the polymerase chain reaction (PCR) to develop a sensitive, specific, and rapid test to diagnose Fusarium keratitis. Fusarium is the most common cause of fungal corneal infection in some parts of the world. It is often difficult to establish that a keratitis is due to fungal infection.
METHODS—Fusarium solani keratitis was induced in three eyes of three rabbits by injection of a suspension of the fungus into the anterior corneal stroma. In one rabbit the contralateral eye served as a control. From four to 28 days after inoculation, the corneas were scraped for culture, then scraped and swabbed for PCR analysis. The PCR was performed with primers directed against a portion of the Fusarium cutinase gene, and the presence or absence of this amplified target sequence was determined by agarose gel.
RESULTS—The amplified DNA sequence was detected in 25 of 28 samples from the corneas infected with Fusarium, for a sensitivity of 89%. Only three of the 14 samples from these eyes with Fusarium keratitis were positive by culture, for a sensitivity of 21%. Seven of eight control samples were negative by the PCR based test, for a specificity of 88%.
CONCLUSION—This PCR based test holds promise of being an effective method of diagnosing Fusarium keratitis as well as Fusarium infections at other sites.

 Keywords: keratitis; Fusarium; ulcer; cornea; polymerase chain reaction PMID:9602631

  10. A 5-Year Retrospective Review of Fungal Keratitis at Hospital Universiti Sains Malaysia

    PubMed Central

    Mohd-Tahir, Fadzillah; Norhayati, A.; Siti-Raihan, Ishak; Ibrahim, M.

    2012-01-01

    Background. Corneal blindness from healed infected keratitis is one of the most preventable causes of monocular blindness in developing countries, including Malaysia. Our objectives were to identify the causative fungi, predisposing risk factors, the proportion of correct clinical diagnosis, and visual outcome of patients treated in our hospital. Methods. A retrospective review of medical and microbiology records was conducted for all patients who were treated for fungal keratitis at Hospital Universiti Sains Malaysia from January 2007 until December 2011. Results. Forty-seven patients (47/186, 25.27%) were treated for fungal keratitis during the study period. This demonstrated that the incidence of fungal keratitis has increased each year from 2007 to 2011 by 12.50%, 17.65%, 21.21%, 26.83%, and 28.57%, respectively. The most common predisposing factors were injury to the eye followed by use of topical steroid, and preexisting ocular surface disease. Fusarium species were the most common fungal isolated, followed by Candida species. Clinical diagnosis of fungal keratitis was made in 26 of the 41 (63.41%) cases of positive isolates. Of these, in eleven cases (23.40%) patients required surgical intervention. Clinical outcome of healed scar was achieved in 34 (72.34%) cases. Conclusions. The percentage of positive fungal isolated has steadily increased and the trend of common fungal isolated has changed. The latest review regarding fungal keratitis is important for us to improve patients' outcome in the future. PMID:23304138

  11. Role of steroids in the treatment of bacterial keratitis.

    PubMed

    Palioura, Sotiria; Henry, Christopher R; Amescua, Guillermo; Alfonso, Eduardo C

    2016-01-01

    Bacterial keratitis can lead to severe visual impairment from corneal ulceration, subsequent scarring, and possible perforation. The mainstay of treatment is topical antibiotics, whereas the use of adjunctive topical corticosteroid drops remains a matter of debate. Herein, we review the rationale for and against the use of topical corticosteroids and we assess their effectiveness and safety in the published randomized controlled trials that have evaluated their role as adjunctive therapy for bacterial corneal ulcers. In the largest study to date, the Steroids for Corneal Ulcers Trial, topical corticosteroid drops were neither helpful nor harmful for the 500 participants as a whole. However, subgroup analyses suggested that topical corticosteroids may be beneficial upon early administration (within 2-3 days after starting antibiotics) for more central corneal ulcers with poorer vision at presentation, for invasive Pseudomonas strains, and for non-Nocardia ulcers. These results are discussed within the limitations of the study.

  12. Interface infectious keratitis following deep anterior lamellar keratoplasty

    PubMed Central

    Kodavoor, Shreesha Kumar; Dandapani, Ramamurthy; Kaushik, Ajay Ramesh

    2016-01-01

    A 32-year-old female patient underwent deep anterior lamellar keratoplasty (DALK) in both eyes for advanced keratoconus. She developed an infiltrate in the nasal paracentral interface of the left eye. In view of the paracentral location, localized epithelial removal/stromal scraping was done and the infiltrate was removed using forceps and sent for microbiological assay, which revealed fungal filaments. The patient successfully responded to intensive topical antifungal therapy with the maintenance of visual acuity. Interface keratitis following DALK frequently needs graft lift/interface wash due to deep location, rapid spread and poor penetration and efficacy of topical medications. In view of a paracentral location, modified debulking with topical therapy resulted in a satisfactory outcome in our case thereby avoiding the need for more invasive treatments. PMID:27688284

  13. Azole Antifungal Agents To Treat the Human Pathogens Acanthamoeba castellanii and Acanthamoeba polyphaga through Inhibition of Sterol 14α-Demethylase (CYP51)

    PubMed Central

    Lamb, David C.; Warrilow, Andrew G. S.; Rolley, Nicola J.; Parker, Josie E.; Nes, W. David; Smith, Stephen N.; Kelly, Diane E.

    2015-01-01

    In this study, we investigate the amebicidal activities of the pharmaceutical triazole CYP51 inhibitors fluconazole, itraconazole, and voriconazole against Acanthamoeba castellanii and Acanthamoeba polyphaga and assess their potential as therapeutic agents against Acanthamoeba infections in humans. Amebicidal activities of the triazoles were assessed by in vitro minimum inhibition concentration (MIC) determinations using trophozoites of A. castellanii and A. polyphaga. In addition, triazole effectiveness was assessed by ligand binding studies and inhibition of CYP51 activity of purified A. castellanii CYP51 (AcCYP51) that was heterologously expressed in Escherichia coli. Itraconazole and voriconazole bound tightly to AcCYP51 (dissociation constant [Kd] of 10 and 13 nM), whereas fluconazole bound weakly (Kd of 2,137 nM). Both itraconazole and voriconazole were confirmed to be strong inhibitors of AcCYP51 activity (50% inhibitory concentrations [IC50] of 0.23 and 0.39 μM), whereas inhibition by fluconazole was weak (IC50, 30 μM). However, itraconazole was 8- to 16-fold less effective (MIC, 16 mg/liter) at inhibiting A. polyphaga and A. castellanii cell proliferation than voriconazole (MIC, 1 to 2 mg/liter), while fluconazole did not inhibit Acanthamoeba cell division (MIC, >64 mg/liter) in vitro. Voriconazole was an effective inhibitor of trophozoite proliferation for A. castellanii and A. polyphaga; therefore, it should be evaluated in trials versus itraconazole for controlling Acanthamoeba infections. PMID:26014948

  14. Ciprofloxacin susceptibility of Pseudomonas aeruginosa isolates from keratitis

    PubMed Central

    Lomholt, J A; Kilian, M

    2003-01-01

    Aim: To examine the ciprofloxacin susceptibility of 106 Pseudomonas aeruginosa eye isolates from the United Kingdom, Denmark, India, the United States, and Australia, and to determine the molecular mechanisms of resistance. Methods: Ciprofloxacin susceptibility was tested by an agar dilution method; genomic DNA corresponding to the quinolone target genes gyrA and parC, and the regulatory genes mexR and nfxB controlling drug efflux systems, was amplified by PCR and sequenced; multilocus enzyme electrophoresis was performed to examine the genetic relation among resistant strains. Results: Three out of 90 keratitis isolates (3.3%), one from the United Kingdom and two from India, exhibited MIC values of 16 mg/l or 32 mg/l. The UK isolate had a mutation in gyrA (Thr83Ile), whereas the two Indian isolates showed mutations in both gyrA (Thr83Ile) and parC (Ser87Leu). The remaining isolates from keratitis, endophthalmitis, contact lens associated red eye (CLARE), and contact lens storage cases showed MIC values below 1 mg/l. Several allelic forms of gyrA and a single variation in the mexR gene product were detected in 10 ciprofloxacin susceptible strains. Conclusions: The vast majority of eye isolates of P aeruginosa from European countries are fully susceptible to ciprofloxacin and the concentration of ciprofloxacin eye drops used for local treatment (3000 mg/l) exceeds MIC values for strains recorded as resistant. Mutations in more than one target gene were associated with higher MIC values. PMID:14507757

  15. Down-regulation of cellulose synthase inhibits the formation of endocysts in Acanthamoeba.

    PubMed

    Moon, Eun-Kyung; Hong, Yeonchul; Chung, Dong-Il; Goo, Youn-Kyoung; Kong, Hyun-Hee

    2014-04-01

    Acanthamoeba cysts are resistant to unfavorable physiological conditions and various disinfectants. Acanthamoeba cysts have 2 walls containing various sugar moieties, and in particular, one third of the inner wall is composed of cellulose. In this study, it has been shown that down-regulation of cellulose synthase by small interfering RNA (siRNA) significantly inhibits the formation of mature Acanthamoeba castellanii cysts. Calcofluor white staining and transmission electron microscopy revealed that siRNA transfected amoeba failed to form an inner wall during encystation and thus are likely to be more vulnerable. In addition, the expression of xylose isomerase, which is involved in cyst wall formation, was not altered in cellulose synthase down-regulated amoeba, indicating that cellulose synthase is a crucial factor for inner wall formation by Acanthamoeba during encystation.

  16. Juglone induces cell death of Acanthamoeba through increased production of reactive oxygen species.

    PubMed

    Jha, Bijay Kumar; Jung, Hui-Jung; Seo, Incheol; Suh, Seong-Il; Suh, Min-Ho; Baek, Won-Ki

    2015-12-01

    Juglone (5-hydroxy-1,4-naphthoquinone) is a major chemical constituent of Juglans mandshruica Maxim. Recent studies have demonstrated that juglone exhibits anti-cancer, anti-bacterial, anti-viral, and anti-parasitic properties. However, its effect against Acanthamoeba has not been defined yet. The aim of this study was to investigate the effect of juglone on Acanthamoeba. We demonstrate that juglone significantly inhibits the growth of Acanthamoeba castellanii at 3-5 μM concentrations. Juglone increased the production of reactive oxygen species (ROS) and caused cell death of A. castellanii. Inhibition of ROS by antioxidant N-acetyl-l-cysteine (NAC) restored the cell viability. Furthermore, our results show that juglone increased the uptake of mitochondrial specific dye. Collectively, these results indicate that ROS played a significant role in the juglone-induced cell death of Acanthamoeba.

  17. Juglone induces cell death of Acanthamoeba through increased production of reactive oxygen species.

    PubMed

    Jha, Bijay Kumar; Jung, Hui-Jung; Seo, Incheol; Suh, Seong-Il; Suh, Min-Ho; Baek, Won-Ki

    2015-12-01

    Juglone (5-hydroxy-1,4-naphthoquinone) is a major chemical constituent of Juglans mandshruica Maxim. Recent studies have demonstrated that juglone exhibits anti-cancer, anti-bacterial, anti-viral, and anti-parasitic properties. However, its effect against Acanthamoeba has not been defined yet. The aim of this study was to investigate the effect of juglone on Acanthamoeba. We demonstrate that juglone significantly inhibits the growth of Acanthamoeba castellanii at 3-5 μM concentrations. Juglone increased the production of reactive oxygen species (ROS) and caused cell death of A. castellanii. Inhibition of ROS by antioxidant N-acetyl-l-cysteine (NAC) restored the cell viability. Furthermore, our results show that juglone increased the uptake of mitochondrial specific dye. Collectively, these results indicate that ROS played a significant role in the juglone-induced cell death of Acanthamoeba. PMID:26358271

  18. Detection of Vibrio cholerae and Acanthamoeba species from same natural water samples collected from different cholera endemic areas in Sudan

    PubMed Central

    2011-01-01

    Background Vibrio cholerae O1 and V. cholerae O139 infect humans, causing the diarrheal and waterborne disease cholera, which is a worldwide health problem. V. cholerae and the free-living amoebae Acanthamoeba species are present in aquatic environments, including drinking water and it has shown that Acanthamoebae support bacterial growth and survival. Recently it has shown that Acanthamoeba species enhanced growth and survival of V. cholerae O1 and O139. Water samples from different cholera endemic areas in Sudan were collected with the aim to detect both V. cholerae and Acanthamoeba species from same natural water samples by polymerase chain reaction (PCR). Findings For the first time both V. cholerae and Acanthamoeba species were detected in same natural water samples collected from different cholera endemic areas in Sudan. 89% of detected V. cholerae was found with Acanthamoeba in same water samples. Conclusions The current findings disclose Acanthamoedae as a biological factor enhancing survival of V. cholerae in nature. PMID:21470437

  19. [Encephalitis due to Naegleria and Acanthamoeba. Comparison of organisms and diseases (author's transl)].

    PubMed

    Martinez, A J; Janitschke, K

    1979-04-01

    Naegleria and Acanthamoeba are ubiquitous, free-living amoebas. Infections with Naegleria are acquired nasally by exposure to water and are characterized by an acute fulminant hemorrhagic necrotizing meningoencephalitis leading to death. Acanthamoeba-infections occur in chronically ill, debilitated individuals. A patchy chronic or subacute granulomatous encephalitis is produced by hematogenous spread of the amoebas. The histological or clinical diagnosis is not difficult. PMID:457190

  20. Vectorial role of Acanthamoeba in Legionella propagation in water for human use.

    PubMed

    Magnet, A; Peralta, R H S; Gomes, T S; Izquierdo, F; Fernandez-Vadillo, C; Galvan, A L; Pozuelo, M J; Pelaz, C; Fenoy, S; Del Águila, C

    2015-02-01

    Legionella spp. is the causative agent of Legionnaires' disease and is transmitted through aerosols emanating from man-made water systems. Legionella resistance to water treatments has been related to its association with environmental amoebae such as Acanthamoeba. Due to the high presence of this protozoon in Spain and the high rate of notification of Legionnaires' disease of this country, the aims of this work were to study the coexistence of these bacteria and protozoa in water as well as their interaction. The usefulness of Acanthamoeba co-culture for the isolation of environmental Legionella was also studied. For this purpose, 70 water samples were collected in 2011 from three Drinking Water Treatment Plants, three Wastewater Treatment Plants and five Natural Pools in Spain. Acanthamoeba was found by PCR in 87.1% (61/70) samples and, by culture in 85.7% (60/70) samples. Legionella was detected by PCR in 58.6% (41/70) of water samples, in 5.7% (4/70) by agar culture and 75.7% (53/70) by Acanthamoeba co-culture. From the 54 Acanthamoeba water isolates, Legionella was detected in 43 of them independently of Acanthamoeba's genotype (T3, T4 and T11). Legionella feeleii, Legionella birminghamiensis, Legionella gresilensis/berliardensis, Legionella fairfieldensis, Legionella drozanski and Legionella falloni were identified. In conclusion, our results showed that environmental Acanthamoeba is infected by Legionella to a high percentage, and due to its ubiquity, high resistance and its pathogenic potential per se, new methods for its elimination should be studied. Also, the high effectivity of Acanthamoeba co-culture for Legionella detection has been shown. PMID:25461091

  1. Vectorial role of Acanthamoeba in Legionella propagation in water for human use.

    PubMed

    Magnet, A; Peralta, R H S; Gomes, T S; Izquierdo, F; Fernandez-Vadillo, C; Galvan, A L; Pozuelo, M J; Pelaz, C; Fenoy, S; Del Águila, C

    2015-02-01

    Legionella spp. is the causative agent of Legionnaires' disease and is transmitted through aerosols emanating from man-made water systems. Legionella resistance to water treatments has been related to its association with environmental amoebae such as Acanthamoeba. Due to the high presence of this protozoon in Spain and the high rate of notification of Legionnaires' disease of this country, the aims of this work were to study the coexistence of these bacteria and protozoa in water as well as their interaction. The usefulness of Acanthamoeba co-culture for the isolation of environmental Legionella was also studied. For this purpose, 70 water samples were collected in 2011 from three Drinking Water Treatment Plants, three Wastewater Treatment Plants and five Natural Pools in Spain. Acanthamoeba was found by PCR in 87.1% (61/70) samples and, by culture in 85.7% (60/70) samples. Legionella was detected by PCR in 58.6% (41/70) of water samples, in 5.7% (4/70) by agar culture and 75.7% (53/70) by Acanthamoeba co-culture. From the 54 Acanthamoeba water isolates, Legionella was detected in 43 of them independently of Acanthamoeba's genotype (T3, T4 and T11). Legionella feeleii, Legionella birminghamiensis, Legionella gresilensis/berliardensis, Legionella fairfieldensis, Legionella drozanski and Legionella falloni were identified. In conclusion, our results showed that environmental Acanthamoeba is infected by Legionella to a high percentage, and due to its ubiquity, high resistance and its pathogenic potential per se, new methods for its elimination should be studied. Also, the high effectivity of Acanthamoeba co-culture for Legionella detection has been shown.

  2. Standardized Method of Measuring Acanthamoeba Antibodies in Sera from Healthy Human Subjects

    PubMed Central

    Chappell, Cynthia L.; Wright, John A.; Coletta, Michael; Newsome, Anthony L.

    2001-01-01

    Acanthamoeba species can cause serious, debilitating, and sometimes life-threatening infections. Three groups have been identified using morphological and immunological comparisons. Previous serological studies have utilized a variety of antigen preparations and assay methods and reported disparate (3 to 100%) results. This study was designed to (i) optimize an enzyme-linked immunosorbent assay for detecting serum antibodies to each of the Acanthamoeba serogroups and (ii) test 55 healthy individuals for specific immunoglobulin G reactivity. The highest signal-to-background ratio was found when 3,000 fixed, intact trophozoites per well were used with a 1:10 serum dilution. Sera yielding optical densities of <0.25 against all three Acanthamoeba serogroups were used to define the cutoff for positive results. The highest background reactivity with these sera was seen with Acanthamoeba polyphaga (serogroup 2), followed by Acanthamoeba culbertsoni (serogroup 3) and Acanthamoeba astronyxis (serogroup 1). Of 55 subjects tested, the highest number of positive results was seen with A. polyphaga (81.8%), followed by A. astronyxis (52.8%) and A. culbertsoni (40%). Seven serum samples (12.7%) were negative for all three Acanthamoeba serogroups, 16 (29.1%) were positive for one serogroup only, 16 were positive for two serogroups, and 16 reacted to all three serogroups. Further analysis showed no significant associations between serogroup reactivity and age or gender. However, some ethnic differences were noted, especially with A. polyphaga antigens. In that case, serum samples from Hispanic subjects were 14.5 times less likely to be positive (P = 0.0025) and had lower mean absorbance values (P = 0.047) than those from Caucasian subjects. Overall, these data suggest that Acanthamoeba colonization or infection is more common than previously thought. Mild or asymptomatic infections may contribute to the observed serum reactivities. PMID:11427418

  3. In the case of transmission of Mycobacterium ulcerans in buruli ulcer disease Acanthamoeba species stand accused.

    PubMed

    Wilson, M D; Boakye, D A; Mosi, L; Asiedu, K

    2011-03-01

    Buruli ulcer disease caused by Mycobacterium ulcerans results in extensive destruction of skin and soft tissue and long-term functional disabilities that ultimately require surgery and rehabilitation. The disease is associated with aquatic and swampy environments with the mycobacterium occurring in biofilms, soil, aquatic insects, fish and wildlife however, the mode of transmission to humans remains an enigma. Current transmission ideas including bites from predatory water bugs and mosquitoes, do not explain satisfactorily the spasmodic disease distribution in human populations. Here we argue that Acanthamoeba species are the natural hosts of M. ulcerans and are mainly responsible for disease transmission because; (i) Acanthamoebae are known natural hosts of several microbial pathogens including M. marinum, M. avium and Legionella pneumophila, (ii) culture of slow-to-grow microbial pathogens hosted in nature by Acanthamoeba spp is enhanced when the media is seeded with the protozoa, (iii) acanthamoebae and M. ulcerans share similar bio-ecological and epidemiological settings, (iv) documented evidence that prior growth of L. pneumophila and M. avium in acanthamoebae influences entry mechanisms, intracellular growth and virulence in human monocytes, (v) Acanthamoeba spp also infect humans and cause diseases via routes of openings including broken skin and sites of trauma similar to M. ulcerans and (vi) M. ulcerans is rather a fastidious intracellular organism as recent analysis of the genome indicate. We argue further that temperature plays a significant role in transmission determining the fate of either the intracellular microbe or the host cells. Also, Acanthamoeba-pathogen association has a long evolutionary history because the same set of bacterial genes and gene products e.g. in L. pneumophila are required for survival in both mammalian and protozoan host cells. We suggest that the involvement of Acanthamoeba in the transmission of M. ulcerans to humans better

  4. Molecular characterization of bacterial endosymbionts of Acanthamoeba isolates from infected corneas of Korean patients.

    PubMed

    Xuan, Ying-Hua; Yu, Hak Sun; Jeong, Hae Jin; Seol, Sung-Yong; Chung, Dong-Il; Kong, Hyun-Hee

    2007-03-01

    The endosymbionts of 4 strains of Acanthamoeba (KA/E9, KA/E21, KA/E22, and KA/E23) isolated from the infected corneas of Korean patients were characterized via orcein stain, transmission electron microscopic examination, and 16S rDNA sequence analysis. Double membrane-bound, rod-shaped endosymbionts were distributed randomly throughout both the trophozoites and cysts of each of Acanthamoeba isolates. The endosymbionts of KA/E9, KA/E22, and KA/E23 were surrounded by electron-translucent areas. No lacunae-like structures were observed in the endosymbionts of KA/E21, the bacterial cell walls of which were studded with host ribosomes. Comparative analyses of the 16S rDNA sequences showed that the endosymbionts of KA/E9, KA/E22 and KA/E23 were closely related to Caedibacter caryophilus, whereas the KA/E21 endosymbiont was assigned to the Cytophaga-Flavobacterium-Bacteroides (CFB) phylum. In the 4 strains of Acanthamoeba, the hosts of the endosymbionts were identified as belonging to the Acanthamoeba castellanii complex, which corresponds to the T4 genotype. Acanthamoeba KA/E21 evidenced characteristics almost identical to those of KA/E6, with the exception of the existence of endosymbionts. The discovery of these endosymbionts from Acanthamoeba may prove essential to future studies focusing on interactions between the endosymbionts and the amoebic hosts. PMID:17374972

  5. Evaluation of Ozone Application in Dental Unit Water Lines Contaminated with Pathogenic Acanthamoeba

    PubMed Central

    HIKAL, Wafaa; ZAKI, Basma; SABRY, Hany

    2015-01-01

    Background: In this study morphological and molecular characterization of Acanthamoeba strains, isolated from dental unit waterlines (DUWLs) were surveyed and the levels of disinfection achievable in vitro by the application of ozone disinfectant to DUWLs were evaluate. Methods: Water samples were collected from air-water syringes, cup fillers and tap water before and at the end of the working day. They were cultured on non-nutrient agar (NNA) plates. Species identification was carried out with a PCR assay based on sequence analysis of the 18S rRNA gene. The cellular response to ozone was tested on Acanthamoeba cyst with different doses at different contact time in vitro twice. Results: Prevalence rates for Acanthamoeba contamination were 100, 100 and 72% for air-water syringes, cup fillers and tap water, respectively. The morphological analysis revealed the presence of A. castellanii, A. griffin, A. hatchitti and A. lenticulata. Phylogenetic analysis of the sequences showed the four strains to be closely related to a sequence type (T3, T4, T5 and T11). Acanthamoeba cells were stained with trypan blue, which revealed killed of Acanthamoeba instantaneously after 10 minutes in ozonized water. There was no growth of Acanthamoeba occurred after ozone treatment in water bottles for 5 minutes with a flow rate of 500 mg/hour. Conclusion : Ozone can play an important role in controlling the problem of contamination of DUWLs as a potent disinfectant. PMID:26622296

  6. [Acanthamoeba isolated from child thymus: Acanthamoeba growth during cocultivation with human and murine eukaryotic cells and induction of cytopathogenic effect on these cells].

    PubMed

    Komogorova, V V; Sharova, N I; Gordeeva, L M; Iarilin, A A

    2003-01-01

    A cell line of the ameba assigned to the genus Acanthamoeba by morphological and cariotypic signs and by the specific features of its life cycle was isolated from the thymus of a child operated on for heart disease. The line received the name CDHT (Cells Derived from Human Thymus). Actively proliferating mammalian cells maintain the multiplication of Acanthamoeba cells in vitro. Using thymocytes as an example, it was shown that death of these cells occurred through the mechanism of apoptosis. The human thymocytes preincubated with the supernatant obtained through the cocultivation of CDHT and thymocytes may also undergo apoptosis. PMID:12886588

  7. Nitrogen Mineralization by Acanthamoeba polyphaga in Grazed Pseudomonas paucimobilis Populations

    PubMed Central

    Sinclair, James L.; McClellan, J. Forbes; Coleman, David C.

    1981-01-01

    Nitrogen mineralization was studied in a simple grazing system in which the protozoan Acanthamoeba polyphaga was grown with the bacterium Pseudomonas paucimobilis (two soil organisms isolated from the shortgrass prairie in northern Colorado). In different experiments, either carbon or nitrogen was adjusted to be in limiting amounts. When carbon was limiting, grazers were almost entirely responsible for nitrogen mineralization, with bacteria themselves contributing little. When nitrogen was limiting, nitrogen mineralization by grazers permitted continued growth by the grazed bacteria and a greater bacterial biomass production. The increased growth of the grazed bacteria did not result in an increased total amount of carbon used, but the grazed bacteria used carbon more efficiently than the ungrazed bacteria. PMID:16345864

  8. Arcobacter butzleri survives within trophozoite of Acanthamoeba castellanii.

    PubMed

    Villanueva, María P; Medina, Gustavo; Fernández, Heriberto

    2016-01-01

    The survival of three Arcobacter butzleri strains inside Acanthamoeba castellanii was assessed using axenic cultures of A. castellanii that were inoculated with the tested strains and incubated at 26°C under aerobic conditions for 240h. The behavior of bacteria in contact with amoebae was monitored using phase contrast microscopy. The bacterial survival rate within amoebae was assessed through counting colony forming units, using the gentamicin protection assay. All A. butzleri strains were able to survive during 240h within the amoebae, thus suggesting that (i) A. butzleri resists the amoebic digestion processes at least for the analyzed time; (ii) that A. castellanii could serve as an environmental reservoir for this bacterium, probably acting as a transmission vehicle for A. butzleri. PMID:26972277

  9. Cryptococcus neoformans: pseudohyphal forms surviving culture with Acanthamoeba polyphaga.

    PubMed Central

    Neilson, J B; Ivey, M H; Bulmer, G S

    1978-01-01

    During experiments on the gastrointestinal tract as a possible portal of entry for Cryptococcus neoformans, we occasionally observed the free-living amoeba, Acanthamoeba polyphaga, growing in the presence of C. neoformans cultured from mouse feces. Examination of the amoebic trophozoites revealed that they were engorged with yeast cells. Over a period of 2 to 3 weeks of incubation, the amoebae apparently killed most of the yeast cells. Some of the surviving C. neoformans cells formed atypical colonies which contained pseudohyphae. Seven other strains have since been cultured with this amoeba. Pseudohyphal forms were found among the surviving colonies in all strains tested. Virulence studies were performed on one randomly selected pseudohyphal isolate from each of the eight strains of C. neoformans. Pseudohyphal isolates from seven of the eight strains failed to kill mice 30 days after intracranial inoculation. The potential role of soil amoebae in the control of C. neoformans in nature is discussed. Images PMID:352931

  10. Acanthamoeba and bacteria produce antimicrobials to target their counterpart

    PubMed Central

    2014-01-01

    Background In the microbial ecosystem, microbes compete for space and nutrients. Consequently, some have developed the ability to kill or inhibit the growth of other competing microbes by producing antimicrobial substances. As the ‘producer’ species are generally immune to these substances, their compounds act on the competing microbial species and give the producer more space and access to nutrients for growth. Many currently used antibiotics were developed by exploiting this potential of certain microbes. Findings Here, the free-living amoeba, Acanthamoeba castellanii, was investigated for its antibacterial activity against representative Gram positive and Gram negative bacteria, while bacterial isolates were tested for their anti-amoebic properties. Conditioned medium from A. castellanii showed remarkable bactericidal properties against methicillin-resistant Staphylococcus aureus (MRSA) exhibiting almost 100% kill rate, but had limited effect against Acinetobacter sp., Pseudomonas aeruginosa and vancomycin-resistant Enterococcus faecalis (VRE). Similarly, the conditioned medium of E. coli K1 and Enterobacter sp., exhibited potent anti-Acanthamoebic effects in a concentration-dependent manner. Conditioned media of Acanthamoeba, E. coli K1 and Enterobacter sp. showed no cytotoxicity in vitro when tested against human brain microvascular endothelial cells. Active molecule/s in aforementioned amoebic and two bacterial conditioned media were 5 – 10 kDa, and <5 kDa respectively. Conclusions A. castellanii conditioned medium showed potent bactericidal properties against MRSA. The active molecule(s) are heat- and pronase-resistant, and in the 5 to 10 kDa molecular mass range. Contrary to this, E. coli K1 and Enterobacter sp., conditioned medium showed anti-amoebic effects that are <5 kDa in molecular mass, suggestive of active metabolites. PMID:24479709

  11. Topical nanoparticulate formulation of drugs for ocular keratitis

    NASA Astrophysics Data System (ADS)

    Yang, Xiaoyan

    The primary objective of this project is to develop drug-loaded polymeric nanoparticles suspended in a biocompatible gel for topical delivery of therapeutic agents commonly employed in the treatment of ocular viral/bacterial keratitis. PART 1: Poly(D,L-lactic-co-glycolic acid) (PLGA) nanoparticles (NP) of dipeptide monoester prodrugs of ganciclovir (GCV) including L-Val-L-Val-GCV (LLGCV), L-Val-D-Val-GCV (LDGCV), D-Val-L-Val-GCV (DLGCV) were formulated and dispersed in thermosensitive PLGA-PEG-PLGA polymer gel for the treatment of herpes simplex virus type 1 (HSV-1) induced viral corneal keratitis. NP containing prodrugs of GCV were prepared by a double-emulsion solvent evaporation technique using various PLGA polymers with different drug/polymer ratios. Cytotoxicity studies suggested that all NP formulations are non-toxic. In vitro release of prodrugs from NP showed a biphasic release pattern with an initial burst phase followed by a sustained phase. Such burst effect was completely eliminated when NP were suspended in thermosensitive gels with near zero-order release kinetics. Prodrugs-loaded PLGA NP dispersed in thermosensitive gels can thus serve as a promising drug delivery system for the treatment of anterior eye diseases. Maximum uptake (around 60%) was noted at 3 h for NP. Cellular uptake and intracellular accumulation of prodrugs are significantly different among three stereoisomeric dipeptide prodrugs. The microscopic images show that NP are avidly internalized by HCEC cells and distributed throughout the cytoplasm instead of being localized on the cell surface. Following cellular uptake, prodrugs released from NP gradually bioreversed into parent drug GCV. LLGCV showed the highest degradation rate, followed by LDGCV and DLGCV. LLGCV, LDGCV and DLGCV released from NP exhibited superior uptake and bioreversion in corneal cells. PART 2: PLGA NP of hydrocortisone butyrate (HB) suspended in thermosensitive PLGA-PEG-PLGA gel were developed for the treatment of

  12. Risk factors for treatment outcome of suspected microbial keratitis

    PubMed Central

    Morlet, N.; Minassian, D.; Butcher, J.; the, O

    1999-01-01

    BACKGROUND—Primary treatment for suspected microbial keratitis is generally successful. Although risks such as contact lens use are well recognised as causative factors for microbial keratitis, little is known about the risk factors that influence treatment outcome. The present study evaluates the risk factors assessed at diagnosis as prognostic indicators of primary treatment failure.
METHODS—Patients were prospectively enrolled in the ofloxacin treatment trial and data concerning symptoms, treatments, past and concurrent eye disease were collected along with the measurement of corneal ulcer size at the slit lamp. All patients were scraped for microbiological investigation, and treated with either ofloxacin (0.3%) or standard therapy of fortified cefuroxime and gentamicin drops. Treatment success was complete healing of the ulcer with zero dimensions of the epithelial defect within 2 weeks of start of treatment. The important prognostic indicators were selected by comparison among those who failed treatment, had delayed healing, or were culture positive with other patients using univariate and stratified analysis. These were then used in a Poisson model for multiple regression analysis to estimate the relative risk of the main prognostic variables.
RESULTS—Of the 118 patients enrolled in the study, 14 were identified as primary treatment failures, 17 had slow healing, and 15 indolent ulcers. There were 49 culture positive patients. The multivariate analysis identified that large culture positive ulcers in patients 60 years or older had 5.5 times the risk of primary treatment failure (p<0.001). Significant predictors of slow healing were previous ocular disease and a positive culture; significant predictors of indolent ulceration were previous ocular disease and steroid use at diagnosis; the main predictor of a culture positive result was ulcer size.
CONCLUSIONS—Elderly patients with large ulcers were more likely to be culture positive, fail

  13. PACK-CXL: Corneal Cross-linking for Treatment of Infectious Keratitis.

    PubMed

    Tabibian, David; Richoz, Olivier; Hafezi, Farhad

    2015-01-01

    This article discusses corneal cross-linking (CXL) and how it transitioned from a modality for treating corneal ectatic disorders to an inventive means of treating infectious keratitis. Initially, CXL was successfully developed to halt the progression of ectatic diseases such as keratoconus, using the standard Dresden protocol. Later, indications were extended to treat iatrogenic ectasia developing after laser-assisted in situ keratomileusis (LASIK) and photo-refractive keratectomy (PRK). At the time, it had been postulated that the combination of ultraviolet light with riboflavin could not only biomechanically strengthen the cornea but also was capable of destroying living cells and organisms including keratocytes and pathogens. Thus a new and innovative concept of treatment for infectious keratitis emerged through the use of CXL technology. Initially only advanced infectious melting ulcers resisting standard microbicidal therapy were treated with CXL in addition to standard therapy. In subsequent studies CXL was also used to treat bacterial keratitis as first line therapy without the use of concomitant antibiotic therapy. With the increasing interest in CXL technology to treat infectious keratitis and to clearly separate its use from the treatment of ectatic disorders, a new term was adopted at the 9(th) CXL congress in Dublin for this specific indication: PACK-CXL (photoactivated chromophore for infectious keratitis). PACK-CXL has the potential to eventually become an interesting alternative to standard antibiotic therapy in treating infectious corneal disorders, and may help reduce the global burden of microbial resistance to antibiotics and other therapeutic agents.

  14. A Multi-Center, Cross-Sectional Study on the Burden of Infectious Keratitis in China

    PubMed Central

    Song, Xiusheng; Xie, Lixin; Tan, Xiaodong; Wang, Zhichong; Yang, Yanning; Yuan, Yuansheng; Deng, Yingping; Fu, Shaoying; Xu, Jianjiang; Sun, Xuguang; Sheng, Xunlun; Wang, Qing

    2014-01-01

    Objective To understand the prevalence and demographic characteristics of infectious keratitis and infectious corneal blindness. Methods A multi-center, population-based cross-sectional study was conducted from January 1 to August 31, 2010. A total of 191,242 individuals of all age groups from 10 geographically representative provinces were sampled using stratified, multi-stage, random and systematic sampling procedures. A majority, 168,673 (88.2%), of those sampled participated in the study. The examination protocol included a structured interview, visual acuity testing, an external eye examination, and an anterior segment examination using a slit lamp. The causes and sequelae of corneal disease were identified using uniform customized protocols. Blindness in one eye caused by infectious keratitis was defined as infectious corneal blindness. Results The prevalence of past and active infectious keratitis was 0.192% (95% confidence interval [CI], 0.171–0.213%), and the prevalence of viral, bacterial, and fungal keratitis was 0.11%, 0.075%, and 0.007%, respectively. There were 138 cases of infectious corneal blindness in at least one eye in the study population (prevalence of 0.082% [95%CI, 0.068%–0.095%]). Statistical analysis suggested that ocular trauma, alcoholic consumption, low socioeconomic levels, advanced age, and poor education were risk factors for infectious corneal blindness. Conclusions Infectious keratitis is the leading cause of corneal blindness in China. Eye care strategies should focus on the prevention and rehabilitation of infectious corneal blindness. PMID:25438169

  15. Synthetic β-sheet forming peptide amphiphiles for treatment of fungal keratitis.

    PubMed

    Wu, Hong; Ong, Zhan Yuin; Liu, Shaoqiong; Li, Yan; Wiradharma, Nikken; Yang, Yi Yan; Ying, Jackie Y

    2015-03-01

    Fungal keratitis is a leading cause of ocular morbidity. It is frequently misdiagnosed as bacterial keratitis, causing a delay in proper treatment. Furthermore, due to the lack of safe and effective anti-fungal agents for clinical use, treatment of fugal keratitis remains a challenge. In recent years, antimicrobial peptides (AMPs) have received considerable attention as potent and broad-spectrum antimicrobial agents with the potential to overcome antibiotics resistance. We previously reported the design of short synthetic β-sheet forming peptides (IKIK)2-NH2 and (IRIK)2-NH2 with excellent antimicrobial activities and selectivities against various clinically relevant microorganisms, including Gram-positive Staphylococcus epidermidis and Staphylococcus aureus, Gram-negative Escherichia coli and Pseudomonas aeruginosa, and yeast Candida albicans (C. albicans). In this study, we evaluated the application of the two most promising synthetic β-sheet forming peptide candidates for in vivo fungal keratitis treatment in comparison with the commercially available amphotericin B. It was found that topical solutions of the designed peptides are safe, and as effective as the clinically used amphotericin B. Compared to the costly and unstable amphotericin B, (IKIK)2-NH2 and (IRIK)2-NH2 are water-soluble, less expensive and stable. Thus, the synthetic β-sheet forming peptides are presented as promising candidates for the treatment of fungal keratitis.

  16. Synthetic β-sheet forming peptide amphiphiles for treatment of fungal keratitis.

    PubMed

    Wu, Hong; Ong, Zhan Yuin; Liu, Shaoqiong; Li, Yan; Wiradharma, Nikken; Yang, Yi Yan; Ying, Jackie Y

    2015-03-01

    Fungal keratitis is a leading cause of ocular morbidity. It is frequently misdiagnosed as bacterial keratitis, causing a delay in proper treatment. Furthermore, due to the lack of safe and effective anti-fungal agents for clinical use, treatment of fugal keratitis remains a challenge. In recent years, antimicrobial peptides (AMPs) have received considerable attention as potent and broad-spectrum antimicrobial agents with the potential to overcome antibiotics resistance. We previously reported the design of short synthetic β-sheet forming peptides (IKIK)2-NH2 and (IRIK)2-NH2 with excellent antimicrobial activities and selectivities against various clinically relevant microorganisms, including Gram-positive Staphylococcus epidermidis and Staphylococcus aureus, Gram-negative Escherichia coli and Pseudomonas aeruginosa, and yeast Candida albicans (C. albicans). In this study, we evaluated the application of the two most promising synthetic β-sheet forming peptide candidates for in vivo fungal keratitis treatment in comparison with the commercially available amphotericin B. It was found that topical solutions of the designed peptides are safe, and as effective as the clinically used amphotericin B. Compared to the costly and unstable amphotericin B, (IKIK)2-NH2 and (IRIK)2-NH2 are water-soluble, less expensive and stable. Thus, the synthetic β-sheet forming peptides are presented as promising candidates for the treatment of fungal keratitis. PMID:25591960

  17. PACK-CXL: Corneal Cross-linking for Treatment of Infectious Keratitis

    PubMed Central

    Tabibian, David; Richoz, Olivier; Hafezi, Farhad

    2015-01-01

    This article discusses corneal cross-linking (CXL) and how it transitioned from a modality for treating corneal ectatic disorders to an inventive means of treating infectious keratitis. Initially, CXL was successfully developed to halt the progression of ectatic diseases such as keratoconus, using the standard Dresden protocol. Later, indications were extended to treat iatrogenic ectasia developing after laser-assisted in situ keratomileusis (LASIK) and photo-refractive keratectomy (PRK). At the time, it had been postulated that the combination of ultraviolet light with riboflavin could not only biomechanically strengthen the cornea but also was capable of destroying living cells and organisms including keratocytes and pathogens. Thus a new and innovative concept of treatment for infectious keratitis emerged through the use of CXL technology. Initially only advanced infectious melting ulcers resisting standard microbicidal therapy were treated with CXL in addition to standard therapy. In subsequent studies CXL was also used to treat bacterial keratitis as first line therapy without the use of concomitant antibiotic therapy. With the increasing interest in CXL technology to treat infectious keratitis and to clearly separate its use from the treatment of ectatic disorders, a new term was adopted at the 9th CXL congress in Dublin for this specific indication: PACK-CXL (photoactivated chromophore for infectious keratitis). PACK-CXL has the potential to eventually become an interesting alternative to standard antibiotic therapy in treating infectious corneal disorders, and may help reduce the global burden of microbial resistance to antibiotics and other therapeutic agents. PMID:26005557

  18. [Use of the presence of cellulose in cellular wall of Acanthamoeba cysts for diagnostic purposes].

    PubMed

    Derda, Monika; Hadaś, Edward

    2009-01-01

    Species identification within the genus Acanthamoeba is based predominantly on morphological and biochemical features. It is labor-intensive and requires cloning and axenization. We described a novel immunocytochemical method for the identification of Acanthamoeba spp. based on selective binding of Clostridium cellulovorans cellulase to protozoan cyst wall cellulose. Free-living amoebae isolated from different water sources by filtration and subsequent cultivation on non-nutrient agar were assigned to genera Acanthamoeba, Naegleria or Hartmannella using morphological taxonomic criteria. Tissues samples from experimentally infected mice were fixed in formalin and for sectioning embedded in paraffin or snap frozen. The Cellulose-Binding Domain of C. cellulovorans cellulase (CBD) obtained as a recombinant protein, were coupled to the fluorescent dye using Alexa Fluor350, 488, 568 - Protein Labelling Kit or labelled with the biotin using EZ-Link Sulfo-NHS-Biotin. All coupling procedures were performed according to the methods provided by manufacturers. For staining with CBD conjugate, slides containing cysts collected from the agar plates or tissue sections were immersed with PBS and incubated with CBD for 30 min at room temperature, washed 3 times with PBS. For staining with CBD-biotin slides containing cysts were incubated with biotinylated CBD for 30 min at room temperature. Subsequent washings in changes of PBS were followed by the incubation with Strept ABComplex/HRP, for 30 min at room temperature, than 3,3 diaminobenzidine tetrahydrochloride was added for 15 min. Slides were rinsed with water, dried and examined in the light microscope. We showed that cellulose could be easily detected by immunofluorescence using conjugated CBD in the inner cyst wall of Acanthamoeba spp. The reference strains of Acanthamoeba spp. and all Acanthamoeba strains isolated from water and from tissues of infected animals gave positive reaction. CBD prepared as a biotynylated protein

  19. Robo 4 Counteracts Angiogenesis in Herpetic Stromal Keratitis

    PubMed Central

    Gimenez, Fernanda; Bhela, Siddheshvar; Rouse, Barry T.

    2015-01-01

    The cornea is a complex tissue that must preserve its transparency to maintain optimal vision. However, in some circumstances, damage to the eye can result in neovascularization that impairs vision. This outcome can occur when herpes simplex virus type 1 (HSV-1) causes the immunoinflammatory lesion stromal keratitis (SK). Potentially useful measures to control the severity of SK are to target angiogenesis which with herpetic SK invariably involves VEGF. One such way to control angiogenesis involves the endothelial receptor Robo4 (R4), which upon interaction with another protein activates an antiangiogenic pathway that counteracts VEGF downstream signaling. In this study we show that mice unable to produce R4 because of gene knockout developed significantly higher angiogenesis after HSV-1 ocular infection than did infected wild type (WT) controls. Moreover, providing additional soluble R4 (sR4) protein by subconjunctival administration to R4 KO HSV-1 infected mice substantially rescued the WT phenotype. Finally, administration of sR4 to WT HSV-1 infected mice diminished the extent of corneal angiogenesis compared to WT control animals. Our results indicate that sR4 could represent a useful therapeutic tool to counteract corneal angiogenesis and help control the severity of SK. PMID:26720197

  20. Congenital Corneal Anesthesia and Neurotrophic Keratitis: Diagnosis and Management

    PubMed Central

    Mantelli, Flavio; Nardella, Chiara; Tiberi, Eloisa; Sacchetti, Marta; Bruscolini, Alice; Lambiase, Alessandro

    2015-01-01

    Neurotrophic keratitis (NK) is a rare degenerative disease of the cornea caused by an impairment of corneal sensory innervation, characterized by decreased or absent corneal sensitivity resulting in epithelial keratopathy, ulceration, and perforation. The aetiopathogenesis of corneal sensory innervation impairment in children recognizes the same range of causes as adults, although they are much less frequent in the pediatric population. Some extremely rare congenital diseases could be considered in the aetiopathogenesis of NK in children. Congenital corneal anesthesia is an extremely rare condition that carries considerable diagnostic and therapeutic problems. Typically the onset is up to 3 years of age and the cornea may be affected in isolation or the sensory deficit may exist as a component of a congenital syndrome, or it may be associated with systemic somatic anomalies. Accurate diagnosis and recognition of risk factors is important for lessening long-term sequelae of this condition. Treatment should include frequent topical lubrication and bandage corneal or scleral contact lenses. Surgery may be needed in refractory cases. The purpose of this review is to summarize and update data available on congenital causes and treatment of corneal hypo/anesthesia and, in turn, on congenital NK. PMID:26451380

  1. Successful treatment of Fusarium keratitis after photo refractive keratectomy

    PubMed Central

    Cavallini, Gian Maria; Ducange, Pietro; Volante, Veronica; Benatti, Caterina

    2013-01-01

    A 39-year-old woman presented to our hospital with a history of photorefractive keratectomy (PRK), performed two weeks prior; slit-lamp examination revealed diffuse conjunctival congestion, corneal ulcer and stromal infiltration. After 5 days of antifungal and antibacteric treatment, the infiltrate progressively increased so that a therapeutic penetrating keratoplasty was necessary. The microbiological analyses revealed the presence of fungal filaments. Twenty days after surgery the patient had recurrent fungal infiltrate in the donor cornea with wound dehiscence. We performed a second penetrating keratoplasty. With the matrix-assisted-laser-desorption-ionization-time-of-flight analysis (MALDI-TOF) we identified a Fusarium solani. Intravenous amphothericine B, a combination of intracameral and intrastromal voriconazole and intracameral amphotericine B were administered. After 6 months from the last surgery the infection was eradicated. The management of fungal keratitis after PRK depends on many factors: In our experience, a prompt keratoplasty and the use of intracameral antifungal medication proved to be very effective. PMID:24178402

  2. Clinical and corneal microbial profile of infectious keratitis in a high HIV prevalence setting in rural South Africa.

    PubMed

    Schaftenaar, E; Peters, R P H; Baarsma, G S; Meenken, C; Khosa, N S; Getu, S; McIntyre, J A; Osterhaus, A D M E; Verjans, G M G M

    2016-09-01

    The purpose of this investigation was to determine the clinical and corneal microbial profile of infectious keratitis in a high human immunodeficiency virus (HIV) prevalence setting in rural South Africa. Data in this cross-sectional study were collected from patients presenting with symptoms of infectious keratitis (n = 46) at the ophthalmology outpatient department of three hospitals in rural South Africa. Corneal swabs were tested for herpes simplex virus type 1 (HSV-1) and 2 (HSV-2), varicella zoster virus (VZV) and adenovirus DNA by real-time polymerase chain reaction (PCR) and for bacteria and fungi by culture. Based on clinical history, disease characteristics and laboratory results, 29 (63 %) patients were diagnosed as viral keratitis, including 14 (48 %) viral keratitis cases complicated by bacterial superinfection, and 17 (37 %) as bacterial keratitis. VZV and HSV-1 DNA was detected in 11 (24 %) and 5 (11 %) corneal swabs, respectively. Among clinically defined viral keratitis cases, a negative viral swab was predominantly (93 %) observed in cases with subepithelial inflammation and was significantly associated with an increased duration of symptoms (p = 0.003). The majority of bacteria cultured were Gram-positive (24/35), including Staphylococcus epidermidis and S. aureus. Viral aetiology was significantly associated with a history of herpes zoster ophthalmicus (p < 0.001) and a trend was observed between viral aetiology and HIV infection (p = 0.06). Twenty-one (47 %) keratitis cases were complicated by anterior uveitis, of which 18 (86 %) were HIV-infected cases with viral keratitis. The data implicate a high prevalence of herpetic keratitis, in part complicated by bacterial superinfection and/or uveitis, in HIV-infected individuals presenting with infectious keratitis in rural South Africa. PMID:27236644

  3. Fungal interface keratitis by Candida orthopsilosis following deep anterior lamellar keratoplasty

    PubMed Central

    Wessel, Julia M; Bachmann, Björn O; Meiller, Ralph; Kruse, Friedrich E

    2013-01-01

    A 39-year-old male patient underwent uncomplicated deep anterior lamellar keratoplasty due to keratoconus. On day 5 after surgery, small whitish infiltrates developed in the corneal interface. The diagnosis of fungal keratitis was made when the culture medium of the graft grew Candida after the surgical intervention. Despite intensive antimycotic treatment and irrigation of the interface, the infiltrates persisted and eventually enlarged. Therefore, revision surgery with penetrating keratoplasty was performed. Microbiological analysis showed Candida orthopsilosis in the culture of the excised graft button. Histopathological staining of the excised graft showed periodic acid–Schiff-positive and Grocott methenamine silver-positive clusters of yeast between Descemet's membrane and the deep corneal stroma with focal perforations through Descemet's membrane. The treatment of mycotic keratitis caused by C orthopsilosis is challenging. Antimycotic treatment was unsuccessful in this case. Progression of the keratitis and perforation of Descemet's membrane suggest that early surgical intervention by penetrating keratoplasty is required. PMID:23349184

  4. Fungal Keratitis Due to Beauveria bassiana in a Contact Lenses Wearer and Review of Published Reports.

    PubMed

    Lara Oya, Ana; Medialdea Hurtado, María Eloisa; Rojo Martín, María Dolores; Aguilera Pérez, Antonia; Alastruey-Izquierdo, Ana; Miranda Casas, Consuelo; Rubio Prats, Marina; Medialdea Marcos, Santiago; Navarro Marí, José María

    2016-10-01

    Fungal keratitis is a severe ocular infection that primarily affects subjects engaged in outdoor activities. Risk factors include allergic conjunctivitis, previous eye surgery, previous treatment with wide-spectrum antimicrobial agents and corticosteroids and using contact lenses. Corneal infection is usually secondary to trauma involving organic material, which is often the only predisposing factor. Early diagnosis based on clinical examination and microbiological investigation (microscopy, cultures and molecular techniques) is crucial to selecting the appropriate antifungal therapy and prevent progression. We report the case of a patient with keratitis due to Beauveria bassiana, an opportunistic and entomopathogenic filamentous fungus that is used as a biological insecticide and which is a rare cause of corneal infection. We review previous cases reports of B. bassiana keratitis published and its main features to compare with our case, a female occasional agriculture worker who had not suffered any trauma involving organic material. The patient received topical and oral antifungal therapy and debridement surgery, with a satisfactory outcome.

  5. Methicillin-resistant Staphylococcus aureus ulcerative keratitis in a Thoroughbred racehorse

    PubMed Central

    KURODA, Taisuke; KINOSHITA, Yuta; NIWA, Hidekazu; MIZOBE, Fumiaki; UENO, Takanori; KUWANO, Atsutoshi; HATAZOE, Takashi; HOBO, Seiji

    2015-01-01

    ABSTRACT We report the first case of methicillin-resistant Staphylococcus aureus (MRSA) keratitis in a racehorse. A 5-year-old mare developed punctate keratitis after racing. The corneal ulcer continued to expand despite ophthalmic antimicrobial therapy. On day 6, a conjunctival graft surgery was performed. The mare was euthanized, following colitis and laminitis development on day 10. MRSA was isolated from the corneal swab taken at the time of euthanasia. Immunohistochemical analysis demonstrated gram-positive and anti-S. aureus monoclonal antibody-positive cocci infiltration of the corneal stroma; and a diagnosis of MRSA ulcerative keratitis was made. An ophthalmic antimicrobial against the isolated MRSA did not improve the ocular lesion. The MRSA strain was found to be staphylococcal cassette chromosome mec type II, a strain frequently isolated from humans in Japan. PMID:26435683

  6. Methicillin-resistant Staphylococcus aureus ulcerative keratitis in a Thoroughbred racehorse.

    PubMed

    Kuroda, Taisuke; Kinoshita, Yuta; Niwa, Hidekazu; Mizobe, Fumiaki; Ueno, Takanori; Kuwano, Atsutoshi; Hatazoe, Takashi; Hobo, Seiji

    2015-01-01

    We report the first case of methicillin-resistant Staphylococcus aureus (MRSA) keratitis in a racehorse. A 5-year-old mare developed punctate keratitis after racing. The corneal ulcer continued to expand despite ophthalmic antimicrobial therapy. On day 6, a conjunctival graft surgery was performed. The mare was euthanized, following colitis and laminitis development on day 10. MRSA was isolated from the corneal swab taken at the time of euthanasia. Immunohistochemical analysis demonstrated gram-positive and anti-S. aureus monoclonal antibody-positive cocci infiltration of the corneal stroma; and a diagnosis of MRSA ulcerative keratitis was made. An ophthalmic antimicrobial against the isolated MRSA did not improve the ocular lesion. The MRSA strain was found to be staphylococcal cassette chromosome mec type II, a strain frequently isolated from humans in Japan. PMID:26435683

  7. A multisystemic Acanthamoeba infection in a dog in Tenerife, Canary Islands, Spain.

    PubMed

    Valladares, María; Reyes-Batlle, María; Mora-Peces, Inmaculada; Martín-Navarro, Carmen M; López-Arencibia, Atteneri; Dorta-Gorrín, Alexis; Comyn-Afonso, Estefanía; Martínez-Carretero, Enrique; Maciver, Sutherland K; Piñero, José E; Valladares, Basilio; Lorenzo-Morales, Jacob

    2014-10-15

    A 22-month-old male Spanish water dog was hospitalized after its physical examination revealed fever and movement difficulty. After 24h, the dog was found to have a high fever (39.5 °C) and was treated empirically with doxycycline/ciprofloxacin. At 48 h, after submission the fever rose to 41 °C and the animal presented with a stiff neck and dehydration. Peripheral blood and cerebrospinal fluid (CSF) were sampled and trophozoites with an Acanthamoeba-like morphology were observed in the CSF. PCR specific for Acanthamoeba, Naegleria fowleri and Balamuthia mandrillaris were performed and the CSF sample found positive for Acanthamoeba. Lungs, kidney, liver and spleen samples were collected post mortem. All collected organ samples were positive for Acanthamoeba by PCR, thus confirming a multisystemic infection. Water samples taken at a suspected site of infection yielded an almost identical PCR fragment to those of the clinical samples, indicating that this was probably where the infection originated. This is the first report of a fatal case of Acanthamoeba disseminated infection in a dog in Spain. PMID:25193180

  8. Detection and quantification of human adenovirus genomes in Acanthamoeba isolated from swimming pools.

    PubMed

    Staggemeier, Rodrigo; Arantes, Thalita; Caumo, Karin S; Rott, Marilise B; Spilki, Fernando R

    2016-01-01

    Acanthamoeba is the most common free-living environmental amoeba, it may serve as an important vehicle for various microorganisms living in the same environment, such as viruses, being pathogenic to humans. This study aimed to detect and quantify human adenoviruses (HAdV) in Acanthamoebas isolated from water samples collected from swimming pools in the city of Porto Alegre, Southern Brazil. Free-living amoebae of the genus Acanthamoeba were isolated from water samples, and isolates (n=16) were used to investigate the occurrence of HAdVs. HAdV detection was performed by quantitative real-time polymerase chain reaction (qPCR). HAdVs were detected in 62.5% (10/16) of Acanthamoeba isolates, ranging from 3.24x103 to 5.14x105 DNA copies per milliliter of isolate. HAdV viral loads found in this study are not negligible, especially because HAdV infections are associated with several human diseases, including gastroenteritis, respiratory distress, and ocular diseases. These findings reinforce the concept that Acanthamoeba may act as a reservoir and promote HAdV transmission through water. PMID:27142544

  9. Chloroquine Has a Cytotoxic Effect on Acanthamoeba Encystation through Modulation of Autophagy

    PubMed Central

    Jha, Bijay Kumar; Jung, Hui-Jung; Seo, Incheol; Kim, Hyun Ah; Suh, Seong-Il; Suh, Min-Ho

    2014-01-01

    Encystation of Acanthamoeba castellanii is associated with resistance to chemotherapeutic agents. Blocking the encystation process could potentiate the efficacy of chemotherapeutic agents and biocides. During encystation, autophagy is highly stimulated and required for proper encystation of Acanthamoeba. In this study, the cytotoxic effect of chloroquine, a well-known autophagy-inhibitory drug, was tested in A. castellanii. Chloroquine was able to selectively reduce cell survival during the encystation of A. castellanii. However, A. castellanii trophozoites and mature cysts were resistant to chloroquine. Chloroquine treatment led to an increase in the number and size of lysosomes in encysting cells. Moreover, chloroquine inhibited the degradation of long-lived proteins in the encysting cells. Decreased autophagic flux, indicated by an increased number of lysosomes and decreased degradation of long-lived proteins, may be the mechanism by which cell death is induced by chloroquine in encysting Acanthamoeba. These results suggest a potential novel therapeutic application of chloroquine as an anti-Acanthamoeba drug. Our findings also suggest that targeting autophagy could be a therapeutic strategy against Acanthamoeba infection. PMID:25114131

  10. Chloroquine has a cytotoxic effect on Acanthamoeba encystation through modulation of autophagy.

    PubMed

    Jha, Bijay Kumar; Jung, Hui-Jung; Seo, Incheol; Kim, Hyun Ah; Suh, Seong-Il; Suh, Min-Ho; Baek, Won-Ki

    2014-10-01

    Encystation of Acanthamoeba castellanii is associated with resistance to chemotherapeutic agents. Blocking the encystation process could potentiate the efficacy of chemotherapeutic agents and biocides. During encystation, autophagy is highly stimulated and required for proper encystation of Acanthamoeba. In this study, the cytotoxic effect of chloroquine, a well-known autophagy-inhibitory drug, was tested in A. castellanii. Chloroquine was able to selectively reduce cell survival during the encystation of A. castellanii. However, A. castellanii trophozoites and mature cysts were resistant to chloroquine. Chloroquine treatment led to an increase in the number and size of lysosomes in encysting cells. Moreover, chloroquine inhibited the degradation of long-lived proteins in the encysting cells. Decreased autophagic flux, indicated by an increased number of lysosomes and decreased degradation of long-lived proteins, may be the mechanism by which cell death is induced by chloroquine in encysting Acanthamoeba. These results suggest a potential novel therapeutic application of chloroquine as an anti-Acanthamoeba drug. Our findings also suggest that targeting autophagy could be a therapeutic strategy against Acanthamoeba infection. PMID:25114131

  11. Trichophyton Spp. fungal keratitis in 22 years old female contact lenses wearer.

    PubMed

    Mravicić, Ivana; Dekaris, Iva; Gabrić, Nikica; Romac, Ivana; Glavota, Vlade; Sviben, Mario

    2010-04-01

    Fungal keratitis represents one of the most difficult forms of microbial keratitis to diagnose and treat successfully. It is difficult to obtain correct diagnosis and topical antifungal preparations. Fungi can cause severe stromal necrosis and enter the anterior chamber by penetrating an intact Descemet membrane. The most common pathogens are filamentous fungi (Aspergillus and Fusarium spp.) and Candida albicans. The incidence of Trichophyton spp. keratitis is 5%. A 22 years old female contact lenses wearer after keratitis developed corneal melting syndrome, spontaneous perforation of the cornea and complicated cataract of the left eye. Conjunctival swab was sterile as well as first sample of corneal tissue and sample from the anterior chamber. Urgent therapeutic perforating keratoplasty (PK), was performed together with extracapsular cataract extraction and the implantation of the intraocular lens in the posterior chamber. The patient was treated with ciprofloxacin and diflucan (systemic therapy); with dexamethason and atropin (subconjunctivaly) and chlorhexidine, brolene, levofloxacin, polimyxin B, and dexamethason/neomycin (topically). Microbiology evaluation was performed once again following excisional biopsy of the intracameral portion of the lesion. The presence of Trichophyton spp. was finally confirmed. Itraconazole and garamycin were included in the systemic therapy. Corneal graft was clear for 17 days but decompensated 28 days after the PK. After two weeks microorganisms invaded the vitreous and caused endophthalmitis. Despite urgent pars plana vitrectomy patient developed endophthalmitis, lost light sensation and developed phthysis. Evisceration and the implantation of silicon prosthesis was done. Perforating keratoplasty is a method of choice in treating severe infectious keratitis unresponsive to conservative treatment but without the eradication of microorganisms it cannot restore the vision or save the eye. Trichophyton spp. may cause a severe

  12. 46 CFR 7.150 - Canadian (BC) and United States (AK) Borders to Cape Spencer, AK.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 1 2014-10-01 2014-10-01 false Canadian (BC) and United States (AK) Borders to Cape Spencer, AK. 7.150 Section 7.150 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY PROCEDURES APPLICABLE TO THE PUBLIC BOUNDARY LINES Alaska § 7.150 Canadian (BC) and United States (AK) Borders to...

  13. 46 CFR 7.150 - Canadian (BC) and United States (AK) Borders to Cape Spencer, AK.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 1 2011-10-01 2011-10-01 false Canadian (BC) and United States (AK) Borders to Cape Spencer, AK. 7.150 Section 7.150 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY PROCEDURES APPLICABLE TO THE PUBLIC BOUNDARY LINES Alaska § 7.150 Canadian (BC) and United States (AK) Borders to...

  14. 46 CFR 7.150 - Canadian (BC) and United States (AK) Borders to Cape Spencer, AK.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 1 2012-10-01 2012-10-01 false Canadian (BC) and United States (AK) Borders to Cape Spencer, AK. 7.150 Section 7.150 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY PROCEDURES APPLICABLE TO THE PUBLIC BOUNDARY LINES Alaska § 7.150 Canadian (BC) and United States (AK) Borders to...

  15. 46 CFR 7.150 - Canadian (BC) and United States (AK) Borders to Cape Spencer, AK.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 1 2013-10-01 2013-10-01 false Canadian (BC) and United States (AK) Borders to Cape Spencer, AK. 7.150 Section 7.150 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY PROCEDURES APPLICABLE TO THE PUBLIC BOUNDARY LINES Alaska § 7.150 Canadian (BC) and United States (AK) Borders to...

  16. 46 CFR 7.150 - Canadian (BC) and United States (AK) Borders to Cape Spencer, AK.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 1 2010-10-01 2010-10-01 false Canadian (BC) and United States (AK) Borders to Cape Spencer, AK. 7.150 Section 7.150 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY PROCEDURES APPLICABLE TO THE PUBLIC BOUNDARY LINES Alaska § 7.150 Canadian (BC) and United States (AK) Borders to...

  17. Isolation and identification of Acanthamoeba species from natural water sources in the northeastern part of Thailand.

    PubMed

    Thammaratana, Thani; Laummaunwai, Porntip; Boonmars, Thidarut

    2016-04-01

    Acanthamoeba are found in the environment, particularly in water, all over the world. The genus is currently classified into 20 different genotypes, T1-T20. In this study, 63 natural water samples from 11 provinces in northeast Thailand were collected and cultured on non-nutrient agar plates. Positive samples by culture were subsequently analyzed by molecular methods. The identification of Acanthamoeba was based on morphological features and molecular techniques using PCR and DNA sequencing. The results showed that 10 samples out of 63 were positive (15.9 %). Phylogenetic analysis revealed that seven samples were T4, one sample was similar to T3, and the other two samples were similar to T5. This is the first report demonstrating the contamination of Acanthamoeba species in natural water sources in northeast Thailand. PMID:26779920

  18. Internalization of Mycobacterium shottsii and Mycobacterium pseudoshottsii by Acanthamoeba polyphaga.

    PubMed

    Gupta, Tuhina; Fine-Coulson, Kari; Karls, Russell; Gauthier, David; Quinn, Frederick

    2013-08-01

    Amoebae serve as environmental hosts to a variety of mycobacteria, including Mycobacterium avium and Mycobacterium marinum. Mycobacterium shottsii and Mycobacterium pseudoshottsii are waterborne species isolated from the spleens and dermal lesions of striped bass (Morone saxatilis) from the Chesapeake Bay. The optimal growth temperature for these fish isolates is 25 °C. In the present study, amoebae were examined as a potential environmental reservoir for these fish pathogens. Several studies demonstrated that M. avium bacilli replicate within the trophozoite stage and reside in large numbers within the cytosol of the cyst of the free-living amoeba Acanthamoeba polyphaga. Results from the present study showed that M. shottsii, M. pseudoshottsii, and M. marinum bacilli were internalized by A. polyphaga trophozoites within 6 h but that intracellular viability decreased by 2 to 3 logs over 10 days. While an average of 25 M. marinum bacilli were identified by electron microscopy in the cytosol of the cyst, <5 M. pseudoshottsii and no M. shottsii bacilli were observed in this location. All Mycobacterium species examined remained viable but did not replicate after encystment and subsequent 48 h incubation in 4% HCl. This concentration of HCl will kill mycobacteria but will not enter amoebal cysts. Bacterial viability studies within stages of the amoeba life cycle indicate fewer M. shottsii and M. pseudoshottsii bacilli within the trophozoite and cyst stages relative to M. marinum. PMID:23899000

  19. Shiga toxins decrease enterohaemorrhagic Escherichia coli survival within Acanthamoeba castellanii.

    PubMed

    Chekabab, Samuel M; Daigle, France; Charette, Steve J; Dozois, Charles M; Harel, Josée

    2013-07-01

    Enterohaemorrhagic Escherichia coli (EHEC) are zoonotic pathogens transmitted to humans through contaminated water or bovine products. One of the strategies used by pathogenic bacteria to survive in aquatic environments is using free-living amoebae as hosts. Acanthamoeba castellanii is an amoeba known to host several waterborne pathogens. This study investigates the survival of EHEC with A. castellanii, which could contribute to its spread and transmission to humans. We used a gentamicin protection assay as well as fluorescence and electron microscopy to monitor the intra-amoebae survival of EHEC O157:H7 over 24 h. The results showed that EHEC were able to survive within A. castellanii and that this survival was reduced by Shiga toxins (Stx) produced by EHEC. A toxic effect mediated by Stx was demonstrated by amoebae mortality and LDH release during co-culture of EHEC and amoeba. This work describes the ability of EHEC to survive within A. castellanii, and this host-pathogen interaction is partially controlled by the Stx. Thus, this ubiquitous amoeba could represent an environmental niche for EHEC survival and transmission. PMID:23581502

  20. Photoinhibition of Growth in Acanthamoeba castellanii Cultures1

    PubMed Central

    Dolphin, Warren D.

    1970-01-01

    Light from 350 to 680 nm at intensities up to 1.62 × 105 ergs per sec per cm2 slowed exponential growth and lowered the maximum yield in axenic cultures of Acanthamoeba castellanii. Photoinhibition was a linear function of light intensity up to 1.25 × 105 ergs per sec per cm2. At higher intensities, growth was too slow to be measured accurately. A photochemical change occurring in the growth medium on irradiation was a function of light dosage and not intensity per se. Light in dosages which appreciably changed the growth-supporting properties of the medium exceeded the dosages received by exponentially growing cultures during irradiation. Consequently, photoinhibition of growth was attributed to a direct effect of light on the amoebae, not to photodegradation of the medium. The growth-supporting properties of irradiated media could be restored by the addition of yeast extract and Proteose peptone. The reduced growth rate in the light was not due to cyst formation or induction of multinuclearity. Light affected the amoebae either through absorption by intracellular pigment(s) or through binding to the amoebae of a photosensitizing compound in the medium. PMID:5474886

  1. Ultrastructural study of encystation and excystation in Acanthamoeba castellanii.

    PubMed

    Chávez-Munguía, Bibiana; Omaña-Molina, Maritza; González-Lázaro, Mónica; González-Robles, Arturo; Bonilla, Patricia; Martínez-Palomo, Adolfo

    2005-01-01

    Encystation and excystation of Acanthamoeba castellanii were studied by transmission electron microscopy. The differentiation process was induced in asynchronous cultures grown axenically. Cytoplasmic vesicles containing a dense fibrous material very similar in appearance to the cyst wall were observed in trophozoites induced to encyst. When these trophozoites were incubated with calcofluor white m2r, fluorescence was observed in cytoplasmic vesicles, suggesting that the material contained in these vesicles corresponded to cyst wall precursors. Semithin cryosections of mature cysts with the same treatment showed fluorescence in the ectocyst and a less intense fluorescence in the endocyst, suggesting the presence of cellulose in both structures of the cyst wall. In mature cysts induced to excystation, small structures very similar to electron-dense granules (EDG) previously described in other amoebae were frequently observed. The EDGs were either sparsely distributed in the cytoplasm or associated with the cytoplasmic face of the plasma membrane. Many of them were located near the ostiole. In advanced phases of excystation, endocytic activity was suggested by the formation of endocytic structures and the presence of vacuoles with fibrous content similar to that of the cyst wall. Electron-dense granules in the process of dissolution were also observed in these vacuoles. Furthermore, the formation of a pseudopod suggests a displacement of the amoeba toward the ostiole. PMID:15817120

  2. Acanthamoeba royreba: morphological features and in vitro cytopathic effect.

    PubMed

    González-Robles, Arturo; Salazar-Villatoro, Lizbeth; Omaña-Molina, Maritza; Lorenzo-Morales, Jacob; Martínez-Palomo, Adolfo

    2013-04-01

    Observations on cultured Acanthamoeba royreba trophozoites and in vitro cytopathogenicity of this amoeba are described. In culture, amoebae were active, pleomorphic and moved on the substrate by producing endocytic structures and emitting slight cytoplasmic microprojections from the cell surface. These projections were formed by hyaline cytoplasm and they were related to motion structures such as acanthopodia and lamellipodia, in which actin provides a framework that allows rapid changes in morphology. In the cytoplasm abundant vacuoles of different size and content were seen. By means of electron microscopy, it was possible to observe the compact fibrogranular appearance of the cytoplasm, along with the main cellular organelles such as the Golgi complex, the endoplasmic reticulum, digestive vacuoles, mitochondria and contractile vacuoles. Incubation of MDCK epithelial cell monolayers with conditioned medium did not produce a significant structural damage to the monolayer, even after 24h of incubation. When the trophozoites were incubated with the target cells the monolayer exhibited a clear injury created by the amoebae, which produced focal damage. Nevertheless, the rest of the monolayer appeared to remain intact, suggesting that a contact-dependent interaction is necessary to damage the target cells. These observations demonstrate the low invasive capacity of this amoeba. PMID:23357648

  3. A comparative, retrospective, observational study of the clinical and microbiological profiles of post-penetrating keratoplasty keratitis

    NASA Astrophysics Data System (ADS)

    Lin, I.-Huang; Chang, Yi-Sheng; Tseng, Sung-Huei; Huang, Yi-Hsun

    2016-09-01

    Infectious keratitis after penetrating keratoplasty (PK) is a devastating condition that may result in graft failure and poor visual outcome. In this study, we retrospectively reviewed the medical records of patients who underwent PK between 2009 and 2014, and recorded those who developed infectious keratitis. We compared the predisposing factors and organisms isolated to those identified in our previous study, conducted between 1989 and 1994. The incidence of post-PK infectious keratitis decreased from 11.6% (41 out of 354 cases, 1989–1994) to 6.5% (9 out of 138 cases, 2009–2014). Graft epithelial defect and suture-related problems remained the leading two risk factors of infectious keratitis after PK. Gram-positive and Gram-negative bacterial infection decreased from 58.5% and 46.3% to 11.1% and 22.2%, respectively (P = 0.023 and P = 0.271). In contrast, fungus infection increased from 9.8% to 66.7% (P = 0.001) fungi have become the major pathogen for post-PK infectious keratitis. In conclusion, while the incidence of post-PK infectious keratitis has decreased over time, the number and frequency of fungal infections have significantly increased in the recent study period. Clinicians should be aware of the shifting trend in pathogens involved in post-PK infectious keratitis.

  4. A comparative, retrospective, observational study of the clinical and microbiological profiles of post-penetrating keratoplasty keratitis

    PubMed Central

    Lin, I-Huang; Chang, Yi-Sheng; Tseng, Sung-Huei; Huang, Yi-Hsun

    2016-01-01

    Infectious keratitis after penetrating keratoplasty (PK) is a devastating condition that may result in graft failure and poor visual outcome. In this study, we retrospectively reviewed the medical records of patients who underwent PK between 2009 and 2014, and recorded those who developed infectious keratitis. We compared the predisposing factors and organisms isolated to those identified in our previous study, conducted between 1989 and 1994. The incidence of post-PK infectious keratitis decreased from 11.6% (41 out of 354 cases, 1989–1994) to 6.5% (9 out of 138 cases, 2009–2014). Graft epithelial defect and suture-related problems remained the leading two risk factors of infectious keratitis after PK. Gram-positive and Gram-negative bacterial infection decreased from 58.5% and 46.3% to 11.1% and 22.2%, respectively (P = 0.023 and P = 0.271). In contrast, fungus infection increased from 9.8% to 66.7% (P = 0.001); fungi have become the major pathogen for post-PK infectious keratitis. In conclusion, while the incidence of post-PK infectious keratitis has decreased over time, the number and frequency of fungal infections have significantly increased in the recent study period. Clinicians should be aware of the shifting trend in pathogens involved in post-PK infectious keratitis. PMID:27587283

  5. War of the microbial worlds: who is the beneficiary in Acanthamoeba-bacterial interactions?

    PubMed

    Siddiqui, Ruqaiyyah; Khan, Naveed Ahmed

    2012-04-01

    Acanthamoeba hosts diverse microbial organisms including viruses, bacteria, yeast and protists, some of which are potential human pathogens. The precise nature of this symbiosis is not clear, but it is suggested that such interactions enable pathogenic microbes to survive hostile conditions and lead to their transmission to susceptible hosts to establish infection. In particular, Acanthamoeba-bacteria interactions have gained significant attention by the scientific and the medical community and have led to speculations of employing anti-amoebic approaches in eradicating 'superbugs' from clinical settings. Here, we discuss the nature of these convoluted interactions and the benefit they represent for the symbionts.

  6. Novel Acanthamoeba 18S rRNA gene sequence type from an environmental isolate.

    PubMed

    Magnet, A; Henriques-Gil, N; Galván-Diaz, A L; Izquiedo, F; Fenoy, S; del Aguila, C

    2014-08-01

    The free-living amoebae, Acanthamoeba, can act as opportunistic parasites on a wide range of vertebrates and are becoming a serious threat to human health due to the resistance of their cysts to harsh environmental conditions, disinfectants, some water treatment practices, and their ubiquitous distribution. Subgenus classification based on morphology is being replaced by a classification based on the sequences of the 18S rRNA gene with a total of 18 different genotypes (T1-T18). A new environmental strain of Acanthamoeba isolated from a waste water treatment plant is presented in this study as a candidate for the description of the novel genotype T19 after phylogenetic analysis.

  7. Microphthalmia, late onset keratitis, and iris coloboma/aniridia in a family with a novel PAX6 mutation.

    PubMed

    Xiao, Xueshan; Li, Shiqiang; Zhang, Qingjiong

    2012-06-01

    Autosomal dominant microphthalmia with late-onset keratitis and iris coloboma/aniridia has not been reported before. Here we report a Chinese family with these phenotypes and a novel PAX6 mutation. Microphthalmia, late-onset keratitis, iris coloboma, and nystagmus were present in the proband. His son had microphthalmia, aniridia, foveal hypoplasia, and nystagmus. A novel c.649C>T (p.Arg217X) mutation in PAX6 was detected in the proband and his affected son. This study expands the phenotypic spectrum of PAX6 mutation and enriched our knowledge of the genetic cause for microphthalmia and late-onset keratitis.

  8. Acyclovir-resistant varicella-zoster virus keratitis in an immunocompetent patient.

    PubMed

    Gueudry, J; Boutolleau, D; Gueudin, M; Burrel, S; Miri, A; Bodaghi, B; Muraine, M

    2013-09-01

    Varicella-zoster virus (VZV) resistance to acyclovir (ACV) has only been reported in rare cases of immunocompromised patients. We report the first case of an immunocompetent patient with ACV-resistant VZV keratitis associated with a nucleotide deletion in the VZV thymidine kinase gene, leading to production of a truncated protein.

  9. Topical voriconazole therapy of Purpureocillium lilacinum keratitis that occurred in disposable soft contact lens wearers.

    PubMed

    Todokoro, Daisuke; Yamada, Norihiro; Fukuchi, Mariko; Kishi, Shoji

    2014-10-01

    The objective of this study was to describe 2 cases of keratitis caused by Purpureocillium lilacinum (formerly Paecilomyces lilacinus) that occurred in disposable contact lens users, which were successfully treated with topical voriconazole. Case 1 was a healthy 44-year-old woman, who wore weekly disposable contact lenses and had developed a superficial corneal infection in her right eye. For diagnosis, corneal scraping and molecular identification of the cultured pathogen were performed. A corneal smear revealed the presence of fungi. The pathogen was identified as P. lilacinum by traditional morphological identification of fungal culture, and this identification was confirmed by DNA sequencing of the ribosomal internal transcribed spacer (ITS) sequence. Therapy with topical fluconazole, topical pimaricin (natamycin), and oral itraconazole were ineffective. Topical voriconazole showed a significant effect, and the keratitis was successfully treated. Case 2 was a 43-year-old woman with bilateral recurrent peripheral corneal ulcers by meibomian gland dysfunction, who used therapeutic bandage contact lenses on her left eye. However, a corneal abscess with hypopyon occurred in the eye after 3 months. The microbial smear examination showed the presence of fungi and the fungal culture, and the DNA sequence of ITS region revealed that the causative agent was P. lilacinum. The susceptibility testing against antifungal agents showed that voriconazole was effective. The lesion improved gradually by topical voriconazole. As a conclusion, P. lilacinum keratitis can occur in disposable soft contact lens wearer. Early and accurate detection of the pathogenic organism is essential. Topical voriconazole was effective against P. lilacinum keratitis.

  10. Tyrosinemia without liver or renal damage with plantar and palmar keratosis and keratitis (hypertyrosinemia type II).

    PubMed

    Pelet, B; Antener, I; Faggioni, R; Spahr, A; Gautier, E

    1979-05-01

    A boy of 3 2/12 years of age with Richner-Hanhart syndrome (plantar and palmar keratosis and chronic keratitis) was found to have hypertyrosinemia and to excrete the hydroxyacids derived from tyrosine. A diet poor in phenylalanine and tyrosine cured the skin and corneal lesions. Clinical and biochemical observations are reported.

  11. Pathogen Induced Changes in the Protein Profile of Human Tears from Fusarium Keratitis Patients

    PubMed Central

    Ananthi, Sivagnanam; Venkatesh Prajna, Namperumalsamy; Lalitha, Prajna; Valarnila, Murugesan; Dharmalingam, Kuppamuthu

    2013-01-01

    Fusarium is the major causative agent of fungal infections leading to corneal ulcer (keratitis) in Southern India and other tropical countries. Keratitis caused by Fusarium is a difficult disease to treat unless antifungal therapy is initiated during the early stages of infection. In this study tear proteins were prepared from keratitis patients classified based on the duration of infection. Among the patients recruited, early infection (n = 35), intermediate (n = 20), late (n = 11), samples from five patients in each group were pooled for analysis. Control samples were a pool of samples from 20 patients. Proteins were separated on difference gel electrophoresis (DIGE) and the differentially expressed proteins were quantified using DeCyder software analysis. The following differentially expressed proteins namely alpha-1-antitrypsin, haptoglobin α2 chain, zinc-alpha-2-glycoprotein, apolipoprotein, albumin, haptoglobin precursor - β chain, lactoferrin, lacrimal lipocalin precursor, cystatin SA III precursor, lacritin precursor were identified using mass spectrometry. Variation in the expression level of some of the proteins was confirmed using western blot analysis. This is the first report to show stage specific tear protein profile in fungal keratitis patients. Validation of this data using a much larger sample set could lead to clinical application of these findings. PMID:23308132

  12. Evaluation of Fungal Keratitis using a Newly Developed Computer Program, Optscore, for Grading Digital Corneal Photographs

    PubMed Central

    Toutain-Kidd, Christine M.; Porco, Travis C.; Kidd, Eric M.; Srinivasan, M; Prajna, Namperumalsamy V.; Acharya, Nisha; Lietman, Thomas; Zegans, Michael E.

    2016-01-01

    Purpose To validate computer software developed to assess digital corneal photographs of fungal keratitis in clinical research. Methods A cornea specialist and five medical students (after training) graded on two occasions 100 corneal photographs of patients with fungal keratitis using Optscore software. Variables assessed were lesion area, location, degree of opacity, percentage of the ulcer lying within a central 4mm circle of the cornea. Intraclass correlation coefficients (ICCs) were used to assess intragrader reliability, agreement of the students with the corneal specialist, and the reliability of the group mean of the student raters. The area determined using Optscore was compared to the area estimated from slit lamp and to visual acuity. Results As a group, medical students achieved an ICC greater than 0.9 for five out of the seven assessed variables. Similar levels of consistency were found after analyzing the graders’ individual results compared to the specialist. The area estimated using slit lamp examination was highly correlated with the mean area determined by Optscore, as was the logarithm of the minimum angle of resolution visual acuity at enrollment. Conclusions Non-expert graders using Optscore to assess digital photographs of fungal keratitis are self-consistent, agree with an expert grader both as a group and individually, and measurements of ulcer area obtained from Optscore are highly correlated with measurements of the same patients obtained on clinical examination. These observations support the validity of Optscore for assessing corneal pathology associated with fungal keratitis and make it a promising clinical research tool. PMID:24467559

  13. Keratitis due to the wood saprobic ascomycete, Auerswaldia lignicola (Family Botryosphaeriaceae), in a carpenter in India.

    PubMed

    Ruban, Vasanthakumar Vasantha; Kaliamurthy, Jayaraman; Dineshkumar, Muniyandi; Jesudasan, Christadoss Arul Nelson; Geraldine, Pitchairaj; Thomas, Philip Aloysius

    2013-12-01

    Keratitis due to Auerswaldia lignicola in a 32-year-old Indian male carpenter is described. At presentation, the patient reported persistent pain and tearing (left eye) in spite of topical antimicrobial therapy for more than 3 weeks. Clinically, mycotic keratitis was suspected, and direct microscopy of corneal scrapings stained by lactophenol cotton blue and Gram stains revealed broad septate hyphae. Intensive topical antifungal therapy was then given for 15 days. The keratitis continued to progress, necessitating therapeutic penetrating keratoplasty. Following the keratoplasty, there was rapid reduction in inflammation and gradual quietening of the eye. Brown-black fungal colonies resembling Lasiodiplodia theobromae were isolated from corneal scrape and corneal button (post-surgery) material on Sabouraud glucose-neopeptone agar; however, sporulation did not occur, so the morphological identification could not be confirmed. Sequence analysis of the 18S rRNA region of extracted fungal genomic DNA yielded an identification of A. lignicola Ariyawansa, J.K. Liu & K.D. Hyde; the sequence data have been deposited in GenBank (A. lignicola strain DK/V4, accession number KC866317.1). Medical management of keratitis due to such rarely reported fungal species may be difficult, necessitating surgical procedures. PMID:24158617

  14. Antimicrobial management of presumed microbial keratitis: guidelines for treatment of central and peripheral ulcers

    PubMed Central

    Bennett, H; Hay, J; Kirkness, C; Seal, D; Devonshire, P.

    1998-01-01

    AIMS—To determine the quantitative relation between the major risk factors for microbial keratitis of previous ocular surface disease and contact lens wear and central and peripheral infiltration, often associated with ulceration, in order to establish a rational chemotherapeutic management algorithm.
METHODS—Data from 55 patients were collected over a 10 month period. All cases of presumed microbial keratitis where corneal scrapes had been subjected to microbiological examination were included. Risk factor data and laboratory outcome were recorded. Antimicrobial regimens used to treat each patient were documented.
RESULTS—57 episodes of presumed microbial keratitis were identified from 55 patients, 24 male and 31 female. There were 30 central infiltrates and 27 peripheral infiltrates of which 28 were culture positive (73% of central infiltrates, 22% of peripheral infiltrates). 26 patients had worn contact lenses of whom 12 had culture positive scrapes (9/14 for central infiltrates, 3/12 for peripheral infiltrates). 31 patients had an ocular surface disease of whom five previous herpes simplex virus keratitis patients developed secondary bacterial infection. Anterior chamber activity and an infiltrate size ⩾ 4 mm2 were more common with culture positive central infiltrates than peripheral infiltrates (χ2 test = 11.98, p<0.001).
CONCLUSIONS—Predisposing factors for "presumed" microbial keratitis, either central or peripheral, were: ocular surface disease (26/57 = 45.6%), contact lens wear (26/57 = 45.6%), and previous trauma (5/57 = 8.8%). Larger ulceration (⩾4 mm2) with inflammation was more often associated with positive culture results for central infiltration. None of these four variables (contact lens wear, ocular surface disease, ulcer size, anterior chamber activity) were of intrinsic value in predicting if a peripheral infiltrate would yield identifiable micro-organisms. Successful management of presumed microbial

  15. Microscopic Evaluation, Molecular Identification, Antifungal Susceptibility, and Clinical Outcomes in Fusarium, Aspergillus and, Dematiaceous Keratitis

    PubMed Central

    Gajjar, Devarshi U.; Pal, Anuradha K.; Ghodadra, Bharat K.; Vasavada, Abhay R.

    2013-01-01

    Purpose. Fusarium, Aspergillus, and Dematiaceous are the most common fungal species causing keratitis in tropical countries. Herein we report a prospective study on fungal keratitis caused by these three fungal species. Methodology. A prospective investigation was undertaken to evaluate eyes with presumed fungal keratitis. All the fungal isolates (n = 73) obtained from keratitis infections were identified using morphological and microscopic characters. Molecular identification using sequencing of the ITS region and antifungal susceptibility tests using microdilution method were done. The final clinical outcome was evaluated in terms of the time taken for resolution of keratitis and the final visual outcome. The results were analyzed after segregating the cases into three groups, namely, Fusarium, Aspergillus, and Dematiaceous keratitis. Results. Diagnosis of fungal keratitis was established in 73 (35.9%) cases out of 208 cases. The spectra of fungi isolated were Fusarium spp. (26.6%), Aspergillus spp. (21.6%), and Dematiaceous fungi (11.6%). The sequence of the ITS region could identify the Fusarium and Aspergillus species at the species complex level, and the Dematiaceous isolates were accurately identified. Using antifungal agents such as fluconazole, natamycin, amphotericin B, and itraconazole, the minimum inhibitory concentrations (MICs) for Fusarium spp. were >32 μg/mL, 4–8 μg/mL, 0.5–1 μg/mL, and >32 μg/mL, respectively. Antifungal susceptibility data showed that Curvularia spp. was highly resistant to all the antifungal agents. Overall, natamycin and amphotericin B were found to be the most effective antifungal agents. The comparative clinical outcomes in all cases showed that the healing response in terms of visual acuity of the Dematiaceous group was significantly good when compared with the Fusarium and Aspergillus groups (P < 0.05). The time required for healing in the Fusarium group was statistically significantly less when compared with

  16. Treatment with intrastromal and intracameral voriconazole in 2 eyes with Lasiodiplodia theobromae keratitis: case reports.

    PubMed

    Lekhanont, Kaevalin; Nonpassopon, Manachai; Nimvorapun, Nutthida; Santanirand, Pitak

    2015-02-01

    To report the clinical presentation and the role of intrastromal and intracameral voriconazole injection in the management of rare cases of fungal keratitis caused by Lasiodiplodia theobromae.Two eyes of 2 patients with Lasiodiplodia keratitis unresponsive to topical and oral antifungal medications were included in this study. Diagnosis of Lasiodiplodia keratitis was confirmed by microbiological analysis, including culture-based (case 1 and 2) and DNA sequencing techniques (case 2 only).The first patient presented with multiple satellite lesions and one of these infiltrates spread deeply into the cornea, forming a stromal abscess. Another patient had a large full-thickness corneal infiltrates with several fungal balls in the anterior chamber, requiring a limbus-to-limbus therapeutic penetrating keratoplasty. Despite aggressive topical therapy, the stromal abscess continued to worsen in the first case and recurrent keratitis was observed postoperatively in the second case. Voriconazole 50 μg/0.1 mL was administered intracamerally and intrastromally around the fungal abscess as adjuncts to topical antimycotics in the first case. The second patient who needed therapeutic keratoplasty was treated with an intracameral injection of 50 μg/0.1 mL voriconazole at the end of surgery. Postoperatively, 100 μg/0.1 mL voriconazole was also injected intracamerally after the recurrence of infection was noted in the graft. Reinjections were given 48 hours apart in both cases. After the injections, all corneal and anterior chamber lesions were reduced in size and density and completely resolved within 4 weeks.Intrastromal and intracameral voriconazole injections may offer safe and effective treatment options for L theobromae keratitis. PMID:25674759

  17. Treatment With Intrastromal and Intracameral Voriconazole in 2 Eyes With Lasiodiplodia theobromae Keratitis

    PubMed Central

    Lekhanont, Kaevalin; Nonpassopon, Manachai; Nimvorapun, Nutthida; Santanirand, Pitak

    2015-01-01

    Abstract To report the clinical presentation and the role of intrastromal and intracameral voriconazole injection in the management of rare cases of fungal keratitis caused by Lasiodiplodia theobromae. Two eyes of 2 patients with Lasiodiplodia keratitis unresponsive to topical and oral antifungal medications were included in this study. Diagnosis of Lasiodiplodia keratitis was confirmed by microbiological analysis, including culture-based (case 1 and 2) and DNA sequencing techniques (case 2 only). The first patient presented with multiple satellite lesions and one of these infiltrates spread deeply into the cornea, forming a stromal abscess. Another patient had a large full-thickness corneal infiltrates with several fungal balls in the anterior chamber, requiring a limbus-to-limbus therapeutic penetrating keratoplasty. Despite aggressive topical therapy, the stromal abscess continued to worsen in the first case and recurrent keratitis was observed postoperatively in the second case. Voriconazole 50 μg/0.1 mL was administered intracamerally and intrastromally around the fungal abscess as adjuncts to topical antimycotics in the first case. The second patient who needed therapeutic keratoplasty was treated with an intracameral injection of 50 μg/0.1 mL voriconazole at the end of surgery. Postoperatively, 100 μg/0.1 mL voriconazole was also injected intracamerally after the recurrence of infection was noted in the graft. Reinjections were given 48 hours apart in both cases. After the injections, all corneal and anterior chamber lesions were reduced in size and density and completely resolved within 4 weeks. Intrastromal and intracameral voriconazole injections may offer safe and effective treatment options for L theobromae keratitis. PMID:25674759

  18. Interactions between Human Norovirus Surrogates and Acanthamoeba spp.

    PubMed

    Hsueh, Tun-Yun; Gibson, Kristen E

    2015-06-15

    Human noroviruses (HuNoVs) are the most common cause of food-borne disease outbreaks, as well as virus-related waterborne disease outbreaks in the United States. Here, we hypothesize that common free-living amoebae (FLA)-ubiquitous in the environment, known to interact with pathogens, and frequently isolated from water and fresh produce-could potentially act as reservoirs of HuNoV and facilitate the environmental transmission of HuNoVs. To investigate FLA as reservoirs for HuNoV, the interactions between two Acanthamoeba species, A. castellanii and A. polyphaga, as well as two HuNoV surrogates, murine norovirus type 1 (MNV-1) and feline calicivirus (FCV), were evaluated. The results showed that after 1 h of amoeba-virus incubation at 25°C, 490 and 337 PFU of MNV-1/ml were recovered from A. castellanii and A. polyphaga, respectively, while only few or no FCVs were detected. In addition, prolonged interaction of MNV-1 with amoebae was investigated for a period of 8 days, and MNV-1 was demonstrated to remain stable at around 200 PFU/ml from day 2 to day 8 after virus inoculation in A. castellanii. Moreover, after a complete amoeba life cycle (i.e., encystment and excystment), infectious viruses could still be detected. To determine the location of virus associated with amoebae, immunofluorescence experiments were performed and showed MNV-1 transitioning from the amoeba surface to inside the amoeba over a 24-h period. These results are significant to the understanding of how HuNoVs may interact with other microorganisms in the environment in order to aid in its persistence and survival, as well as potential transmission in water and to vulnerable food products such as fresh produce. PMID:25841006

  19. Interactions between Human Norovirus Surrogates and Acanthamoeba spp.

    PubMed Central

    Hsueh, Tun-Yun

    2015-01-01

    Human noroviruses (HuNoVs) are the most common cause of food-borne disease outbreaks, as well as virus-related waterborne disease outbreaks in the United States. Here, we hypothesize that common free-living amoebae (FLA)—ubiquitous in the environment, known to interact with pathogens, and frequently isolated from water and fresh produce—could potentially act as reservoirs of HuNoV and facilitate the environmental transmission of HuNoVs. To investigate FLA as reservoirs for HuNoV, the interactions between two Acanthamoeba species, A. castellanii and A. polyphaga, as well as two HuNoV surrogates, murine norovirus type 1 (MNV-1) and feline calicivirus (FCV), were evaluated. The results showed that after 1 h of amoeba-virus incubation at 25°C, 490 and 337 PFU of MNV-1/ml were recovered from A. castellanii and A. polyphaga, respectively, while only few or no FCVs were detected. In addition, prolonged interaction of MNV-1 with amoebae was investigated for a period of 8 days, and MNV-1 was demonstrated to remain stable at around 200 PFU/ml from day 2 to day 8 after virus inoculation in A. castellanii. Moreover, after a complete amoeba life cycle (i.e., encystment and excystment), infectious viruses could still be detected. To determine the location of virus associated with amoebae, immunofluorescence experiments were performed and showed MNV-1 transitioning from the amoeba surface to inside the amoeba over a 24-h period. These results are significant to the understanding of how HuNoVs may interact with other microorganisms in the environment in order to aid in its persistence and survival, as well as potential transmission in water and to vulnerable food products such as fresh produce. PMID:25841006

  20. Interactions between Human Norovirus Surrogates and Acanthamoeba spp.

    PubMed

    Hsueh, Tun-Yun; Gibson, Kristen E

    2015-06-15

    Human noroviruses (HuNoVs) are the most common cause of food-borne disease outbreaks, as well as virus-related waterborne disease outbreaks in the United States. Here, we hypothesize that common free-living amoebae (FLA)-ubiquitous in the environment, known to interact with pathogens, and frequently isolated from water and fresh produce-could potentially act as reservoirs of HuNoV and facilitate the environmental transmission of HuNoVs. To investigate FLA as reservoirs for HuNoV, the interactions between two Acanthamoeba species, A. castellanii and A. polyphaga, as well as two HuNoV surrogates, murine norovirus type 1 (MNV-1) and feline calicivirus (FCV), were evaluated. The results showed that after 1 h of amoeba-virus incubation at 25°C, 490 and 337 PFU of MNV-1/ml were recovered from A. castellanii and A. polyphaga, respectively, while only few or no FCVs were detected. In addition, prolonged interaction of MNV-1 with amoebae was investigated for a period of 8 days, and MNV-1 was demonstrated to remain stable at around 200 PFU/ml from day 2 to day 8 after virus inoculation in A. castellanii. Moreover, after a complete amoeba life cycle (i.e., encystment and excystment), infectious viruses could still be detected. To determine the location of virus associated with amoebae, immunofluorescence experiments were performed and showed MNV-1 transitioning from the amoeba surface to inside the amoeba over a 24-h period. These results are significant to the understanding of how HuNoVs may interact with other microorganisms in the environment in order to aid in its persistence and survival, as well as potential transmission in water and to vulnerable food products such as fresh produce.

  1. Impact of free-living amoebae on presence of Parachlamydia acanthamoebae in the hospital environment and its survival in vitro without requirement for amoebae.

    PubMed

    Fukumoto, Tatsuya; Matsuo, Junji; Hayashi, Yasuhiro; Hayashi, Masahiro; Oguri, Satoshi; Nakamura, Shinji; Mizutani, Yoshihiko; Yao, Takashi; Akizawa, Kouzi; Suzuki, Haruki; Shimizu, Chikara; Matsuno, Kazuhiko; Yamaguchi, Hiroyuki

    2010-09-01

    Parachlamydia acanthamoebae is an obligately intracellular bacterium that infects free-living amoebae and is a potential human pathogen in hospital-acquired pneumonia. We examined whether the presence of P. acanthamoebae is related to the presence of Acanthamoeba in an actual hospital environment and assessed the in vitro survival of P. acanthamoebae. Ninety smear samples were collected between November 2007 and March 2008 (trial 1, n = 52) and between October 2008 and February 2009 (trial 2, n = 38) from the floor (dry conditions, n = 56) and sink outlets (moist conditions, n = 34) of a hospital. The prevalences of P. acanthamoebae DNA in the first and second trials were 64.3% and 76%, respectively. The prevalences of Acanthamoeba DNA in the first and second trials were 48% and 63.1%, respectively. A statistical correlation between the prevalence of P. acanthamoebae and that of Acanthamoeba was found (trial 1, P = 0.011; trial 2, P = 0.022), and that correlation increased when samples from just the dry area (floor smear samples, P = 0.002) were analyzed but decreased when samples from a moist area were analyzed (P = 0.273). The in vitro experiment showed that, without Acanthamoeba, P. acanthamoebae could not survive in dry conditions for 3 days at 30 degrees C or 15 days at 15 degrees C. Thus, both organisms were coincidentally found in an actual hospital environment, with the presence of Acanthamoeba having a significant effect on the long-term survival of P. acanthamoebae, suggesting that this potential human pathogen could spread through a hospital environment via Acanthamoeba.

  2. Essential Role for an M17 Leucine Aminopeptidase in Encystation of Acanthamoeba castellanii

    PubMed Central

    Lee, Yu-Ran; Na, Byoung-Kuk; Moon, Eun-Kyung; Song, Su-Min; Joo, So-Young; Kong, Hyun-Hee; Goo, Youn-Kyoung; Chung, Dong-Il; Hong, Yeonchul

    2015-01-01

    Encystation of Acanthamoeba leads to the formation of resilient cysts from vegetative trophozoites. This process is essential for parasite survival under unfavorable conditions such as starvation, low temperatures, and exposure to biocides. During encystation, a massive turnover of intracellular components occurs, and a large number of organelles and proteins are degraded by proteases. Previous studies with specific protease inhibitors have shown that cysteine and serine proteases are involved in encystation of Acanthamoeba, but little is known about the role of metalloproteases in this process. Here, we have biochemically characterized an M17 leucine aminopeptidase of Acanthamoeba castellanii (AcLAP) and analyzed its functional involvement in encystation of the parasite. Recombinant AcLAP shared biochemical properties such as optimal pH, requirement of divalent metal ions for activity, substrate specificity for Leu, and inhibition profile by aminopeptidase inhibitors and metal chelators with other characterized M17 family LAPs. AcLAP was highly expressed at a late stage of encystation and mainly localized in the cytoplasm of A. castellanii. Knockdown of AcLAP using small interfering RNA induced a decrease of LAP activity during encystation, a reduction of mature cyst formation, and the formation of abnormal cyst walls. In summary, these results indicate that AcLAP is a typical M17 family enzyme that plays an essential role during encystation of Acanthamoeba. PMID:26075721

  3. Isolation of Acanthamoeba Spp. from Drinking Waters in Several Hospitals of Iran

    PubMed Central

    Bagheri, HR; Shafiei, R; Shafiei, F; Sajjadi, SA

    2010-01-01

    Background Acanthamoeba is an opportunistic amphizoic protozoan found in different water sources including swimming pool as well as in sewage. The aim of this study was to investigate the prevalence of Acanthamoeba in tap-water samples in Iran. Method In this descriptive cross-sectional study, 94 samples of cold and warm tap-water were collected from different wards of hospitals in 13 cities of Iran in 2007–2008. Free residual chlorine, pH, and temperature of samples were measured. After filtration through multipore nylon membrane, samples were cultured on non-nutrient agar. Then we investigated existence of Acanthamoeba by reverse contrast phase microscope. Results Acanthamoeba was found in 45 samples (48%). Thirty-four and 11 positive samples were collected from cold and warm tap water, respectively. The samples belonged to the category of 20–30°C temperature with 0–2 ppm free residual chlorine and pH 6–7.4 showed the most coincidence to the positive cases. The greatest proportion of positive samples was obtained from Mashhad hospitals, while all samples collected from Arak and Semnan hospitals were negative. Conclusion considering the results of this study and the pathogenic role of this protozoan on patients with immunodeficiency, as well as capability of this microorganism in carrying other pathogens such as Legionella, further studies are needed. What is more important, potable water in hospitals should follow the procedure of treatment and sanitation, in order to prevent the relevant nosocomial infections. PMID:22347240

  4. Occurrence of pathogenic Acanthamoeba genotypes in nasal swabs of cancer patients in Iran.

    PubMed

    Memari, Fatemeh; Niyyati, Maryam; Haghighi, Ali; Seyyed Tabaei, Seyyed Javad; Lasjerdi, Z

    2015-05-01

    Incidences of Acanthamoeba granulomatous encephalitis (AGE) have been increased due to a rise in the number of high-risk people, such as immunodeficient patients. Indeed, immunosuppress situation can render the patient in acquiring opportunistic Acanthamoeba infections. In this study, analysis was carried out to verify the presence of free-living amoebae of Acanthamoeba genus in nasal swabs of cancer patients in hospitals of Tehran, Iran. Detection of isolates was based on morphotyping and PCR sequencing of the Diagnostic Fragment 3 (DF3) to identify strains at the genotype level. In addition, the pathogenic potential of the isolates was assayed using temperature and osmotolerance assays. The obtained results revealed that nine isolated strains belonging to T4 genotype-exhibited pathogenic potential. After sequencing, genotype T4 was found to be the most common one in the samples included in this study. Genotype T3 and T5 were also identified. To the best of our knowledge, this is the first study on the typing of Acanthamoeba strains at the genotype level in cancer patients in Iran and worldwide.

  5. Developmental changes in the localization of calcium binding sites in Acanthamoeba castellanii.

    PubMed

    Sobota, A; Przelecka, A

    1981-01-01

    Vegetative cells of Acanthamoeba castellanii have the ability to bind calcium on the plasma membrane in form of the electron-dense deposits. The appearance of the deposits depends on the age of Acanthamoeba culture. In 24-h-old culture the deposits are very small, with diameter of 26 nm. During aging of culture, at both logarithmic and stationary growth phases, the diameter of deposits is larger (70-80 nm), while the deposits are localized only on the plasma membrane. During differentiation of Acanthamoeba cells into cysts electron-dense deposits with a diameter of about 170 nm appear in the mitochondria, whereas no deposits are observed on the plasma membrane. However, at the first stage of differentiation electron-dense material together with extruded membraneous fragments are also observed outside of some newly-formed young cysts. These results suggest that in Acanthamoeba cells, depending on the stage of life cycle, either plasma membrane or mitochondria may be involved in storage of excess cellular calcium. PMID:7228741

  6. Acanthamoeba Encephalitis: Isolation of Genotype T1 in Mycobacterial Liquid Culture Medium

    PubMed Central

    Azzam, Rula; Badenoch, Paul R.; Francis, Michelle J.; Fernandez, Charles; Adamson, Penelope J.; Dendle, Claire; Woolley, Ian; Robson, Jenny; Korman, Tony M.

    2014-01-01

    We report a case of Acanthamoeba encephalitis diagnosed from an antemortem brain biopsy specimen, where the organism was first isolated in mycobacterial liquid medium and first identified by using a sequence generated by a commercial panfungal sequencing assay. We correlate susceptibility results with clinical outcome. PMID:25502534

  7. Is blood agar an alternative to sabouraud dextrose agar for the isolation of fungi in patients with mycotic keratitis.

    PubMed

    Reddy, Ashok Kumar; Brahmaiah, Upputuri; Narayen, Nitesh; Reddy, Ravi Kumar; Reddy, Rupak Kumar; Chitta, Meghraj; Prasad, Srinivas; Swarup, Rishi; Mohiuddin, Syed Maaz; Reddy, Madhukar; Aasuri, Murali K; Murthy, B S R; Bhide, Milind; Ahmed, Sajid

    2013-06-01

    To compare the blood agar (BA), sabouraud dextrose agar (SDA) and chocolate agar (CA) for the isolation of fungi in patients with mycotic keratitis. Corneal Scrapings of 229 patients with clinically diagnosed microbial keratitis were inoculated on BA, SDA, CA. The culture media were evaluated for the rate and time taken for the fungal growth. Seventy six of 229 patients had fungal keratitis. Fungus grew on BA in 60/76(78.9 %), on SDA in 76/76 (100 %), on CA in 40/76(52.6 %) patients. The fungi which grew on BA (60/76) also grown on SDA at the same time. The colony morphologies of different fungi were better on SDA than BA/CA. Among the different culture media, SDA is essential for the isolation fungi in patients with mycotic keratitis.

  8. Anti-amoebic properties of a Malaysian marine sponge Aaptos sp. on Acanthamoeba castellanii.

    PubMed

    Nakisah, M A; Ida Muryany, M Y; Fatimah, H; Nor Fadilah, R; Zalilawati, M R; Khamsah, S; Habsah, M

    2012-03-01

    Crude methanol extracts of a marine sponge, Aaptos aaptos, collected from three different localities namely Kapas, Perhentian and Redang Islands, Terengganu, Malaysia, were tested in vitro on a pathogenic Acanthamoeba castellanii (IMR isolate) to examine their anti-amoebic potential. The examination of anti-Acanthamoebic activity of the extracts was conducted in 24 well plates for 72 h at 30 °C. All extracts possessed anti-amoebic activity with their IC(50) values ranging from 0.615 to 0.876 mg/mL. The effect of the methanol extracts on the surface morphology of A. castellanii was analysed under scanning electron microscopy. The ability of the extracts to disrupt the amoeba cell membrane was indicated by extensive cell's blebbing, changes in the surface morphology, reduced in cell size and with cystic appearance of extract-treated Acanthamoeba. Number of acanthapodia and food cup was also reduced in this Acanthamoeba. Morphological criteria of apoptosis in Acanthamoeba following treatment with the sponge's extracts was determined by acridine orange-propidium iodide staining and observed by fluorescence microscopy. By this technique, apoptotic and necrotic cells can be visualized and quantified. The genotoxic potential of the methanol extracts was performed by the alkaline comet assay. All methanol extracts used were significantly induced DNA damage compared to untreated Acanthamoeba by having high percentage of scores 1, 2, and 3 of the DNA damage. Results from cytotoxicity and genotoxicity studies carried out in the present study suggest that all methanol extracts of A. aaptos have anti-amoebic properties against A. castellanii. PMID:22805843

  9. Preliminary data suggest a role for bacterial superinfection of a viral keratitis after zosteriform spread of herpes simplex virus to the eye of the mouse.

    PubMed Central

    Claoué, C

    1988-01-01

    After inoculation of herpes simplex virus (HSV) on the snout of the mouse, spread to the eye occurs via neural pathways. The keratitis which ensued was clinically atypical for a pure herpetic keratitis, leading to a search for other causative agents. Preliminary data suggests that bacteriological super-infection of the viral keratitis occurs frequently, producing a unique experimental model. Images Figure 1. Figure 2. Figure 3. Figure 4. Figure 5. Figure 6. PMID:3418659

  10. Necrotizing Meningoencephalitis in a Captive Black and White Ruffed Lemur (Varecia variegata variegata) Caused by Acanthamoeba T4 Genotype.

    PubMed

    Gaide, N; Pelandakis, M; Robveille, C; Albaric, O; Jouvion, G; Souchon, M; Risler, A; Abadie, J

    2015-11-01

    A mature male, black and white ruffed lemur (Varecia variegata variegata) died in a zoological garden after a 4-day history of lethargy and non-responsive convulsions. Necropsy and histopathological examinations revealed acute necrotizing and haemorrhagic meningoencephalitis with intralesional amoebas confirmed by immunohistochemistry. Acanthamoeba T4 genotype was identified as the causative agent of the brain lesion, based on amplification and sequencing of 18S ribosomal RNA genes. The presence of free-living amoebas in water and mud from the lemur's environment was investigated by morphological and molecular analyses. The two predominant genera, representing 80% of isolated amoebas, were Naegleria spp. and Acanthamoeba spp. All Acanthamoeba isolates belonged to the T4 genotype. To the author's knowledge, this is the first report of a meningoencephalitis due to Acanthamoeba T4 genotype in Lemuridae with concurrent analysis of pathological tissues and environment.

  11. Status of free-living amoebae (Acanthamoeba spp., Naegleria fowleri, Balamuthia mandrillaris) in drinking water supplies in Karachi, Pakistan.

    PubMed

    Yousuf, Farzana Abubakar; Siddiqui, Ruqaiyyah; Subhani, Faysal; Khan, Naveed Ahmed

    2013-06-01

    The ability of pathogenic free-living amoebae to produce infections is a growing concern. In this study, we investigated the presence of free-living amoebae (Acanthamoeba spp., Naegleria fowleri, Balamuthia mandrillaris) in drinking water supplies in Karachi, Pakistan. Fifty-two domestic tap water samples were examined. Amoebae were identified by morphological characteristics and polymerase chain reaction. Thirty percent of the examined samples were positive for Acanthamoeba spp., 8% for N. fowleri while B. mandrillaris were not recovered. Additionally we examined secretory IgA antibody to Acanthamoeba and B. mandrillaris. Acanthamoeba antibody prevalence rate was 100% in both males and females, while B. mandrillaris antibody prevalence rate was 5.5% in males only (females were negative). Our findings suggest that free-living amoebae are a potential health hazard in domestic water supplies in Karachi, Pakistan. PMID:23708583

  12. Status of free-living amoebae (Acanthamoeba spp., Naegleria fowleri, Balamuthia mandrillaris) in drinking water supplies in Karachi, Pakistan.

    PubMed

    Yousuf, Farzana Abubakar; Siddiqui, Ruqaiyyah; Subhani, Faysal; Khan, Naveed Ahmed

    2013-06-01

    The ability of pathogenic free-living amoebae to produce infections is a growing concern. In this study, we investigated the presence of free-living amoebae (Acanthamoeba spp., Naegleria fowleri, Balamuthia mandrillaris) in drinking water supplies in Karachi, Pakistan. Fifty-two domestic tap water samples were examined. Amoebae were identified by morphological characteristics and polymerase chain reaction. Thirty percent of the examined samples were positive for Acanthamoeba spp., 8% for N. fowleri while B. mandrillaris were not recovered. Additionally we examined secretory IgA antibody to Acanthamoeba and B. mandrillaris. Acanthamoeba antibody prevalence rate was 100% in both males and females, while B. mandrillaris antibody prevalence rate was 5.5% in males only (females were negative). Our findings suggest that free-living amoebae are a potential health hazard in domestic water supplies in Karachi, Pakistan.

  13. Infection in a rat model reactivates attenuated virulence after long-term axenic culture of Acanthamoeba spp

    PubMed Central

    Veríssimo, Carolina De Marco; Maschio, Vinícius José; Correa, Ana Paula Folmer; Brandelli, Adriano; Rott, Marilise Brittes

    2013-01-01

    Prolonged culturing of many microorganisms leads to the loss of virulence and a reduction of their infective capacity. However, little is known about the changes in the pathogenic strains of Acanthamoeba after long culture periods. Our study evaluated the effect of prolonged culturing on the invasiveness of different isolates of Acanthamoeba in an in vivo rat model. ATCC strains of Acanthamoeba, isolates from the environment and clinical cases were evaluated. The in vivo model was effective in establishing the infection and differentiating the pathogenicity of the isolates and re-isolates. The amoebae cultured in the laboratory for long periods were less virulent than those that were recently isolated, confirming the importance of passing Acanthamoeba strains in animal models. PMID:24271042

  14. Necrotizing Meningoencephalitis in a Captive Black and White Ruffed Lemur (Varecia variegata variegata) Caused by Acanthamoeba T4 Genotype.

    PubMed

    Gaide, N; Pelandakis, M; Robveille, C; Albaric, O; Jouvion, G; Souchon, M; Risler, A; Abadie, J

    2015-11-01

    A mature male, black and white ruffed lemur (Varecia variegata variegata) died in a zoological garden after a 4-day history of lethargy and non-responsive convulsions. Necropsy and histopathological examinations revealed acute necrotizing and haemorrhagic meningoencephalitis with intralesional amoebas confirmed by immunohistochemistry. Acanthamoeba T4 genotype was identified as the causative agent of the brain lesion, based on amplification and sequencing of 18S ribosomal RNA genes. The presence of free-living amoebas in water and mud from the lemur's environment was investigated by morphological and molecular analyses. The two predominant genera, representing 80% of isolated amoebas, were Naegleria spp. and Acanthamoeba spp. All Acanthamoeba isolates belonged to the T4 genotype. To the author's knowledge, this is the first report of a meningoencephalitis due to Acanthamoeba T4 genotype in Lemuridae with concurrent analysis of pathological tissues and environment. PMID:26297109

  15. Real-time PCR method for the detection and quantification of Acanthamoeba species in various types of water samples.

    PubMed

    Kao, Po-Min; Tung, Min-Che; Hsu, Bing-Mu; Tsai, Hsien-Lung; She, Cheng-Yu; Shen, Shu-Min; Huang, Wen-Chien

    2013-03-01

    In this study, a quantitative real-time PCR was developed to detect and quantify Acanthamoeba spp. in various environmental water samples. The water samples were taken from watershed, water treatment plant, and three thermal spring recreation areas. The overall detection rate was 14.2 % (25/176) for Acanthamoeba spp. The percentages of samples containing Acanthamoeba spp. from river water, raw drinking water, and thermal spring water were 13 % (13/100), 25 % (7/28), and 10.4 % (5/48), respectively. Acanthamoeba spp. concentrations were determined according to SYBR Green quantitative real-time PCR. A plasmid-based standard curve was constructed to determine the Acanthamoeba concentration using dilution factors for achieving 1.36 × 10(9) gene copies per PCR for 18S rRNA gene in Acanthamoeba spp. The resulting concentrations varied by the type of water, in the range of 46-2.6 × 10(2) cells/l in positive raw drinking water, 2.7 × 10(2)-1.5 × 10(4) cells/l in river water, and 54-1.7 × 10(3) cells/l in thermal spring water. The presence of Acanthamoeba spp. in the raw drinking water samples was also found to have a significant difference with heterotrophic plate count. The presence of Acanthamoeba spp. in various aquatic environments may be a potential health hazard and must be further evaluated.

  16. Isolation and molecular characterization of Acanthamoeba genotypes in recreational and domestic water sources from Jamaica, West Indies.

    PubMed

    Todd, Cheridah D; Reyes-Batlle, María; Piñero, José E; Martínez-Carretero, Enrique; Valladares, Basilio; Streete, Don; Lorenzo-Morales, Jacob; Lindo, John F

    2015-09-01

    Free living amoebae (FLA) are amphizoic protozoa that are ubiquitous in nature. Infection with FLA may result in neurological, ocular and skin infections. Exposure to Acanthamoeba occurs frequently through water contact and knowledge of the presence of the organisms in water sources is important in understanding transmission dynamics. The distribution of Acanthamoeba was studied in recreational and domestic water samples collected from across Jamaica. Morphological assessment and polymerase chain reaction revealed Acanthamoeba spp. isolates in 50.6% (42/83) and 17.3% (14/81) of recreational and domestic water, respectively. Sequencing of the DF3 region of the 18S rDNA resulted in the identification of genotypes T3, T4, T5, T10 and T11 corresponding to Acanthamoeba spp: A. griffini, A. triangularis, A. lenticulata, A. culbertsoni and A. hatchetti. Moreover, T4 was the most frequently isolated genotype in both recreational and domestic water. Thermotolerance and osmotolerance assays indicated that most isolates were potentially pathogenic. This is the first report of T3 and T10 genotypes in the Caribbean and the first report of these Acanthamoeba spp. in Jamaican waters. The study shows that there is potential risk of infection to contact wearers who practise poor lens care. Further, Acanthamoeba should be considered as a cause of neurological infections in Jamaica.

  17. Isolation and genotyping of free-living environmental isolates of Acanthamoeba spp. from bromeliads in Southern Brazil.

    PubMed

    Landell, Melissa Fontes; Salton, Juliana; Caumo, Karin; Broetto, Leonardo; Rott, Marilise B

    2013-07-01

    Species of Acanthamoeba are frequently isolated from distinct environmental sources such as water, soil, dust and air. They are responsible to cause infections and disease in humans and animals. In addition, Acanthamoeba sp. are considered an important reservoir of bacteria, virus and fungi, which act as "Trojan horses" to protect these microorganisms of harsh environmental conditions. In this study, nine Acanthamoeba isolates from bromeliads phylloplane were identified based on the morphology of cyst and trophozoite forms. The genotype level was accessed by the sequence analysis of Acanthamoeba small-subunit rRNA gene. Genotypic characterization grouped five isolates in the genotype T2/T6, three in the T4 genotype and one in the genotype T16. The results obtained indicate that the genotype T2/T6 is common on phylloplane. To predict the pathogenic potential of the Acanthamoeba isolates, thermo and osmotolerance assays were employed, although all isolates were capable of surviving at temperatures of 37°C, other tests will be conducted in the future to determine the potential pathogenic of the isolates. Altogether, our results revealed the importance of the presence of Acanthamoeba associated with bromeliads in Rio Grande do Sul, Brazil, and the necessity for further studies to determine the environmental distribution and the role of these species. PMID:23562883

  18. Imported Pythium insidiosum Keratitis After a Swim in Thailand by a Contact Lens-Wearing Traveler

    PubMed Central

    Lelievre, Lucie; Borderie, Vincent; Garcia-Hermoso, Dea; Brignier, Anne C.; Sterkers, Margaret; Chaumeil, Christine; Lortholary, Olivier; Lanternier, Fanny

    2015-01-01

    A 30-year-old woman with a history of contact lens wear and exposure to swimming pool water in Thailand presented with a non-responsive, progressive corneal ulcer of the right eye. Confocal microscopy evidenced septate linear branching structures, raising suspicion of fungal keratitis. She was promptly treated with topical antibiotics and both topical and intravenous caspofungin plus voriconazole. Worsening of the clinical picture after 1 month of intensive medical therapy led to a large therapeutic penetrating keratoplasty being performed. Corneal cultures grew a mold-like organism, which was identified by sequencing as Pythium insidiosum, an aquatic oomycete. After 4 years of follow-up, the graft exhibits no infection relapse, but graft transparency has been lost after two rejection episodes. Keratoplasty combined with antifungal treatment may offer a cure to P. insidiosum keratitis, although long-term preservation of corneal transparency is difficult to obtain. PMID:25535313

  19. Long-term follow-up after successful treatment of Pythium insidiosum keratitis in Israel.

    PubMed

    Barequet, Irina S; Lavinsky, Fabio; Rosner, Mordechai

    2013-07-01

    The purpose of this project is to report a case of severe Pythium insidiosum keratitis confirmed by polymerase chain reaction (PCR), and its long-term cure after therapeutic penetrating keratoplasty. A 24-year-old woman with a history of contact lens wear and exposure to swimming pool water presented with a severe corneal abscess. She was treated with intensive fortified topical antibiotics and natamycin with limited response. Initial cultures suggested the presence of a septate mold, unclearly identified; therefore, both topical and intravenous voriconazole were administered. Despite the above treatment, there was worsening of the clinical picture. PCR assay revealed homology to Pythium insidiosum. Promptly, the patient underwent a large therapeutic penetrating keratoplasty. After five years of follow-up, the graft exhibits neither signs of rejection nor any recurrence of infection. We conclude that prompt identification of Pythium insidiosum keratitis and aggressive treatment by therapeutic penetrating keratoplasty may offer a cure to this disease. PMID:23627654

  20. Imported pythium insidiosum keratitis after a swim in Thailand by a contact lens-wearing traveler.

    PubMed

    Lelievre, Lucie; Borderie, Vincent; Garcia-Hermoso, Dea; Brignier, Anne C; Sterkers, Margaret; Chaumeil, Christine; Lortholary, Olivier; Lanternier, Fanny

    2015-02-01

    A 30-year-old woman with a history of contact lens wear and exposure to swimming pool water in Thailand presented with a non-responsive, progressive corneal ulcer of the right eye. Confocal microscopy evidenced septate linear branching structures, raising suspicion of fungal keratitis. She was promptly treated with topical antibiotics and both topical and intravenous caspofungin plus voriconazole. Worsening of the clinical picture after 1 month of intensive medical therapy led to a large therapeutic penetrating keratoplasty being performed. Corneal cultures grew a mold-like organism, which was identified by sequencing as Pythium insidiosum, an aquatic oomycete. After 4 years of follow-up, the graft exhibits no infection relapse, but graft transparency has been lost after two rejection episodes. Keratoplasty combined with antifungal treatment may offer a cure to P. insidiosum keratitis, although long-term preservation of corneal transparency is difficult to obtain. PMID:25535313

  1. Bacterial Keratitis: Perspective on Epidemiology, Clinico-Pathogenesis, Diagnosis and Treatment

    PubMed Central

    Al-Mujaini, Abdullah; Al-Kharusi, Nadia; Thakral, Archana; Wali, Upender K

    2009-01-01

    Bacterial keratitis is an acute or chronic, transient or recurrent infection of the cornea with varying predilection for anatomical and topographical parts of the cornea like marginal or central. It is a potentially sight-threatening corneal infection in humans that is generally found in eyes with predisposing elements, the most common of which is contact lens wear. The epidemiological data reveals the universal occurrence of this disease. With advances in the understanding of its pathogenesis, laboratory investigations like immunohistochemistry, fluorescent microscopy, enzyme immunoassays and molecular biology, and the availability of fourth generation antibiotics, the overall visual outcome in bacterial keratitis has improved with time. Particular attention should be given to this condition as it can progress very rapidly with complete corneal destruction occurring within 24–48 hours. Early diagnosis, which is primarily clinical and substantiated largely by microbiological data, and prompt treatment are needed to minimise the possibility of permanent visual loss and reduce structural damage to the cornea. PMID:21509299

  2. Geographic Variations in Microbial Keratitis: An analysis of the Peer-Reviewed Literature

    PubMed Central

    Shah, Ameet; Sachdev, Arun; Coggon, David; Hossain, Parwez

    2012-01-01

    summary The epidemiology of microbial keratitis has been investigated in several studies by analysis of organisms cultured from corneal scrapes. However, a comparison of the frequency of different organisms causing keratitis in different parts of the world is lacking. We present a review incorporating an analysis of data from studies worldwide. The data provide a comparison of the frequency of culture-positive organisms found in different parts of the world. The highest proportion of bacterial corneal ulcers was reported in studies from North America, Australia, the Netherlands and Singapore. The highest proportion of staphylococcal ulcers was found in a study from Paraguay whilst the highest proportion of pseudomonas ulcers was reported in a study from Bangkok. The highest proportions of fungal infections were found in studies from India and Nepal. Possible explanations for these observed geographic variations are discussed. PMID:21478201

  3. Autologous eye drops for the treatment of dry eye and neurotrophic keratitis.

    PubMed

    Mixon, William; Angelle, Patricea Patsy; Chang, Richard I

    2009-01-01

    Tears-which contain fibronectin, growth facors, and vitamins that support the migration, proliferation, and differentiation of the conjunctival and corneal epithelium-nourish, protect, and refresh the eye. An inadequate tear film produces a variety of symptoms (ocular irritation, itching, burning, eye strain or fatigue, photophobia, a foreign-body sensation, a sharp stabbing pain, blurred vision) that wax and wane in intensity and are characteristic of disorders such as dry eye (keratoconjunctivitis sicca) and neurotrophic keratitis. In patients for whom commercially manufactured eye drops fail to provide relief from the symptoms of those conditions, autologous serum eye drops (plasma eye drops, serum eye drops) have proven very effective. In this report, we examine the present case studies that reveal the efects of autologous serum eye drops on the signs and symptoms of both dry eye and neurotrophic keratitis. A procedure for preparing autologous eye drops is included.

  4. Identification of pathogenic factors potentially involved in Staphylococcus aureus keratitis using proteomics.

    PubMed

    Khan, Shamila; Cole, Nerida; Hume, Emma B H; Garthwaite, Linda L; Nguyen-Khuong, Terry; Walsh, Bradley J; Willcox, Mark D P

    2016-10-01

    Staphylococcus is a leading cause of microbial keratitis, characterized by destruction of the cornea by bacterial exoproteins and host-associated factors. The aim of this study was to compare extracellular and cell-associated proteins produced by two different isolates of S. aureus, a virulent clinical isolate (Staph 38) and a laboratory strain (Staphylococcus aureus 8325-4) of weaker virulence in the mouse keratitis model. Proteins were analyzed using 2D polyacrylamide gel electrophoresis and identified by subsequent mass spectrometry. Activity of staphylococcal adhesins was assessed by allowing strains to bind to various proteins adsorbed onto polymethylmethacrylate squares. Thirteen proteins in the extracellular fraction and eight proteins in the cell-associated fractions after bacterial growth were produced in increased amounts in the clinical isolate Staph 38. Four of these proteins were S. aureus virulence factor adhesins, fibronectin binding protein A, staphopain, glyceraldehyde-3-phosphate dehydrogenase 2 and extracellular adherence protein. The clinical isolate Staph 38 adhered to a greater extent to all mammalian proteins tested, indicating the potential of the adhesins to be active on its surface. Other proteins with increased expression in Staph 38 included potential moonlighting proteins and proteins involved in transcription or translation. This is the first demonstration of the proteome of S. aureus isolates from keratitis. These results indicate that the virulent clinical isolate produces more potentially important virulence factors compared to the less virulent laboratory strain and these may be associated with the ability of a S. aureus strain to cause more severe keratitis.

  5. Topical voriconazole therapy of Purpureocillium lilacinum keratitis that occurred in disposable soft contact lens wearers.

    PubMed

    Todokoro, Daisuke; Yamada, Norihiro; Fukuchi, Mariko; Kishi, Shoji

    2014-10-01

    The objective of this study was to describe 2 cases of keratitis caused by Purpureocillium lilacinum (formerly Paecilomyces lilacinus) that occurred in disposable contact lens users, which were successfully treated with topical voriconazole. Case 1 was a healthy 44-year-old woman, who wore weekly disposable contact lenses and had developed a superficial corneal infection in her right eye. For diagnosis, corneal scraping and molecular identification of the cultured pathogen were performed. A corneal smear revealed the presence of fungi. The pathogen was identified as P. lilacinum by traditional morphological identification of fungal culture, and this identification was confirmed by DNA sequencing of the ribosomal internal transcribed spacer (ITS) sequence. Therapy with topical fluconazole, topical pimaricin (natamycin), and oral itraconazole were ineffective. Topical voriconazole showed a significant effect, and the keratitis was successfully treated. Case 2 was a 43-year-old woman with bilateral recurrent peripheral corneal ulcers by meibomian gland dysfunction, who used therapeutic bandage contact lenses on her left eye. However, a corneal abscess with hypopyon occurred in the eye after 3 months. The microbial smear examination showed the presence of fungi and the fungal culture, and the DNA sequence of ITS region revealed that the causative agent was P. lilacinum. The susceptibility testing against antifungal agents showed that voriconazole was effective. The lesion improved gradually by topical voriconazole. As a conclusion, P. lilacinum keratitis can occur in disposable soft contact lens wearer. Early and accurate detection of the pathogenic organism is essential. Topical voriconazole was effective against P. lilacinum keratitis. PMID:24974133

  6. Dental Treatments under the General Anesthesia in a Child with Keratitis, Ichthyosis, and Deafness Syndrome

    PubMed Central

    Derelioglu, Sera Sımsek; Yılmaz, Yücel; Keles, Sultan

    2013-01-01

    KID syndrome is a rare genodermatosis characterized by keratitis, ichthyosis, and sensorineural deafness. Although the dermatological, ophthalmologic, and sensorineural defects are emphasized in the literature, oral and dental evaluations are so superficial. In this case report, dental and oral symptoms of a three year and five months old boy with KID syndrome, suffering severe Early Childhood Caries (s-ECC) and dental treatments done under General Anesthesia (GA) were reported. PMID:24151560

  7. Big bubble deep anterior lamellar keratoplasty for management of deep fungal keratitis.

    PubMed

    Gao, Hua; Song, Peng; Echegaray, Jose J; Jia, Yanni; Li, Suxia; Du, Man; Perez, Victor L; Shi, Weiyun

    2014-01-01

    Objective. To evaluate the therapeutic effect of big bubble deep anterior lamellar keratoplasty (DALK) in patients with deep fungal keratitis. Methods.Consecutive patients who had DALK for deep fungal keratitis at Shandong Eye Hospital between July 2011 and December 2012 were included. In all patients, the infiltration depth was more than 4/5ths of the corneal thickness. DALK surgery was performed with bare Descemet membrane (DM) using the big bubble technique. Corrected distance visual acuity (CDVA), graft status, and intraoperative and postoperative complications were monitored. Results. Big bubble DALK was performed in 23 patients (23 eyes). Intraoperative perforation of the DM occurred in two eyes (8.7%) during stromal dissection. The patients received lamellar keratoplasty with an air bubble injected into the anterior chamber. Double anterior chamber formed in 3 eyes (13.0%). Mean CDVA of the patients without cataract, amblyopia, and fungal recurrence was improved from preoperative HM/20 cm-1.0 (LogMAR) to 0.23 ± 0.13 (LogMAR) at the last followup (P < 0.01). Fungal recurrence was found in two patients (8.7%). Corneal stromal graft rejection was noted in one patient (4.3%). Conclusions. DALK using the big bubble technique seems to be effective and safe in the treatment of deep fungal keratitis unresponsive to medication. PMID:25105019

  8. Rheumatoid polyarthritis suspected in an HIV patient with scleritis, peripheral ulcerative keratitis, and anterior uveitis

    PubMed Central

    Domngang Noche, Christelle; Singwé-Ngandeu, Madeleine; Bella, Assumpta Lucienne

    2016-01-01

    Introduction Scleritis and peripheral ulcerative keratitis are ocular manifestations found in many inflammations and infections. Therefore, their association should prompt a search for inflammatory or infectious causes that may be life-threatening, especially in the context of AIDS due to HIV infection. Findings We report the case of a 37-year-old female, first seen in 2011 with a nodular scleritis in the right eye and a peripheral ulcerative keratitis, a necrotizing scleritis, and a granulomatous anterior uveitis in the left eye, in the context of chronic polyarthropathies that had evolved over 6 months. The patient was diagnosed with AIDS (HIV) in 2008 and was on antiretroviral therapy for the past 2 years. Ophthalmic workup was negative for opportunistic infections and potential causes of scleritis and peripheral ulcerative keratitis, and the patient was unresponsive to topical antibacterial and anti-inflammatory treatment. Ocular lesion resolution and articular swelling improvement was observed less than 6 weeks after sulfasalazine treatment. Based on American College of Rheumatology/European League Against Rheumatism classification criteria, and considering the good response to the treatment (sulfasalazine), diagnosis of rheumatoid arthritis was made in the absence of confirmatory lab tests results. Conclusion In the context of ocular manifestations associated with polyarthropathies, coexisting pathologies should be considered. Diagnostic workup of chronic inflammatory rheumatism should be carried out, even in the context of HIV/AIDS. PMID:26893583

  9. Multi-drug resistant Pseudomonas aeruginosa keratitis and its effective treatment with topical colistimethate

    PubMed Central

    Chatterjee, Samrat; Agrawal, Deepshikha

    2016-01-01

    The purpose was to evaluate the clinical outcome in multi-drug resistant Pseudomonas aeruginosa (MDR-PA) bacterial keratitis and report the successful use of an alternative antibiotic, topical colistimethate in some of them. The medical records of 12 culture-proven MDR-PA keratitis patients, all exhibiting in vitro resistance by Kirby–Bauer disc diffusion method to ≥ three classes of routinely used topical antibiotics were reviewed. Eight patients were treated with 0.3% ciprofloxacin or ofloxacin, 1 patient with 5% imipenem/cilastatin and 3 patients with 1.6% colistimethate. The outcomes in 8 eyes treated with only fluoroquinolones were evisceration in 4 eyes, therapeutic corneal graft in 1 eye, phthisis bulbi in 1 eye, and no improvement in 2 eyes. The eye treated with imipenem/cilastin required a therapeutic corneal graft. All the three eyes treated with 1.6% colistimethate healed. Colistimethate may prove to be an effective alternative antibiotic in the treatment of MDR-PA keratitis. PMID:27050354

  10. Big bubble deep anterior lamellar keratoplasty for management of deep fungal keratitis.

    PubMed

    Gao, Hua; Song, Peng; Echegaray, Jose J; Jia, Yanni; Li, Suxia; Du, Man; Perez, Victor L; Shi, Weiyun

    2014-01-01

    Objective. To evaluate the therapeutic effect of big bubble deep anterior lamellar keratoplasty (DALK) in patients with deep fungal keratitis. Methods.Consecutive patients who had DALK for deep fungal keratitis at Shandong Eye Hospital between July 2011 and December 2012 were included. In all patients, the infiltration depth was more than 4/5ths of the corneal thickness. DALK surgery was performed with bare Descemet membrane (DM) using the big bubble technique. Corrected distance visual acuity (CDVA), graft status, and intraoperative and postoperative complications were monitored. Results. Big bubble DALK was performed in 23 patients (23 eyes). Intraoperative perforation of the DM occurred in two eyes (8.7%) during stromal dissection. The patients received lamellar keratoplasty with an air bubble injected into the anterior chamber. Double anterior chamber formed in 3 eyes (13.0%). Mean CDVA of the patients without cataract, amblyopia, and fungal recurrence was improved from preoperative HM/20 cm-1.0 (LogMAR) to 0.23 ± 0.13 (LogMAR) at the last followup (P < 0.01). Fungal recurrence was found in two patients (8.7%). Corneal stromal graft rejection was noted in one patient (4.3%). Conclusions. DALK using the big bubble technique seems to be effective and safe in the treatment of deep fungal keratitis unresponsive to medication.

  11. Keratitis-ichthyosis-deafness syndrome in association with follicular occlusion triad.

    PubMed

    Maintz, Laura; Betz, Regina C; Allam, Jean-Pierre; Wenzel, Jörg; Jaksche, Axel; Friedrichs, Nicolaus; Bieber, Thomas; Novak, Natalija

    2005-01-01

    Keratitis-Ichthyosis-Deafness syndrome is a rare congenital disorder of the ectoderm caused by mutations in the connexin-26 gene (GJB2) on chromosome 13q11-q12, giving rise to keratitis, erythrokeratoderma and neurosensory deafness. We report the case of a 31-year-old black male diagnosed as having KID syndrome. Sequencing analysis showed a heterozygous missense mutation D50N (148G > A) in the GJB2 gene. In addition to the classical features of vascularizing keratitis, erythrokeratoderma and congenital deafness, our patient presented a follicular occlusion triad with hidradenitis suppurativa (HS, alias acne inversa), acne conglobata and dissecting cellulitis of the scalp, leading to cicatricial alopecia and disfiguring, inflammatory vegetations of his scalp. Conservative therapy such as a keratolytic, rehydrating and antiseptic external therapy, antibiotic, antimycotic and retinoids were only of moderate benefit, so we finally chose the curative possibility of surgery therapy of the axillar papillomas and of the scalp. The inflammatory papillomatous regions of the axillae and of the scalp were radically debrided. Clean granulation was awaited and covered in a second session with a mesh graft from the thigh, achieving a satisfactory result. To our knowledge, only one case of KID syndrome occurring in association with follicular occlusion triad has been reported before. PMID:16172043

  12. Fungal Keratitis Due to Beauveria bassiana in a Contact Lenses Wearer and Review of Published Reports.

    PubMed

    Lara Oya, Ana; Medialdea Hurtado, María Eloisa; Rojo Martín, María Dolores; Aguilera Pérez, Antonia; Alastruey-Izquierdo, Ana; Miranda Casas, Consuelo; Rubio Prats, Marina; Medialdea Marcos, Santiago; Navarro Marí, José María

    2016-10-01

    Fungal keratitis is a severe ocular infection that primarily affects subjects engaged in outdoor activities. Risk factors include allergic conjunctivitis, previous eye surgery, previous treatment with wide-spectrum antimicrobial agents and corticosteroids and using contact lenses. Corneal infection is usually secondary to trauma involving organic material, which is often the only predisposing factor. Early diagnosis based on clinical examination and microbiological investigation (microscopy, cultures and molecular techniques) is crucial to selecting the appropriate antifungal therapy and prevent progression. We report the case of a patient with keratitis due to Beauveria bassiana, an opportunistic and entomopathogenic filamentous fungus that is used as a biological insecticide and which is a rare cause of corneal infection. We review previous cases reports of B. bassiana keratitis published and its main features to compare with our case, a female occasional agriculture worker who had not suffered any trauma involving organic material. The patient received topical and oral antifungal therapy and debridement surgery, with a satisfactory outcome. PMID:27300341

  13. Real-time polymerase chain reaction for the diagnosis of necrotizing herpes stromal keratitis

    PubMed Central

    Ma, Jun-Xin; Wang, Lin-Nong; Zhou, Ru-Xia; Yu, Yang; Du, Tong-Xin

    2016-01-01

    AIM To design, optimize and validate a rapid, internally controlled real-time polymerase chain reaction (RT-PCR) test for herpes simplex virus (HSV) in the diagnosis of necrotizing herpes stromal keratitis. METHODS Tears alone or together with corneal epithelium scrapings from 30 patients (30 eyes) suspected of necrotizing herpes stromal keratitis were tested for HSV DNA by RT-PCR. The samples were collected during the first visit and then on the subsequent 7, 14, 28, 42, and 56d. The symptoms of the patients were scored before treatment to determine the correlation between HSV concentration in the corneal epithelium scrapings and clinical scores. RESULTS The positive rate (46.4%) in the corneal epithelium group before the therapy was significantly higher than that (13.3%) in the tears group (P=0.006). There were 13 positive HSV patients before the therapy, the concentration of HSV DNA in corneal epithelium scrapings group was significantly higher than that in the tears group (paired t-test, P=0.0397). Multilevel mixed-effects model analysis showed that the difference between the corneal epithelium scrapings group and the tears group was statistically significant (P=0.0049). The Spearman rank correlation analysis indicated a positive correlation between the HSV concentration in the corneal epithelium scrapings and clinical scores before the treatment (r=0.844, P<0.0001). CONCLUSION RT-PCR appears to be a powerful molecular tool for the diagnosis of necrotizing herpes stromal keratitis. PMID:27275421

  14. Acanthamoeba misidentification and multiple labels: redefining genotypes T16, T19, and T20 and proposal for Acanthamoeba micheli sp. nov. (genotype T19).

    PubMed

    Corsaro, Daniele; Walochnik, Julia; Köhsler, Martina; Rott, Marilise B

    2015-07-01

    Acanthamoeba species are ubiquitous amoebae able to cause important infections in humans and other vertebrates. The full/near-full sequences (>2000 bp) of the small subunit ribosomal RNA gene (SSU rDNA or 18S rDNA) are used to cluster Acanthamoeba as genotypes, labeled T1 to T20. Genotype T15 remains an exception, being described only partially on a 1500-bp fragment. Strains are thus usually identified based on their 18S identity matches with reference strains, often using shorter (<500 bp) diagnostic fragments of the gene. Nevertheless, short fragments (<1000 bp) have been used to propose genotypes. This has been criticized, and doubts arise therefore on possible confusion leading to classify distinct partial sequences with a same label(s). We demonstrate herein that several partial sequences misassigned either to T16 or to T4, actually belong to at least two separate and distinct genotypes. We obtained the full 18S rDNA of a strain previously typed as T16 on the basis of a small fragment and demonstrated that it actually belongs to the recently described T19. We propose the name Acanthamoeba micheli sp. nov., for this strain. Furthermore, partial molecular phylogenies were performed to show that several other misassigned T16 partial sequences belong to a new genotype. This latter includes also misassigned T4 partial sequences, only recently available as full sequences and labeled as T20. We thus reassign these partial sequences to the genotype T20. Longer sequences, ideally at least 90 % of the total gene length, should be obtained from strains to ensure reliable diagnostic and phylogenetic results.

  15. [Postoperative therapy after penetrating keratoplasty in herpes simplex keratitis].

    PubMed

    Süveges, Ildikó; Füst, Ágnes; Imre, László

    2013-12-29

    Bevezetés: A herpes simplex vírus által okozott szaruhártya-gyulladás a leggyakoribb oka a cornea centrumában kialakuló hegnek, amely látásvesztést okozhat. Célkitűzés: A szerzők célul tűzték ki a perforáló keratoplasztika eredményességének felmérését a szisztémás antiherpeses és immunszuppresszív terápia alkalmazásának tükrében. Módszer: Perforáló keratoplasztikán átesett 12 betegen végezték a retrospektív randomizált vizsgálatot. A műtéti beavatkozásig eltelt idő az első keratitis megjelenésétől számítva átlag 18 év volt (5–40 év). A műtéti indikáció 9 esetben a látás javítása, 3 esetben a cornea perforációjának megelőzése volt. Szisztémás kezelésként 9 beteg herpeszvírus elleni (acyclovir) és immunszuppresszív (mycophenolat mofetil), 2 beteg csak herpeszvírus elleni kezelést kapott, egy betegnél nem alkalmaztak szisztémás terápiát. Az átlagos követési idő 53,1 hónap volt (16–84 hó). Eredmények: A látásjavító célú 9 műtét közül 8 esetben a transzplantátum átlátszóan, ereződés nélkül gyógyult. Mind a 8 beteg acyclovir és mycophenolat mofetil kezelésben részesült. Egy esetben – amikor a beteg szisztémás kezelést nem kapott – recidíva és rejectio is fellépett. Az akut gyulladásos tünetekben végzett műtétek közül egyben gyógyult a transzplantátum átlátszóan, recidíva- és rejectiomentesen; a beteg acyclovir és mycophenolat mofetil terápiában részesült. Két esetben recidíva és rejectio is fellépett. Ezek közül egyben a beteg acyclovir és mycophenolat mofetil, egyben csak acyclovirkezelést kapott. A látóélesség minden esetben javult, 3 esetben a látást egyéb tényezők befolyásolták. Következtetések: A szisztémás acyclovir és mycophenolat mofetil terápia sikerrel alkalmazható herpes simplex keratitisben végzett perforáló keratoplasztikák után. Az acyclovir csökkenti a recidívák számát, a

  16. Distribution of Acanthamoeba Genotypes Isolated from Recreational and Therapeutic Geothermal Water Sources in Southwestern Iran

    PubMed Central

    Niyyati, Maryam; Saberi, Reza; Latifi, Alireza; Lasjerdi, Zohreh

    2016-01-01

    A comprehensive survey was conducted along 10 km of geothermal rivers in southwestern Iran. A total of 40 water samples were tested for the presence of Acanthamoeba spp., and genotypes were determined by targeting the diagnostic fragment 3 region of the 18S rRNA gene. The pathogenic potential of all positive isolates was also identified using tolerance ability test. High occurrences of Acanthamoeba (50%) were detected in the sampling areas. Based on sequencing analysis, isolates belonging to T4 (93.7%) and T2 (6.25%) genotypes were reported. Thermo- and osmotolerance tests revealed that five strains are highly pathogenic. Since every collection site of this study was associated with high human activity, posting of warning signs, monitoring of recreational water sources, and awareness of high-risk people are of utmost importance. To the best of our knowledge, the present research is the first to report T2 genotype from geothermal water sources in Iran. PMID:27127409

  17. Superdiffusion dominates intracellular particle motion in the supercrowded cytoplasm of pathogenic Acanthamoeba castellanii

    NASA Astrophysics Data System (ADS)

    Reverey, Julia F.; Jeon, Jae-Hyung; Bao, Han; Leippe, Matthias; Metzler, Ralf; Selhuber-Unkel, Christine

    2015-06-01

    Acanthamoebae are free-living protists and human pathogens, whose cellular functions and pathogenicity strongly depend on the transport of intracellular vesicles and granules through the cytosol. Using high-speed live cell imaging in combination with single-particle tracking analysis, we show here that the motion of endogenous intracellular particles in the size range from a few hundred nanometers to several micrometers in Acanthamoeba castellanii is strongly superdiffusive and influenced by cell locomotion, cytoskeletal elements, and myosin II. We demonstrate that cell locomotion significantly contributes to intracellular particle motion, but is clearly not the only origin of superdiffusivity. By analyzing the contribution of microtubules, actin, and myosin II motors we show that myosin II is a major driving force of intracellular motion in A. castellanii. The cytoplasm of A. castellanii is supercrowded with intracellular vesicles and granules, such that significant intracellular motion can only be achieved by actively driven motion, while purely thermally driven diffusion is negligible.

  18. Superdiffusion dominates intracellular particle motion in the supercrowded cytoplasm of pathogenic Acanthamoeba castellanii.

    PubMed

    Reverey, Julia F; Jeon, Jae-Hyung; Bao, Han; Leippe, Matthias; Metzler, Ralf; Selhuber-Unkel, Christine

    2015-01-01

    Acanthamoebae are free-living protists and human pathogens, whose cellular functions and pathogenicity strongly depend on the transport of intracellular vesicles and granules through the cytosol. Using high-speed live cell imaging in combination with single-particle tracking analysis, we show here that the motion of endogenous intracellular particles in the size range from a few hundred nanometers to several micrometers in Acanthamoeba castellanii is strongly superdiffusive and influenced by cell locomotion, cytoskeletal elements, and myosin II. We demonstrate that cell locomotion significantly contributes to intracellular particle motion, but is clearly not the only origin of superdiffusivity. By analyzing the contribution of microtubules, actin, and myosin II motors we show that myosin II is a major driving force of intracellular motion in A. castellanii. The cytoplasm of A. castellanii is supercrowded with intracellular vesicles and granules, such that significant intracellular motion can only be achieved by actively driven motion, while purely thermally driven diffusion is negligible.

  19. Distribution of Acanthamoeba Genotypes Isolated from Recreational and Therapeutic Geothermal Water Sources in Southwestern Iran.

    PubMed

    Niyyati, Maryam; Saberi, Reza; Latifi, Alireza; Lasjerdi, Zohreh

    2016-01-01

    A comprehensive survey was conducted along 10 km of geothermal rivers in southwestern Iran. A total of 40 water samples were tested for the presence of Acanthamoeba spp., and genotypes were determined by targeting the diagnostic fragment 3 region of the 18S rRNA gene. The pathogenic potential of all positive isolates was also identified using tolerance ability test. High occurrences of Acanthamoeba (50%) were detected in the sampling areas. Based on sequencing analysis, isolates belonging to T4 (93.7%) and T2 (6.25%) genotypes were reported. Thermo- and osmotolerance tests revealed that five strains are highly pathogenic. Since every collection site of this study was associated with high human activity, posting of warning signs, monitoring of recreational water sources, and awareness of high-risk people are of utmost importance. To the best of our knowledge, the present research is the first to report T2 genotype from geothermal water sources in Iran.

  20. Assessment of the efficacy of benzalkonium chloride and sodium hypochlorite against Acanthamoeba polyphaga and Tetrahymena spp.

    PubMed

    Vaerewijck, M J M; Sabbe, K; Baré, J; Spengler, H-P; Favoreel, H W; Houf, K

    2012-03-01

    The efficacy of benzalkonium chloride and sodium hypochlorite against Acanthamoeba polyphaga and two Tetrahymena spp. was determined based on the European Standard EN 1276:2009 suspension test. Trophozoite viability was assessed by determination of the membrane integrity using flow cytometry as a fast screening technique. Bovine serum albumin was added to simulate clean (0.3 g/liter) and dirty (3 g/liter) conditions. Benzalkonium chloride caused cell lysis at concentrations above 50 mg/liter under clean and dirty conditions. A concentration of 50 mg of free chlorine per liter had a strong biocidal effect on acanthamoebae and tetrahymenae after 15 min under clean and dirty conditions. Our results suggest that benzalkonium chloride and sodium hypochlorite were effective against the three microorganisms at concentrations commonly applied in the food industry. PMID:22410229

  1. Effect of Parachlamydia acanthamoebae on pulmonary function parameters in a bovine respiratory model.

    PubMed

    Lohr, M; Prohl, A; Ostermann, C; Diller, R; Greub, G; Reinhold, P

    2016-07-01

    The aim of this study was to evaluate pulmonary dysfunction induced by experimental infection with Parachlamydia acanthamoebae in calves. Intrabronchial inoculation with P. acanthamoebae was performed in 31 calves aged 2-3 months old at two different challenge doses of 10(8) and 10(10) inclusion-forming units (IFU) per animal. Control animals received heat inactivated bacteria. The effects on pulmonary gas exchange were determined by arterial blood gas analysis and haemoximetry during the 7 days post inoculation (DPI). For pulmonary function testing (PFT), impulse oscillometry, capnography, and measurement of O2 uptake were undertaken in spontaneously breathing animals 7 and 3 days before inoculation and were repeated until 10 DPI. In the early phase after challenge (1-3 DPI), mild hypoxaemia occurred, which was accompanied by a significant reduction in both tidal and alveolar volumes (each related to bodyweight, BW). In parallel, expiratory flow rate and specific ventilation (i.e. minute ventilation related to O2 uptake) were significantly increased. Minute and alveolar ventilations (each related to metabolic BW) increased significantly due to higher respiratory rates, lasting until 4 and 5 DPI, respectively. Oxygen uptake was slightly reduced during the first 2 days after challenge, but increased significantly during the recovery phase, from 4 to 8 DPI. No deterioration in respiratory mechanics or acid-base balance was observed. Respiratory infection with 10(10) IFU P. acanthamoebae per calf induced mild respiratory dysfunction, mainly characterised by hypoxaemia. The study's findings do not indicate severe pathophysiological consequences of P. acanthamoebae infection on pulmonary function in the bovine host. PMID:27240907

  2. Survival and transfer ability of phylogenetically diverse bacterial endosymbionts in environmental Acanthamoeba isolates.

    PubMed

    Matsuo, Junji; Kawaguchi, Kouhei; Nakamura, Shinji; Hayashi, Yasuhiro; Yoshida, Mitsutaka; Takahashi, Kaori; Mizutani, Yoshihiko; Yao, Takashi; Yamaguchi, Hiroyuki

    2010-08-01

    Obligate intracellular bacteria are commonly found as endosymbionts of acanthamoebae; however, their survival in and ability to transfer to amoebae are currently uncharacterized. In this study, six bacterial endosymbionts, found in five environmental Acanthamoeba isolates (S13, R18, S23, S31, S40) from different locations of Sapporo city, Japan, were characterized. Phylogenetic analysis revealed that three bacterial endosymbionts (eS23, eS31, eS40a) belonged to α- and β-Proteobacteria phyla and the remaining endosymbionts (eS13, eR18, eS40b) belonged to the order Chlamydiales. The Acanthamoeba isolate (S40) contained two phylogenetically different bacterial endosymbionts (eS40a, eS40b). Fluorescent in situ hybridization analysis showed that all bacterial endosymbionts were diffusely localized within amoebae. Transmission electron microscopy also showed that the endosymbionts were rod-shaped (eS23, eS31, eS40a) or sphere- or crescent-shaped (eS13, eR18, eS40b). No successful culture of these bacteria was achieved using conventional culture methods, but the viability of endosymbionts was confirmed by live/dead staining and RT-PCR methods. However, endosymbionts (except eR18) derived from original host cells lost the ability to be transferred to another Acanthamoebae strains [ATCC strain (C3), environmental strains (S14, R23, S24)]. Thus, our data demonstrate that phylogenetically diverse bacterial endosymbionts found in amoebae maintain a stable interaction with amoebae, but the transferability is limited.

  3. Identification and properties of proteases from an Acanthamoeba isolate capable of producing granulomatous encephalitis

    PubMed Central

    Sissons, James; Alsam, Selwa; Goldsworthy, Graham; Lightfoot, Mary; Jarroll, Edward L; Khan, Naveed Ahmed

    2006-01-01

    Background Granulomatous amoebic encephalitis due to Acanthamoeba is often a fatal human disease. However, the pathogenesis and pathophysiology of Acanthamoeba encephalitis remain unclear. In this study, the role of extracellular Acanthamoeba proteases in central nervous system pathogenesis and pathophysiology was examined. Results Using an encephalitis isolate belonging to T1 genotype, we observed two major proteases with approximate molecular weights of 150 KD and 130 KD on SDS-PAGE gels using gelatin as substrate. The 130 KD protease was inhibited with phenylmethylsulfonyl fluoride (PMSF) suggesting that it is a serine protease, while the 150 KD protease was inhibited with 1, 10-phenanthroline suggesting that it is a metalloprotease. Both proteases exhibited maximal activity at neutral pH and over a range of temperatures, indicating their physiological relevance. These proteases degrade extracellular matrix (ECM), which provide structural and functional support to the brain tissue, as shown by the degradation of collagen I and III (major components of collagenous ECM), elastin (elastic fibrils of ECM), plasminogen (involved in proteolytic degradation of ECM), as well as casein and haemoglobin. The proteases were purified partially using ion-exchange chromatography and their effects were tested in an in vitro model of the blood-brain barrier using human brain microvascular endothelial cells (HBMEC). Neither the serine nor the metalloprotease exhibited HBMEC cytotoxicity. However, the serine protease exhibited HBMEC monolayer disruptions (trypsin-like) suggesting a role in blood-brain barrier perturbations. Conclusion Overall, these data suggest that Acanthamoeba proteases digest ECM, which may play crucial role(s) in invasion of the brain tissue by amoebae. PMID:16672059

  4. Identification of causative pathogens in mouse eyes with bacterial keratitis by sequence analysis of 16S rDNA libraries

    PubMed Central

    Song, Hong-Yan; Qiu, Bao-Feng; Liu, Chun; Zhu, Shun-Xing; Wang, Sheng-Cun; Miao, Jin; Jing, Jing; Shao, Yi-Xiang

    2014-01-01

    The clone library method using PCR amplification of the 16S ribosomal RNA (rRNA) gene was used to identify pathogens from corneal scrapings of C57BL/6-corneal opacity (B6-Co) mice with bacterial keratitis. All 10 samples from the eyes with bacterial keratitis showed positive PCR results. All 10 samples from the normal cornea showed negative PCR results. In all 10 PCR-positive samples, the predominant and second most predominant species accounted for 20.9 to 40.6% and 14.7 to 26.1%, respectively, of each clone library. The predominant species were Staphylococcus lentus, Pseudomonas aeruginosa, and Staphylococcus epidermidis. The microbiota analysis detected a diverse group of microbiota in the eyes of B6-Co mice with bacterial keratitis and showed that the causative pathogens could be determined based on percentages of bacterial species in the clone libraries. The bacterial species detected in this study were mostly in accordance with results of studies on clinical bacterial keratitis in human eyes. Based on the results of our previous studies and this study, the B6-Co mouse should be considered a favorable model for studying bacterial keratitis. PMID:25312507

  5. Anaerobic respiration: In vitro efficacy of Nitazoxanide against mitochondriate Acanthamoeba castellanii of the T4 genotype.

    PubMed

    Aqeel, Yousuf; Siddiqui, Ruqaiyyah; Farooq, Maria; Khan, Naveed Ahmed

    2015-10-01

    Acanthamoeba is an opportunistic protist pathogen that is responsible for serious human and animal infection. Being one of the most frequently isolated protists from the environment, it is likely that it readily encounters microaerophilic environments. For respiration under anaerobic or low oxygen conditions in several amitochondriate protists, decarboxylation of pyruvate is catalyzed by pyruvate ferredoxin oxidoreductase instead of pyruvate dehydrogenase. In support, Nitazoxanide, an inhibitor of pyruvate ferredoxin oxidoreductase, is effective and non-mutagenic clinically against a range of amitochondriate protists, Giardia intestinalis, Entamoeba histolytica and Trichomonas vaginalis. The overall aim of the present study was to determine in vitro efficacy of Nitazoxanide against Acanthamoeba castellanii. At micromolar concentrations, the findings revealed that Nitazoxanide neither affected A. castellanii growth or viability nor amoeba-mediated host cell monolayer damage in vitro or extracellular proteolytic activities. Similarly, microaerophilic conditions alone had no significant effects. In contrast, microaerophilic conditions together with Nitazoxanide showed amoebicidal effects and inhibited A. castellanii-mediated host cell monolayer damage as well as extracellular proteases. Using encystation assays, it was observed that Nitazoxanide inhibited trophozoite transformation into cysts both under aerophilic and microaerophilic conditions. Furthermore, pre-treatment of cysts with Nitazoxanide inhibited A. castellanii excystation. These findings are important in the identification of potential targets that could be useful against parasite-specific respiration as well as to understand the basic biology of the life cycle of Acanthamoeba.

  6. Prevalence of acanthamoeba from tap water in rio grande do Sul, Brazil.

    PubMed

    Winck, Mari Aline Todero; Caumo, Karin; Rott, Marilise Brittes

    2011-11-01

    A total of 136 samples of tap water were collected from state and municipal schools between March and November 2009. The samples were filtered through cellulose nitrate membranes that were seeded at non-nutrient agar 1.5% containing an overlayer of Escherichia coli suspension. Thirty-one (22.79%) tap water samples investigated were found positive for free-living amoebae (FLA). From these, 13 presented as FLA that seems to belong to the genus Acanthamoeba. All samples of FLA were cloned and identified as belonging to the genus Acanthamoeba by the morphology of cysts and trophozoites and by PCR using genus-specific primers that amplify the ASA.S1 region of 18S rDNA gene. Physiological tests of thermotolerance and osmotolerance were used to evaluate the pathogenicity of the isolates. The sequencing analysis by comparing the sequences submitted to GenBank, showed genotype distribution into groups T2, T2/T6, T6, and T4. In tests of thermotolerance and osmotolerance, 50% of the isolates had a low pathogenic potential. The results indicated the presence of Acanthamoeba in tap water in Rio Grande do Sul, Brazil, revealing its importance and the need for more epidemiological studies to determine their distribution in the environment and its pathogenic potential.

  7. The role of actin in the temperature-dependent gelation and contraction of extracts of Acanthamoeba

    PubMed Central

    1976-01-01

    The temperature-dependent assembly and the interaction of Acanthamoeba contractile proteins have been studied in a crude extract. A cold extract of soluble proteins from Acanthamoeba castellanii is prepared by homogenizing the cells in a sucrose-ATP-ethyleneglycol-bis-(beta- aminoethyl ether) N,N'-tetraacetic acid buffer and centrifuging at 136,000 g for 1 h. When this supernate of soluble proteins is warmed to room temperature, it forms a solid gel. Upon standing at room temperature, the gel slowly contracts and squeezes out soluble components. The rates of gelation and contraction are both highly temperature dependent, with activation energies of about 20 kcal per mol. Gel formation is dependent upon the presence of ATP and Mg++. Low concentrations of Ca++ accelerate the contractile phase of this phenomenon. The major protein component of the gel is actin. It is associated with myosin, cofactor, a high molecular weight protein tentatively identfied as actin-binding protein, and several other unidentified proteins. Actin has been purified from these gels and was found to be capable of forming a solid gel when polymerized in the presence of ATP, MgCl3, and KCL. The rate of purified actin polymerication is very temperature dependent and is accelerated by the addition of fragments of muscle actin filaments. These data suggest that Acanthamoeba contractile proteins have a dual role in the cell; they may generate the forces for cellular movements and also act as cytoskeletal elements by controlling the consistency of the cytoplasm. PMID:1030705

  8. Prevalence of Acanthamoeba and superbugs in a clinical setting: coincidence or hyperparasitism?

    PubMed

    Siddiqui, Ruqaiyyah; Sagheer, Mehwish; Khan, Naveed Ahmed

    2013-03-01

    Antibacterial strategies to eradicate superbugs from hospitals/nursing homes have had limited success, suggesting the need for employing innovative preventative measures and better understanding of the prevalence of microbial pathogens in close proximity of susceptible populations. A total of 120 environmental samples were collected from the Aga Khan University hospital. Amoebae were identified using morphological characteristics as well as PCR using genus-specific primers, while bacteria were identified using standard biochemical testing. Out of 120 samples tested, 52 (43.3 %) samples were positive for Acanthamoeba, while all 120 (100 %) samples were positive for bacteria. Following bacterial identification, samples showed mixed bacterial populations. Out of 120 samples, 76 (63.3 %) samples were positive for Bacillus spp., 64 (53.3 %) samples were positive for Corynebacterium spp., 32 (26.6 %) samples were positive for Staphylococcus spp., and 9 (7.5 %) samples were positive for Micrococcus spp. The antibiotic susceptibility showed that all bacterial isolates recovered were multiple drug-resistant. The current findings suggest that Acanthamoeba and bacteria coexist in a clinical environment. Given that Acanthamoeba can harbor bacteria, anti-amoebic approaches may represent a strategy in eradicating "superbugs" from the clinical setting in addition to the current measures.

  9. Expression levels of encystation mediating factors in fresh strain of Acanthamoeba castellanii cyst ESTs.

    PubMed

    Moon, Eun-Kyung; Chung, Dong-Il; Hong, Yeonchul; Kong, Hyun-Hee

    2011-04-01

    The life cycle of Acanthamoeba consists of two stages, trophozoite and cyst. The cyst form is resistant to almost all antibiotics. By long term cultivation, Acanthamoeba severely attenuated the encysting ability. To determine the changing of gene expression by the long term cultivation, especially focusing an encystation mediating factors, this study compared the ESTs of the fresh strain and the old strain, and trophozoite. Comparison of the KOG (euKaryotic Orthologous Groups) analysis relative to trophozoite revealed higher percentages of cyst ESTs related to G (Carbohydrate transport and metabolism), H (Coenzyme transport and metabolism), I (Lipid transport and metabolism), D (Cell cycle control, cell division, chromosome partitioning), T (signal transduction mechanisms), and O (Posttranslational modification, protein turnover, chaperones). In addition to this result, KOG analysis of fresh strain relative to old strain showed higher percentage of cyst ESTs related to metabolism category and T (signal transduction mechanisms) article. ESTs of the fresh strain revealed more various gene profiles compared to the old strain including encystation mediating factors like autophagy related proteins (Z article) and signal transduction proteins (T article). Twenty seven kinds of protein kinase C (PKC) like genes were detected in cyst or trophozoite ESTs and twenty one of them were highly expressed during encystation. The information of the expressed genes during encystation in only the fresh strain will provide new clues to understanding the encystation mechanism of encysting protozoa including Acanthamoeba. PMID:21276446

  10. Necrotizing meningoencephalitis and pneumonitis in a simian immunodeficiency virus-infected rhesus macaque due to Acanthamoeba.

    PubMed

    Westmoreland, S V; Rosen, J; MacKey, J; Romsey, C; Xia, D-L; Visvesvera, G S; Mansfield, K G

    2004-07-01

    Free-living amoebae of the genus Acanthamoeba can cause a fatal disease of the brain in humans called granulomatous amoebic encephalitis. We present a case of meningoencephalitis and pneumonitis in a simian immunodeficiency virus (SIV)-infected rhesus macaque caused by Acanthamoeba sp. The animal became ill 176 days after intravenous inoculation with SIVmac251 after a short history of weight loss and a sudden onset of hind limb paresis and abnormal head movements. Histopathologic examination of hematoxylin and eosin-stained tissues revealed multifocal to coalescing necrotizing neutrophilic meningoencephalitis and pneumonitis. Immunofluorescence and polymerase chain reaction were used to identify the genus of amoeba as Acanthamoeba. Immunohistochemistry of immune cell markers was used to characterize the animal's immune response to the opportunistic amoebic infection with features of both innate and adaptive cell-mediated immunity. Although not previously reported, the potential transmission to humans, either through environmental contamination or contact with an infected animal, makes this disease a threat to laboratory animal care staff and pathologists. PMID:15232140

  11. Acanthamoeba meningoencephalitis in immunocompetent: A case report and review of literature

    PubMed Central

    Khanna, Vinay; Shastri, BA; Anusha, G; Mukhopadhayay, Chiranjay; Khanna, Ruchee

    2014-01-01

    A 30-year-old manual laborer from Karnataka, India presented with intermittent low grade fever and diffuse headache for 1 month. On examination, patient had enlarged supraclavicular and cervical lymph nodes. Patient had positive Kernig's sign and neck stiffness. Motor, sensory and cranial nerve examinations were within the normal limits. Abdominal, cardiovascular and chest examination did not yield any positive findings. Contrast enhanced computed tomography head was normal. Patient was suspected to have extrapulmonary tuberculosis. Patient was started on antitubercular drugs. Diagnostic lumbar puncture was performed. Wet mount and Giemsa smear preparation of cerebrospinal fluid (CSF) showed trophozoites suggestive of Acanthamoeba. CSF was cultured onto non-nutrient agar with an overlay of Escherichia coli. Wet mount made from the culture media yielded cysts and trophozoites of Acanthamoeba spp. Patient was diagnosed with Acanthamoeba meningitis and was started on specific therapy with Rifampicin 600 mg once a day, Cotrimoxazole 960 mg twice-a-day and Fluconazole 400 mg once daily for 2 weeks. Patient had a complete recovery and was discharged from the hospital. PMID:25250233

  12. Seroprevalence of Acanthamoeba Antibodies in Rheumatoid Arthritis Patients by IFAT, Tehran, Iran 2007

    PubMed Central

    Eftekhar, M; Athari, A; Haghighi, A; Mosaffa, N; Shahram, F; Abadi, A

    2010-01-01

    Background This preliminary study was conducted to discriminate the prevalence of Acanthamoeba antibodies in rheumatoid arthritis (RA) patients and healthy controls to analyze the correlation between these two groups. Methods From October 2006 to August 2007 a total of 121 serum samples from RA patients attending the Rheumatolgy Department at Shariati Hospital in Tehran were obtained and stored at -20°C until using by indirect fluorescent-antibody test (IFAT). RA was diagnosed according to the American Collage of Rheumatology classification criteria. The organism used in this study was isolated from various water resources in Tehran, Iran cultured axenically and then went on a PCR assay based on 18S rRNA to identify the genus Acanthomoeba. Indirect immunofluorescence antibody (IFA) staining of serum samples was carried out to detect anti Acanthomoeba antibodies. Results In culture, out of 22 samples, 13(59%) were grown in xenic but only two in axenic medium. PCR amplified a 904bp fragment, specific for Acanthamoeba. Of examined serum samples, Acanthamoeba antibodies were present in 70 (57.8%) and 52 (41.2%), respectively. The highest titer of antibodies (1:320) was detected in one patient with RA. Conclusion Our study supports the hypothesis that some parasitic microorganisms can involve and contribute toward the development of rheumatoid syndromes. PMID:22347233

  13. Targeting cyst wall is an effective strategy in improving the efficacy of marketed contact lens disinfecting solutions against Acanthamoeba castellanii cysts.

    PubMed

    Abjani, Farhat; Khan, Naveed Ahmed; Yousuf, Farzana Abubakar; Siddiqui, Ruqaiyyah

    2016-06-01

    Acanthamoeba cysts are highly resistant to contact lens disinfecting solutions. Acanthamoeba cyst wall is partially made of 1,4 β-glucan (i.e., cellulose) and other complex polysaccharides making it a hardy shell that protects the resident amoeba. Here, we hypothesize that targeting the cyst wall structure in addition to antiamoebic compound would improve the efficacy of marketed contact lens disinfecting solutions. Using chlorhexidine as an antiamoebic compound and cellulase enzyme to disrupt cyst wall structure, the findings revealed that combination of both agents abolished viability of Acanthamoeba castellanii cysts and trophozoites. When tested alone, none of the agents nor contact lens disinfecting solutions completely destroyed A. castellanii cysts and trophozoites. The absence of cyst wall-degrading enzymes in marketed contact lens disinfecting solutions render them ineffective against Acanthamoeba cysts. It is concluded that the addition of cyst wall degrading molecules in contact lens disinfecting solutions will enhance their efficacy in decreasing the incidence of Acanthamoeba effectively. PMID:26675112

  14. Targeting cyst wall is an effective strategy in improving the efficacy of marketed contact lens disinfecting solutions against Acanthamoeba castellanii cysts.

    PubMed

    Abjani, Farhat; Khan, Naveed Ahmed; Yousuf, Farzana Abubakar; Siddiqui, Ruqaiyyah

    2016-06-01

    Acanthamoeba cysts are highly resistant to contact lens disinfecting solutions. Acanthamoeba cyst wall is partially made of 1,4 β-glucan (i.e., cellulose) and other complex polysaccharides making it a hardy shell that protects the resident amoeba. Here, we hypothesize that targeting the cyst wall structure in addition to antiamoebic compound would improve the efficacy of marketed contact lens disinfecting solutions. Using chlorhexidine as an antiamoebic compound and cellulase enzyme to disrupt cyst wall structure, the findings revealed that combination of both agents abolished viability of Acanthamoeba castellanii cysts and trophozoites. When tested alone, none of the agents nor contact lens disinfecting solutions completely destroyed A. castellanii cysts and trophozoites. The absence of cyst wall-degrading enzymes in marketed contact lens disinfecting solutions render them ineffective against Acanthamoeba cysts. It is concluded that the addition of cyst wall degrading molecules in contact lens disinfecting solutions will enhance their efficacy in decreasing the incidence of Acanthamoeba effectively.

  15. Glycyrrhizin Reduces HMGB1 and Bacterial Load in Pseudomonas aeruginosa Keratitis

    PubMed Central

    Ekanayaka, Sandamali A.; McClellan, Sharon A.; Barrett, Ronald P.; Kharotia, Shikhil; Hazlett, Linda D.

    2016-01-01

    Purpose High mobility group box 1 (HMGB1) contributes to poor disease outcome in Pseudomonas aeruginosa keratitis. This study tests the prophylactic effect of treatment with HMGB1 inhibitors, glycyrrhizin (GLY) and its derivative, carbenoxolone (CBX), for Pseudomonas keratitis. Methods We treated C57BL/6 (B6) mice subconjunctivally with GLY or CBX, infected with a noncytotoxic clinical isolate (KEI 1025) or a cytotoxic strain (ATCC 19660) of P. aeruginosa, and injected intraperitoneally with either agent. Clinical score, photography with a slit lamp, real-time RT-PCR, ELISA, myeloperoxidase (MPO) assay, bacterial plate count, histopathology, and absorbance assays were used to assess treatment efficacy and bacteriostatic activity. Results After KEI 1025 infection, GLY treatment reduced HMGB1 (mRNA and protein levels) and improved disease outcome with significant reduction in mRNA levels of IL-1β, TLR4, CXCL2, and IL-12; protein expression (IL-1β, CXCL2); neutrophil infiltrate; and bacterial load. Treatment with GLY enhanced antimicrobial proteins, including CRAMP and mBD2, but not mBD3. Glycyrrhizin also reduced clinical scores and improved disease outcome in corneas infected with strain 19660. However, neither HMGB1 mRNA or protein levels were reduced, but rather, CXCL2 expression (mRNA and protein), neutrophil infiltrate, and bacterial load were reduced statistically. Treatment with GLY initiated 6 hours after infection reduced plate count; GLY also was bacteriostatic for KEI 1025 and ATCC 19660. Conclusions Glycyrrhizin reduces HMGB1 and is protective against P. aeruginosa–induced keratitis with a clinical isolate that is noncytotoxic. It was similar, but less effective when used after infection with a cytotoxic strain, which did not reduce HMGB1. PMID:27792814

  16. Impact of Microbiota on Resistance to Ocular Pseudomonas aeruginosa-Induced Keratitis

    PubMed Central

    Kugadas, Abirami; Christiansen, Stig Hill; Kunz, Ryan; Fichorova, Raina; Gadjeva, Mihaela

    2016-01-01

    The existence of the ocular microbiota has been reported but functional analyses to evaluate its significance in regulating ocular immunity are currently lacking. We compared the relative contribution of eye and gut commensals in regulating the ocular susceptibility to Pseudomonas aeruginosa–induced keratitis. We find that in health, the presence of microbiota strengthened the ocular innate immune barrier by significantly increasing the concentrations of immune effectors in the tear film, including secretory IgA and complement proteins. Consistent with this view, Swiss Webster (SW) mice that are typically resistant to P. aeruginosa–induced keratitis become susceptible due to the lack of microbiota. This was exemplified by increased corneal bacterial burden and elevated pathology of the germ free (GF) mice when compared to the conventionally maintained SW mice. The protective immunity was found to be dependent on both eye and gut microbiota with the eye microbiota having a moderate, but significant impact on the resistance to infection. These events were IL-1ß–dependent as corneal IL-1ß levels were decreased in the infected GF and antibiotic-treated mice when compared to the SPF controls, and neutralization of IL-1ß increased the ocular bacterial burden in the SPF mice. Monocolonizing GF mice with Coagulase Negative Staphylococcus sp. isolated from the conjunctival swabs was sufficient to restore resistance to infection. Cumulatively, these data underline a previously unappreciated role for microbiota in regulating susceptibility to ocular keratitis. We predict that these results will have significant implications for contact lens wearers, where alterations in the ocular commensal communities may render the ocular surface vulnerable to infections. PMID:27658245

  17. Association Between In Vitro Susceptibility to Natamycin and Voriconazole and Clinical Outcomes in Fungal Keratitis

    PubMed Central

    Sun, Catherine Q.; Lalitha, Prajna; Prajna, N. Venkatesh; Karpagam, Rajarathinam; Geetha, Manoharan; O’Brien, Kieran S.; Oldenburg, Catherine E.; Ray, Kathryn J.; McLeod, Stephen D.; Acharya, Nisha R.; Lietman, Thomas M.

    2014-01-01

    Purpose To assess the association between minimum inhibitory concentration (MIC) and clinical outcomes in a fungal keratitis clinical trial. Design Experimental study using data from a randomized comparative trial. Participants Of the 323 patients enrolled in the trial, we were able to obtain MIC values from 221 patients with monocular fungal keratitis. Methods The Mycotic Ulcer Treatment Trial I (MUTT I) was a randomized, double-masked clinical trial comparing clinical outcomes of monotherapy with topical natamycin versus voriconazole for the treatment of fungal keratitis. Speciation and determination of MIC to natamycin and voriconazole were performed according to Clinical and Laboratory Standards Institute guidelines. The relationship between MIC and clinical outcome was assessed. Main Outcome Measures The primary outcome was 3-month best spectacle-corrected visual acuity. Secondary outcomes included 3-month infiltrate/scar size, corneal perforation and/or therapeutic penetrating keratoplasty (TPK), and time to re-epithelialization. Results A 2-fold increase in MIC was associated with a larger 3-month infiltrate/scar size (0.21 mm, 95% confidence interval [CI] 0.10–0.31, P <0.001) and increased odds of perforation (odds ratio [OR] 1.32, 95% CI 1.04–1.69, P=0.02). No correlation was found between MIC and 3-month visual acuity. For natamycin-treated cases, an association was found between higher natamycin MIC with larger 3-month infiltrate/scar size (0.29 mm, 95% CI 0.15–0.43, P<0.001) and increased perforations (OR 2.41, 95% CI 1.46–3.97, P<0.001). Among voriconazole-treated cases, the voriconazole MIC did not correlate with any of the measured outcomes in the study. Conclusion Decreased susceptibility to natamycin was associated with increased infiltrate/scar size and increased odds of perforation. There was no association between susceptibility to voriconazole and outcome. PMID:24746358

  18. Prevalence of Acanthamoeba spp. and other free-living amoebae in household water, Ohio, USA--1990-1992.

    PubMed

    Stockman, Lauren J; Wright, Carolyn J; Visvesvara, Govinda S; Fields, Barry S; Beach, Michael J

    2011-03-01

    Knowledge of the prevalence of free-living amoebae (FLA) in US household water can provide a focus for prevention of amoeba-associated illnesses. Household water samples from two Ohio counties, collected and examined for amoebae during 1990-1992, were used to describe the prevalence of Acanthamoeba and other FLA in a household setting. Amoebae were isolated and identified by morphologic features. A total of 2,454 samples from 467 households were examined. Amoebae were found in water samples of 371 (79%) households. Sites most likely to contain amoeba were shower heads (52%) and kitchen sprayers (50%). Species of Hartmannella, Acanthamoeba, or Vahlkampfia were most common. Detection was higher in biofilm swab samples than in water samples. Detection of FLA and Acanthamoeba, at 79% and 51%, respectively, exceed estimates that have been published in previous surveys of household sources. We believe FLA are commonplace inhabitants of household water in this sample as they are in the environment.

  19. Acanthamoeba encystment: multifactorial effects of buffers, biocides, and demulcents present in contact lens care solutions

    PubMed Central

    Kovacs, Christopher J; Lynch, Shawn C; Rah, Marjorie J; Millard, Kimberly A; Morris, Timothy W

    2015-01-01

    Purpose To determine whether agents which are purportedly capable of inducing encystment of Acanthamoeba can recapitulate the signal when tested in differing formulations. Methods In accordance with the International Standard ISO 19045, Acanthamoeba castellanii ATCC 50370 trophozoites were cultured in antibiotic-free axenic medium, treated with test solutions, and encystment rates plus viability were measured via bright field and fluorescent microscopy. Test solutions included phosphate-buffered saline (PBS), borate-buffered saline, biguanide- and hydrogen peroxide (H2O2)-based biocides, propylene glycol (PG) and povidone (POV) ophthalmic demulcents, and one-step H2O2-based contact lens disinfection systems. Results Only PBS solutions with 0.25 ppm polyaminopropyl biguanide (PAPB) and increasing concentrations of PG and POV stimulated A. castellanii encystment in a dose-dependent manner, whereas PBS solutions containing 3% H2O2 and increasing concentrations of PG and POV did not stimulate encystment. Borate-buffered saline and PBS/citrate solutions containing PG also did not stimulate encystment. In addition, no encystment was observed after 24 hours, 7 days, or 14 days of exposures of trophozoites to one-step H2O2 contact lens disinfection products or related solutions. Conclusion The lack of any encystment observed when trophozoites were treated with existing or new one-step H2O2 contact lens care products, as well as when trophozoites were exposed to various related test solutions, confirms that Acanthamoeba encystment is a complex process which depends upon simultaneous contributions of multiple factors including buffers, biocides, and demulcents. PMID:26508829

  20. Meningoencephalitis due to Acanthamoeba SP. Pathogenesis and clinico-pathological study.

    PubMed

    Martínez, A J; Sotelo-Avila, C; Garcia-Tamayo, J; Morón, J T; Willaert, E; Stamm, W P

    1977-03-31

    Amebic Meningoencephalitis (AM) and Primary Amebic Meningoencephalitis (PAM) are infectious diseases essentially confined to the Central Nervous System (CNS) and caused by free-living amebas of the genus Acanthamoeba (A.) and Naegleria (N.) respectively. AM due to A. sp. (Acanthamoeba castellanii and Acanthamoeba culbertsoni) have been reported in chronically ill debilitated individuals, some of them under immunosuppressive therapy, or in immunologically impaired patients without a history of recent swimming in contrast to cases due to N. sp. which usually occurs in healthy, young individuals with a recent history of swimming in man-made lakes or heated swimming pools. AM due to A.sp. is characterized by a subacute or chronic granulomatous meningoencephalitis involving mainly the midbrain, basal areas of the temporal and occipital lobes and posterior fossa structures. CNS lesions in AM are perhaps secondary and the portal of entry in humans is probably from the lower respiratory tract, genitourinary system or skin reaching the CNS by hematogenous spread. The predominant host reaction is usually composed of lymphocytes, plasma cells, monocytes and multinucleated foreign body giant cells. Necrosis is moderate and hemorrhage scant or absent. Cysts as well as trophozoites may be seen within the CNS lesions. PAM is due to Naegleria fowleri and is characterized by an hemorrhagic necrotizing meningoencephalities with an acute inflammatory response. Only trophozoites are found in lesions. The portal of entry is through the olfactory neuroepithelium. CNS tissues fixed in formalin may be used for further identification and taxonomical classification of the causative protoza using immunofluorescent antibody techniques (IFAT) and electron microscopic methods. PMID:857580

  1. Interaction of Escherichia coli K1 and K5 with Acanthamoeba castellanii Trophozoites and Cysts

    PubMed Central

    Matin, Abdul

    2011-01-01

    The existence of symbiotic relationships between Acanthamoeba and a variety of bacteria is well-documented. However, the ability of Acanthamoeba interacting with host bacterial pathogens has gained particular attention. Here, to understand the interactions of Escherichia coli K1 and E. coli K5 strains with Acanthamoeba castellanii trophozoites and cysts, association assay, invasion assay, survival assay, and the measurement of bacterial numbers from cysts were performed, and nonpathogenic E. coli K12 was also applied. The association ratio of E. coli K1 with A. castellanii was 4.3 cfu per amoeba for 1 hr but E. coli K5 with A. castellanii was 1 cfu per amoeba for 1 hr. By invasion and survival assays, E. coli K5 was recovered less than E. coli K1 but still alive inside A. castellanii. E. coli K1 and K5 survived and multiplied intracellularly in A. castellanii. The survival assay was performed under a favourable condition for 22 hr and 43 hr with the encystment of A. castellanii. Under the favourable condition for the transformation of trophozoites into cysts, E. coli K5 multiplied significantly. Moreover, the pathogenic potential of E. coli K1 from A. castellanii cysts exhibited no changes as compared with E. coli K1 from A. castellanii trophozoites. E. coli K5 was multiplied in A. castellanii trophozoites and survived in A. castellanii cysts. Therefore, this study suggests that E. coli K5 can use A. castellanii as a reservoir host or a vector for the bacterial transmission. PMID:22355201

  2. Excimer Laser Phototherapeutic Keratectomy for the Treatment of Clinically Presumed Fungal Keratitis

    PubMed Central

    Li, Liang-Mao; Zhao, Li-Quan; Qu, Ling-Hui; Li, Peng

    2014-01-01

    This retrospective study was to evaluate treatment outcomes of excimer laser phototherapeutic keratectomy (PTK) for clinically presumed fungal keratitis. Forty-seven eyes of 47 consecutive patients underwent manual superficial debridement and PTK. All corneal lesions were located in the anterior stroma and were resistant to medication therapy for at least one week. Data were collected by a retrospective chart review with at least six months of follow-up data available. After PTK, infected corneal lesions were completely removed and the clinical symptoms resolved in 41 cases (87.2%). The mean ablation depth was 114.39 ± 45.51 μm and diameter of ablation was 4.06 ± 1.07 mm. The mean time for healing of the epithelial defect was 8.8 ± 5.6 days. Thirty-four eyes (82.9%) showed an improvement in best spectacle-corrected visual acuity of two or more lines. PTK complications included mild to moderate corneal haze, hyperopic shift, irregular astigmatism, and thinning cornea. Six eyes (12.8%) still showed progressed infection, and conjunctival flap covering, amniotic membrane transplantation, or penetrating keratoplasty were given. PTK is a valuable therapeutic alternative for superficial infectious keratitis. It can effectively eradicate lesions, hasten reepithelialization, and restore and preserve useful visual function. However, the selection of surgery candidates should be conducted carefully. PMID:24891945

  3. Viral keratitis-inhibitory effect of 9-beta-D-arabinofuranosylhypoxanthine 5'-monophosphate.

    PubMed

    Sidwell, R W; Allen, L B; Huffman, J H; Revankar, G R; Robins, R K; Tolman, R L

    1975-10-01

    Topical application of 9-beta-d-arabinofuranosylhypoxanthine 5'-monophosphate (ara-HxMP) significantly inhibited the development of keratitis induced by types 1 and 2 herpes simplex virus and vaccinia virus in the eyes of rabbits. Parameters for evaluation of efficacy were infectivity (corneal opacity, lesion size, and type), Draize (erythema, conjunctival swelling, and discharge), and reduction in titer of recoverable virus from the eye. When the relative efficacy of the related compounds 9-beta-d-arabinofuranosyladenine (ara-A), ara-A 5'-monophosphate (ara-AMP), and ara-Hx was determined against type 1 herpes simplex virus in a parallel experiment, the more water-soluble compounds (ara-HxMP, ara-AMP) were the most effective. The relative efficacy of ara-A was also determined against type 2 herpes and vaccinia virus-induced keratitis. Mortality in rabbits due to central nervous system involvement caused by types 1 and 2 herpes simplex virus was inhibited. Ara-HxMP was not discernibly toxic to the eye at concentrations of at least 20%; efficacy was still discernible with a 0.1% solution.

  4. Clinical efficacy of oral and topical acyclovir in herpes simplex virus stromal necrotizing keratitis

    PubMed Central

    Dutt, Surabhi; Acharya, Manisha; Gour, Abha; Sapra, Neelam; Chauhan, Lokesh; Mathur, Umang

    2016-01-01

    Purpose: To evaluate the efficacy of systemic and topical antiviral therapy in the treatment of active herpes simplex virus (HSV) necrotizing stromal keratitis (NSK). Design: Prospective interventional case series. Methodology: Patients with a diagnosis of HSV NSK based on history and clinical findings were enrolled in the study. A standard protocol was used for microbiologic investigations. Ten weeks regime of systemic acyclovir and 2 weeks of topical acyclovir was given. Complete ophthalmic examination was performed at every visit. Outcome measures were a reduction in the area of infiltration and improvement in visual acuity. Results: Fifteen patients were enrolled in the study. The mean age of presentation was 51.53 years. The duration of symptoms at presentation ranged from 2 to 8 weeks. HSV1 DNA polymerase chain reaction was positive in 70% cases of those tested. Area of infiltration at trial entry and at the end of 2 weeks of antiviral treatment reduced significantly (P = 0.007). All patients showed a complete resolution of keratitis at the end of study. Conclusion: Topical and systemic acyclovir for treatment of NSK facilitates healing of ulceration. Topical steroids after initial antiviral therapy are safe and decreases inflammation and improve visual recovery. Early initiation of therapy has better outcomes as compared to late presentations. PMID:27221681

  5. A Hydrogel-Based Hybrid Theranostic Contact Lens for Fungal Keratitis.

    PubMed

    Huang, Jian-Fei; Zhong, Jing; Chen, Guo-Pu; Lin, Zuan-Tao; Deng, Yuqing; Liu, Yong-Lin; Cao, Piao-Yang; Wang, Bowen; Wei, Yantao; Wu, Tianfu; Yuan, Jin; Jiang, Gang-Biao

    2016-07-26

    Fungal keratitis, a severe ocular disease, is one of the leading causes of ocular morbidity and blindness, yet it is often neglected, especially in developing countries. Therapeutic efficacy of traditional treatment such as eye drops is very limited due to poor bioavailability, whereas intraocular injection might cause serious side effects. Herein, we designed and fabricated a hybrid hydrogel-based contact lens which comprises quaternized chitosan (HTCC), silver nanoparticles, and graphene oxide (GO) with a combination of antibacterial and antifungal functions. The hydrogel is cross-linked through electrostatic interactions between GO and HTCC, resulting in strong mechanical properties. Voriconazole (Vor), an antifungal drug, can be loaded onto GO which retains the drug and promotes its sustained release from the hydrogel-based contact lenses. The contact lenses also exhibited good antimicrobial functions in view of glycidyltrimethylammonium chloride and silver nanoparticles. The results from in vitro and in vivo experiments demonstrate that contact lenses loaded with Vor have excellent efficacy in antifungal activity in vitro and could significantly enhance the therapeutic effects on a fungus-infected mouse model. The results indicate that this hydrogel contact lenses-based drug delivery system might be a promising therapeutic approach for a rapid and effective treatment of fungal keratitis. PMID:27244244

  6. Actinobacillus actinomycetemcomitans Keratitis After Glaucoma Infiltration Surgery: A Clinical Report and Literature Review.

    PubMed

    Hong, Jiaxu; Xu, Jianjiang; Cao, Wenjun; Ji, Jian; Sun, Xinghuai

    2016-01-01

    Actinobacillus actinomycetemcomitans infection is a rare and easily misdiagnosed ocular disease. In this article, the authors report a chronic, purulent, and difficult-to-treat case of A actinomycetemcomitans keratitis following a glaucoma infiltration surgery.A 56-year-old man with a long-standing history of open-angle glaucoma in both eyes presented with a 12-week history of ocular pain, redness, and blurred vision in his right eye. He underwent a glaucoma infiltration surgery in his right eye 6 months ago. Three months postoperatively, he developed peripheral corneal stromal opacities associated with a white, thin, cystic bleb, and conjunctival injection. These opacities grew despite topical treatment with topical tobramycin, levofloxacin, natamycin, amikacin, and metronidazole eye drops.Multiple corneal scrapings revealed no organisms, and no organisms grew on aerobic, anaerobic, fungal, or mycobacterial cultures. The patient's right eye developed a severe purulent corneal ulcer with a dense hypopyon and required a corneal transplantation. Histopathologic analysis and 16S ribosomalribonucleic acid polymerase chain reaction sequencing revealed A actinomycetemcomitans as the causative organism. Postoperatively, treatment was initiated with topical levofloxacin and cyclosporine, as well as oral levofloxacin and cyclosporine. Graft and host corneal transparency were maintained at the checkup 1 month after surgery.Although it is a rare cause of corneal disease, A actinomycetemcomitans should be suspected in patients with keratitis refractory to topical antibiotic therapy. Delay in diagnosis and appropriate treatment can result in vision loss. PMID:26817919

  7. A New Combination Formula for Treatment of Fungal Keratitis: An Experimental Study

    PubMed Central

    El-Mofty, Hala Mohamed; Abdelhakim, Mohamad Amr Salah Eddin; El-Miligi, Mohamed Farid; El-Nabarawi, Mohamed A.; Khalil, Islam Ahmed Hamed

    2014-01-01

    Objective. To formulate and evaluate slow release ketoconazole and ketorolac to treat fungal keratitis and associated inflammation. Methods. Experimental study with the following outcome measures. Pharmaceutical Evaluation. Mucoadhesive gels containing ketoconazole and ketorolac were used. Microbiological in vitro evaluation was performed using cup method. In vivo evaluation was performed on 24 rabbits divided into 2 groups, 12 rabbits each, group A (fast release formula; 6 times daily) and group B (slow release formula; 3 times daily). Each group was divided into two subgroups (6 rabbits each). Both eyes of rabbits were inoculated with Candida albicans. The left eye of all rabbits received the combination formulae. The right eye for one subgroup received ketoconazole as control 1 while the other subgroup received placebo as control 2. Clinical follow-up was done and, finally, the corneas were used for microbiological and pathological evaluation. Results. Gels containing high polymer concentration showed both high viscosity and mucoadhesion properties with slower drug release. The infected eyes treated with slow release formula containing both drugs showed better curing of the cornea and pathologically less inflammation than eyes treated with fast release formula. Conclusion. Slow release formula containing ketoconazole and ketorolac showed higher activity than fast release formula against fungal keratitis and associated inflammation. PMID:24872888

  8. Actinobacillus actinomycetemcomitans Keratitis After Glaucoma Infiltration Surgery: A Clinical Report and Literature Review.

    PubMed

    Hong, Jiaxu; Xu, Jianjiang; Cao, Wenjun; Ji, Jian; Sun, Xinghuai

    2016-01-01

    Actinobacillus actinomycetemcomitans infection is a rare and easily misdiagnosed ocular disease. In this article, the authors report a chronic, purulent, and difficult-to-treat case of A actinomycetemcomitans keratitis following a glaucoma infiltration surgery.A 56-year-old man with a long-standing history of open-angle glaucoma in both eyes presented with a 12-week history of ocular pain, redness, and blurred vision in his right eye. He underwent a glaucoma infiltration surgery in his right eye 6 months ago. Three months postoperatively, he developed peripheral corneal stromal opacities associated with a white, thin, cystic bleb, and conjunctival injection. These opacities grew despite topical treatment with topical tobramycin, levofloxacin, natamycin, amikacin, and metronidazole eye drops.Multiple corneal scrapings revealed no organisms, and no organisms grew on aerobic, anaerobic, fungal, or mycobacterial cultures. The patient's right eye developed a severe purulent corneal ulcer with a dense hypopyon and required a corneal transplantation. Histopathologic analysis and 16S ribosomalribonucleic acid polymerase chain reaction sequencing revealed A actinomycetemcomitans as the causative organism. Postoperatively, treatment was initiated with topical levofloxacin and cyclosporine, as well as oral levofloxacin and cyclosporine. Graft and host corneal transparency were maintained at the checkup 1 month after surgery.Although it is a rare cause of corneal disease, A actinomycetemcomitans should be suspected in patients with keratitis refractory to topical antibiotic therapy. Delay in diagnosis and appropriate treatment can result in vision loss.

  9. Treatment with meliacine, a plant derived antiviral, prevents the development of herpetic stromal keratitis in mice.

    PubMed

    Alché, L E; Berra, A; Veloso, M J; Coto, C E

    2000-08-01

    Herpetic stromal keratitis is caused by ocular infection with herpes simplex virus type 1 (HSV-1) and constitutes a leading cause of human blindness. The effect of meliacine, an antiviral compound isolated from leaves of Melia azedarach L. that inhibits HSV-1 replication in vitro, was examined on experimental corneal HSV-1 inoculation in Balb/c mice. Mice were inoculated with HSV-1 strain KOS at their corneas after abrasion. Meliacine was administered topically 3 times a day for 4 days beginning 1 day before inoculation. Infected animals treated or not with meliacine were observed carefully for the development of stromal keratitis and the clinical scoring was done 14 days post-infection. Histological examination of corneas and viral isolation from eyes from HSV-1 infected mice treated or not with meliacine were also carried out. It was found that the treatment of HSV-1-induced ocular disease in Balb/c mice with meliacine reduced significantly the development of clinical disease, as well as the histological damage in corneas. The viral titers detected in eyes of infected and treated mice were 2-orders-of-magnitude lower than those corresponding to HSV-1 infected control animals. Mock-infected and treated mice did not reveal any corneal alteration due to the administration of the compound. Meliacine was found to exert a strong antiviral action on HSV-1-induced ocular disease in mice with no evidence of toxic effects. PMID:10897066

  10. Assessment of Rose Bengal vs. Riboflavin Photodynamic Therapy for Inhibition of Fungal Keratitis Isolates

    PubMed Central

    Arboleda, Alejandro; Miller, Darlene; Cabot, Florence; Taneja, Mukesh; Aguilar, Mariela C.; Alawa, Karam; Amescua, Guillermo; Yoo, Sonia H.; Parel, Jean-Marie

    2014-01-01

    Purpose To compare the in vitro effect of rose bengal and riboflavin as photosensitizing agents for photodynamic therapy (PDT) on fungal isolates that are common causes of fungal keratitis Design Experimental study Methods Three isolates (Fusarium solani, Aspergillus fumigatus, Candida albicans) recovered from patients with confirmed fungal keratitis were used in the experiments. Isolates were grown on Sabouraud-Dextrose agar, swabbed and prepared in suspension, and one milliliter aliquots were inoculated onto test plates in triplicate. Test plates were separated into 5 groups: Group 1 - no treatment, Group 2 - 0.1% rose bengal alone, Group 3 - 518 nm irradiation alone, Group 4 - riboflavin PDT (riboflavin + 375 nm irradiation), and Group 5 - rose bengal PDT (rose bengal + 518 nm irradiation). Irradiation was performed over a circular area using either a green LED array (peak wavelength: 518 nm) or a UV-A LED array (peak wavelength: 375 nm). Test plates were irradiated with an energy density of 5.4 J/cm2. Later, plates were placed in a 30° C incubator and observed for growth. Results Rose bengal-mediated PDT successfully inhibited the growth of all three fungal isolates in the irradiated area. All other groups exhibited unrestricted growth throughout the plate. Conclusions Rose bengal-mediated PDT successfully inhibited the growth of three types of fungi. No other experimental groups, including riboflavin-mediated PDT, had any inhibitory effect on the isolates. The results might be useful for the treatment of patients suffering from corneal infection. PMID:24792103

  11. Clinical features of infectious keratitis at a tertiary referral center in a rural area of Korea

    PubMed Central

    Lee, Seung-Jun; Lee, Jang Hun; Kim, Moosang; Han, Sang Beom; Hyon, Joon Young

    2015-01-01

    Purpose To evaluate clinical features of infectious keratitis at a tertiary referral center in a rural area of Korea. Results This was a retrospective study. Medical records of 70 patients who were diagnosed with infectious keratitis and underwent corneal culture were reviewed. Data including demographics, predisposing factors, and microbiological profiles were collected and analyzed. Results Age distribution showed one peak at sixth and seventh decade, and 48 patients (68.6%) were ≥50 years old. The male to female ratio was 43 (61.4%):27 (38.6%). Ocular surface disease was the most frequently found (48 cases, 68.6%). Gram-positive bacteria were most common (ten patients, 66.7%), followed by Gram-negative bacteria (three patients, 20%) and fungi (two patients, 13.3%). In this study, 34.3% of patients were treated by at least one topical antimicrobial before culture. Patients who received topical therapy before culture showed positive culture rate of 16.7%, and patients who underwent corneal culture without topical antibiotics showed positive culture rate of 23.9%. However, there was no statistically significant difference between the two groups in positive culture rate (P=0.554). Conclusion The proportions of the elderly and male patients were higher than the young and female patients, respectively. Gram-positive bacteria were the most common pathogens, and ocular surface disease was the most common predisposing factor. PMID:26675320

  12. A new combination formula for treatment of fungal keratitis: an experimental study.

    PubMed

    El-Mofty, Hala Mohamed; Abdelhakim, Mohamad Amr Salah Eddin; El-Miligi, Mohamed Farid; El-Nabarawi, Mohamed A; Khalil, Islam Ahmed Hamed

    2014-01-01

    Objective. To formulate and evaluate slow release ketoconazole and ketorolac to treat fungal keratitis and associated inflammation. Methods. Experimental study with the following outcome measures. Pharmaceutical Evaluation. Mucoadhesive gels containing ketoconazole and ketorolac were used. Microbiological in vitro evaluation was performed using cup method. In vivo evaluation was performed on 24 rabbits divided into 2 groups, 12 rabbits each, group A (fast release formula; 6 times daily) and group B (slow release formula; 3 times daily). Each group was divided into two subgroups (6 rabbits each). Both eyes of rabbits were inoculated with Candida albicans. The left eye of all rabbits received the combination formulae. The right eye for one subgroup received ketoconazole as control 1 while the other subgroup received placebo as control 2. Clinical follow-up was done and, finally, the corneas were used for microbiological and pathological evaluation. Results. Gels containing high polymer concentration showed both high viscosity and mucoadhesion properties with slower drug release. The infected eyes treated with slow release formula containing both drugs showed better curing of the cornea and pathologically less inflammation than eyes treated with fast release formula. Conclusion. Slow release formula containing ketoconazole and ketorolac showed higher activity than fast release formula against fungal keratitis and associated inflammation. PMID:24872888

  13. Keratitis after Implantation of Intrastromal Corneal Rings with Spontaneous Extrusion of the Segment.

    PubMed

    Ibáñez-Alperte, Juan; Pérez-García, Diana; Cristóbal, José A; Mateo, Antonio J; Río, Beatriz Jiménez-Del; Mínguez, Enrique

    2010-09-13

    PURPOSE: To report a case of bacterial keratitis in a patient with a history of intrastromal corneal ring segments (INTACS®) implantation to correct keratoconus. METHODS: The patient's history, clinical presentation, pathological analysis and therapeutic management were reviewed. RESULTS: A 36-year-old-man was referred to our department due to decreased vision and intense pain in his left eye, 40 days after INTACS® implantation for keratoconus. Slit-lamp examination revealed epithelial defects and stromal infiltrates in the lower channel without evidence of the inferior ring. The anterior chamber also showed a significant fibrin reaction to hypopyon. A low-tension suture was removed at the site of the incision. Microbiological study of the conjunctival swab was positive for Staphylococcus epidermidis, but the corneal culture was sterile. The patient was treated with topical fortified and systemic antibiotics. The infection slowly resolved, leaving opacity at the inferior segment site. CONCLUSIONS: Infectious keratitis following INTACS implantation is an infrequent complication that can have important consequences without suitable and early therapeutic management.

  14. In-vivo immunofluorescence confocal microscopy of herpes simplex virus type 1 keratitis

    NASA Astrophysics Data System (ADS)

    Kaufman, Stephen C.; Laird, Jeffery A.; Beuerman, Roger W.

    1996-05-01

    The white-light confocal microscope offers an in vivo, cellular-level resolution view of the cornea. This instrument has proven to be a valuable research and diagnostic tool for the study of infectious keratitis. In this study, we investigate the direct visualization of herpes simplex virus type 1 (HSV-1)-infected corneal epithelium, with in vivo confocal microscopy, using HSV-1 immunofluorescent antibodies. New Zealand white rabbits were infected with McKrae strain of HSV-1 in one eye; the other eye of each rabbit was used as an uninfected control. Four days later, the rabbits were anesthetized and a cellulose sponge was applied to each cornea, and a drop of direct HSV fluorescein-tagged antibody was placed on each sponge every 3 to 5 minutes for 1 hour. Fluorescence confocal microscopy was then performed. The HSV-infected corneas showed broad regions of hyperfluorescent epithelial cells. The uninfected corneas revealed no background fluorescence. Thus, using the confocal microscope with a fluorescent cube, we were able to visualize HSV-infected corneal epithelial cells tagged with a direct fluorescent antibody. This process may prove to be a useful clinical tool for the in vivo diagnosis of HSV keratitis.

  15. Evaluation of nested pcr technique for detection of Pythium insidiosum in pathological specimens from patients with suspected fungal keratitis.

    PubMed

    Kosrirukvongs, Panida; Chaiprasert, Angkana; Uiprasertkul, Mongkol; Chongcharoen, Metinee; Banyong, Ramrada; Krajaejun, Theerapong; Wanachiwanawin, Wanchai

    2014-01-01

    Diagnosis of Pythium keratitis is problematic due to the difficulty in obtaining a culture report resulting in unnecessarily prolonged usage of antimicrobial medication due to misdiagnosis. This study evaluated and compared nested PCR technique with culture and immunoperoxidase staining assays of Pythium insidiosum in paraffin-embedded corneal tissues from patients with suspected fungal keratitis. Six of 51 pathological reports compatible with fungal infection and 6 of 48 culture-proven fungal keratitis were identified as Pythium. Twenty-seven specimens were PCR-positive for Pythium insidiosum. In comparison with fungal culture for P. insidiosum, PCR had 83% sensitivity and 77% specificity with fair agreement (Kappa score of 0.227, p = 0.001). The mean age of PCR-positive is younger than PCR-negative group and there is a female preponderance in Pythium-infected group (p = 0.002 and p = 0.004, respectively). Nineteen specimens had positive results using immunoperoxidase staining assay with fair agreement to culture method (Kappa 0.340, p < 0.001), and 83% sensitivity, 85% specificity and 85% accuracy (95% CI: 76.7-90.7). PCR-based technique compared with culture and/or immunoperoxidase staining assay had 91.7% sensitivity, 81.8% specificity and 83% accuracy (95% CI: 74.5-89.1) with moderate agreement (Kappa 0.477, p < 0.001). Thus nested PCR detection of P. insidiosum should be employed in preliminary diagnosis of Pythium keratitis in order to initiate proper management. PMID:24964666

  16. Therapeutic potential of the antimicrobial peptide OH-CATH30 for antibiotic-resistant Pseudomonas aeruginosa keratitis.

    PubMed

    Li, Sheng-An; Liu, Jie; Xiang, Yang; Wang, Yan-Jie; Lee, Wen-Hui; Zhang, Yun

    2014-06-01

    The therapeutic potential of antimicrobial peptides (AMPs) has been evaluated in many infectious diseases. However, the topical application of AMPs for ocular bacterial infection has not been well investigated. The AMP OH-CATH30, which was identified in the king cobra, exhibits potent antimicrobial activity. In this study, we investigated the therapeutic potential of OH-CATH30 for Pseudomonas aeruginosa keratitis. Ten isolates of P. aeruginosa from individuals with keratitis were susceptible to OH-CATH30 but not to cefoperazone, ciprofloxacin, gentamicin, and levofloxacin. The microdilution checkerboard assay showed that OH-CATH30 exhibited synergistic activity with ciprofloxacin and levofloxacin against antibiotic-resistant P. aeruginosa. Meanwhile, P. aeruginosa did not develop resistance to OH-CATH30, even after exposure at 0.5× the MIC for up to 25 subcultures. Furthermore, treatment with OH-CATH30, alone or in combination with levofloxacin, significantly improved the clinical outcomes of rabbit keratitis induced by antibiotic-resistant P. aeruginosa. Taken together, our data indicate that the topical application of OH-CATH30 is efficacious against drug-resistant P. aeruginosa keratitis. In addition, our study highlights the potential application of AMPs in treating ocular bacterial infections. PMID:24637683

  17. Conjunctival Flap Covering Combined with Antiviral and Steroid Therapy for Severe Herpes Simplex Virus Necrotizing Stromal Keratitis

    PubMed Central

    Jia, Yanni; Li, Suxia; Wang, Ting; Tan, Yaohong; Shi, Weiyun

    2015-01-01

    Herpes simplex virus (HSV) necrotizing stromal keratitis is a common type of herpetic stromal keratitis (HSK). Antiviral medication alone cannot control the disease, and corticosteroid eye drops may aggravate the ulcer and result in corneal perforation. Amniotic membrane transplantation effectively treats superficial corneal ulcer resulting from necrotizing stromal HSK. However, the efficacy of this approach seems to be limited for more serious cases. This study presented the clinical treatment of severe HSV necrotizing stromal keratitis (ulcer depth greater than half of the corneal stroma) by conjunctival flap covering surgery in 25 patients (25 eyes) combined with antivirus and corticosteroid treatment at Shandong Eye Hospital from January 2007 to December 2013. Clinical results showed that the mean best spectacle-corrected visual acuity improved from preoperative 20/333 to postoperative 20/40 (P < 0.05). All patients recovered ocular surface stabilization. There was recurrence in two eyes, which was cured with antiviral medication. Conjunctival flap covering combined with antivirus and corticosteroid treatment is effective in treating severe HSV necrotizing stromal keratitis. PMID:25785282

  18. [Incurable keratitis and chronic palmoplantar hyperkeratosis with hypertyrosinemia. Cure using a tyrosine-restricted diet. Type II tyrosinemia].

    PubMed

    Hervé, F; Moreno, J L; Ogier, H; Saudubray, J M; De Prost, Y; Duffier, J L; Charpentier, C; Lemonnier, F; Frézal, J

    1986-01-01

    One should henceforth systematically search for hypertyrosinemia which, too often, goes unrecognized for years, in patients presenting chronic keratitis associated with palmar and plantar hyperkeratosis. As a matter of fact, this highly crippling disease may be cured with an appropriate diet and the diagnosis, once suspected, is easily confirmed by simple investigations.

  19. Comparison of reflectance confocal microscopy and two-photon second harmonic generation microscopy in fungal keratitis rabbit model ex vivo.

    PubMed

    Lee, Jun Ho; Lee, Seunghun; Yoon, Calvin J; Park, Jin Hyoung; Tchah, Hungwon; Kim, Myoung Joon; Kim, Ki Hean

    2016-02-01

    Fungal keratitis is an infection of the cornea by fungal pathogens. Diagnosis methods based on optical microscopy could be beneficial over the conventional microbiology method by allowing rapid and non-invasive examination. Reflectance confocal microscopy (RCM) and two-photon second harmonic generation microscopy (TPSHGM) have been applied to pre-clinical or clinical studies of fungal keratitis. In this report, RCM and TPSHGM were characterized and compared in the imaging of a fungal keratitis rabbit model ex vivo. Fungal infection was induced by using two strains of fungi: aspergillus fumigatus and candida albicans. The infected corneas were imaged in fresh condition by both modalities sequentially and their images were analyzed. Both RCM and TPSHGM could detect both fungal strains within the cornea based on morphology: aspergillus fumigatus had distinctive filamentous structures, and candida albicans had round structures superficially and elongated structures in the corneal stroma. These imaging results were confirmed by histology. Comparison between RCM and TPSHGM showed several characteristics. Although RCM and TPSHGM images had good correlation each other, their images were slightly different due to difference in contrast mechanism. RCM had relatively low image contrast with the infected turbid corneas due to high background signal. TPSHGM visualized cells and collagen in the cornea clearly compared to RCM, but used higher laser power to compensate low autofluorescence. Since these two modalities provide complementary information, combination of RCM and TPSHGM would be useful for fungal keratitis detection by compensating their weaknesses each other. PMID:26977371

  20. Comparison of reflectance confocal microscopy and two-photon second harmonic generation microscopy in fungal keratitis rabbit model ex vivo

    PubMed Central

    Lee, Jun Ho; Lee, Seunghun; Yoon, Calvin J.; Park, Jin Hyoung; Tchah, Hungwon; Kim, Myoung Joon; Kim, Ki Hean

    2016-01-01

    Fungal keratitis is an infection of the cornea by fungal pathogens. Diagnosis methods based on optical microscopy could be beneficial over the conventional microbiology method by allowing rapid and non-invasive examination. Reflectance confocal microscopy (RCM) and two-photon second harmonic generation microscopy (TPSHGM) have been applied to pre-clinical or clinical studies of fungal keratitis. In this report, RCM and TPSHGM were characterized and compared in the imaging of a fungal keratitis rabbit model ex vivo. Fungal infection was induced by using two strains of fungi: aspergillus fumigatus and candida albicans. The infected corneas were imaged in fresh condition by both modalities sequentially and their images were analyzed. Both RCM and TPSHGM could detect both fungal strains within the cornea based on morphology: aspergillus fumigatus had distinctive filamentous structures, and candida albicans had round structures superficially and elongated structures in the corneal stroma. These imaging results were confirmed by histology. Comparison between RCM and TPSHGM showed several characteristics. Although RCM and TPSHGM images had good correlation each other, their images were slightly different due to difference in contrast mechanism. RCM had relatively low image contrast with the infected turbid corneas due to high background signal. TPSHGM visualized cells and collagen in the cornea clearly compared to RCM, but used higher laser power to compensate low autofluorescence. Since these two modalities provide complementary information, combination of RCM and TPSHGM would be useful for fungal keratitis detection by compensating their weaknesses each other. PMID:26977371

  1. Growth phase dependent alterations in the surface coat of Acanthamoeba castellanii.

    PubMed

    Przełecka, A; Sobota, A

    1982-01-01

    Application of ruthenium red, cationized ferritin and concanavalin A to exponentially growing trophozoites reveals on their plasma membrane negatively charged surface coat bearing sugar residues. In the coat of trophozoites from advanced stationary growth phase no sugar residues can be visualized. In mature cysts the external layer of their wall is negatively charged, however, on their protoplast surface no terminals reacting with the 2 polycations, or with concanavalin A can be revealed, even though the penetration of the reagents has been ensured by enzymatic impairing of the cyst wall. The results are confronted with the known facts concerning alterations of physiological properties of plasma membrane occurring during the life cycle of Acanthamoeba.

  2. Detection of free living amoebae, Acanthamoeba and Naegleria, in swimming pools, Malaysia.

    PubMed

    Init, I; Lau, Y L; Arin Fadzlun, A; Foead, A I; Neilson, R S; Nissapatorn, V

    2010-12-01

    This study reports the detection of Acanthamoeba and Naegleria species in 14 swimming pools around Petaling Jaya and Kuala Lumpur, Malaysia. Sampling was carried out at 4 sites (the platforms (P), wall (W), 1 meter from the wall (1) and middle (2)) of each swimming pool. These free living amoebae (FLA) were detected under light and inverted microscopes after being cultured on the surface of non-nutrient agar lawned with Escherichia coli. Acanthamoeba species were detected in higher number of culture plates from all sampling sites of all the swimming pools. While Naegleria, were detected in fewer culture plates at 3 sampling sites (absent at site P) of 8 swimming pools. This suggested that the thick double-walled cysts of Acanthamoeba were more resistant, thus remaining viable in the dry-hot areas of the platforms and in chlorinated water of the swimming pools whereas Naegleria cysts, that are fragile and susceptible to desiccation, preferred watery or moist areas for growth and proliferation. The prevalence of both FLA was highest at site W (76.2%), followed by site 1 (64.7%), lowest at site 2 (19.4%), and could be detected at all 3 sampling levels (top, middle and bottom) of these 3 sites. The surface of site W might act as a bio-film that accumulated all kinds of microbes providing sufficient requirement for the FLA to develop and undergo many rounds of life cycles as well as moving from top to bottom in order to graze food. Other factors such as human activities, the circulating system which was fixed at all swimming pools, blowing wind which might carry the cysts from surroundings and the swimming flagellate stage of Naegleria could also contribute to the distribution of the FLA at these sampling sites. Both FLA showed highest growth (80.4%) at room temperature (25-28 ºC) and lesser (70.0%) at 37 ºC which might be due to the overgrowth of other microbes (E. coli, fungi, algae, etc). While at 44 ºC, only Acanthamoeba species could survive thus showing that

  3. Acanthamoeba polyphaga mimivirus and other giant viruses: an open field to outstanding discoveries.

    PubMed

    Abrahão, Jônatas S; Dornas, Fábio P; Silva, Lorena C F; Almeida, Gabriel M; Boratto, Paulo V M; Colson, Phillipe; La Scola, Bernard; Kroon, Erna G

    2014-06-30

    In 2003, Acanthamoeba polyphaga mimivirus (APMV) was first described and began to impact researchers around the world, due to its structural and genetic complexity. This virus founded the family Mimiviridae. In recent years, several new giant viruses have been isolated from different environments and specimens. Giant virus research is in its initial phase and information that may arise in the coming years may change current conceptions of life, diversity and evolution. Thus, this review aims to condense the studies conducted so far about the features and peculiarities of APMV, from its discovery to its clinical relevance.

  4. Acanthamoeba polyphaga mimivirus and other giant viruses: an open field to outstanding discoveries

    PubMed Central

    2014-01-01

    In 2003, Acanthamoeba polyphaga mimivirus (APMV) was first described and began to impact researchers around the world, due to its structural and genetic complexity. This virus founded the family Mimiviridae. In recent years, several new giant viruses have been isolated from different environments and specimens. Giant virus research is in its initial phase and information that may arise in the coming years may change current conceptions of life, diversity and evolution. Thus, this review aims to condense the studies conducted so far about the features and peculiarities of APMV, from its discovery to its clinical relevance. PMID:24976356

  5. 77 FR 16314 - Alaska Disaster #AK-00024

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-03-20

    ... ADMINISTRATION Alaska Disaster AK-00024 AGENCY: U.S. Small Business Administration. ACTION: Notice. SUMMARY: This is a notice of an Administrative declaration of a disaster for the State of Alaska dated 03/13/2012... INFORMATION CONTACT: A. Escobar, Office of Disaster Assistance, U.S. Small Business Administration, 409...

  6. 78 FR 39822 - Alaska Disaster #AK-00028

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-02

    ... ADMINISTRATION Alaska Disaster AK-00028 AGENCY: U.S. Small Business Administration. ACTION: Notice SUMMARY: This is a Notice of the Presidential declaration of a major disaster for the State of Alaska (FEMA-4122-DR... INFORMATION CONTACT: A. Escobar, Office of Disaster Assistance, U.S. Small Business Administration, 409...

  7. 78 FR 39821 - Alaska Disaster #AK-00029

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-02

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  9. Interleukin 4 and T helper type 2 cells are required for development of experimental onchocercal keratitis (river blindness)

    PubMed Central

    1995-01-01

    Inflammation of the corneal stroma (stromal keratitis) is a serious complication of infection with the nematode parasite Onchocerca volvulus. Because stromal keratitis is believed to be immunologically mediated in humans, we used a murine model to examine the role of T cells and T helper cell cytokines in the immunopathogenesis of these eye lesions. BALB/c mice immunized subcutaneously and injected intrastromally with soluble O. volvulus antigens (OvAg) developed pronounced corneal opacification and neovascularization. The corneal stroma was edematous and contained numerous eosinophils and mononuclear cells. Stromal keratitis in immunized mice was determined to be T cell dependent based on the following observations: (a) T cell-deficient nude mice immunized and injected intrastromally with OvAg fail to develop corneal pathology; and (b) adoptive transfer of spleen cells from OvAg-immunized BALB/c mice to naive nude mice before intrastromal injection of OvAg results in development of keratitis. OvAg-stimulated lymph node and spleen cell cytokine production was dependent on CD4 cells and included interleukin (IL)-4 and IL-5, but not interferon gamma, indicating a predominant T helper type 2 cell-like response. Inflamed corneas from immunized BALB/c mice and from reconstituted nude mice had greatly elevated CD4 and IL-4 gene expression compared with interferon gamma. Mice in which the IL-4 gene was disrupted failed to develop corneal disease, demonstrating that IL-4 is essential in the immunopathogenesis of O. volvulus-mediated stromal keratitis. PMID:7561696

  10. Phylogenetic analysis and the evolution of the 18S rRNA gene typing system of Acanthamoeba.

    PubMed

    Fuerst, Paul A; Booton, Gregory C; Crary, Monica

    2015-01-01

    Species of Acanthamoeba were first described using morphological characters including cyst structure and cytology of nuclear division. More than 20 nominal species were proposed using these methods. Morphology, especially cyst shape and size, has proven to be plastic and dependent upon culture conditions. The DNA sequence of the nuclear small-subunit (18S) rRNA, the Rns gene, has become the most widely accepted method for rapid diagnosis and classification of Acanthamoeba. The Byers-Fuerst lab first proposed an Rns typing system in 1996. Subsequent refinements, with an increasing DNA database and analysis of diagnostic fragments within the gene, have become widely accepted by the Acanthamoeba research community. The development of the typing system, including its current state of implementation is illustrated by three cases: (i) the division between sequence types T13 and T16; (ii) the diversity within sequence supertype T2/T6, and (iii) verification of a new sequence type, designated T20. Molecular studies make clear the disconnection between phylogenetic relatedness and species names, as applied for the genus Acanthamoeba. Future reconciliation of genetic types with species names must become a priority, but the possible shortcomings of the use of a single gene when reconstructing the evolutionary history of the acanthamoebidae must also be resolved. PMID:25284310

  11. Acanthamoeba belonging to T3, T4, and T11: genotypes isolated from air-conditioning units in Santiago, Chile.

    PubMed

    Astorga, Berbeli; Lorenzo-Morales, Jacob; Martín-Navarro, Carmen M; Alarcón, Verónica; Moreno, Johanna; González, Ana C; Navarrete, Elizabeth; Piñero, José E; Valladares, Basilio

    2011-01-01

    Free-living amoebae (FLA) of the genus Acanthamoeba are widely distributed in the environment, in the air, soil, and water, and have also been isolated from air-conditioning units. The objective of this work was to investigate the presence of this genus of FLA in the air-conditioning equipment at the Institute of Public Health of Chile in Santiago, Chile. Water and air samples were collected from air-conditioning systems and were checked for the presence of Acanthamoeba spp. Positive samples were further classified at the genotype level after sequencing the highly variable diagnostic fragment 3 (DF3) region of the 18S rRNA gene. This is the first report of the T3, T4, and T11 genotypes of Acanthamoeba in air-conditioning units from Chile. Overall, the widespread distribution of potentially pathogenic Acanthamoeba strains in the studied source demands more awareness within the public and health professionals in Chile as this pathogen is emerging as a risk for human health worldwide.

  12. Phylogenetic analysis and the evolution of the 18S rRNA gene typing system of Acanthamoeba.

    PubMed

    Fuerst, Paul A; Booton, Gregory C; Crary, Monica

    2015-01-01

    Species of Acanthamoeba were first described using morphological characters including cyst structure and cytology of nuclear division. More than 20 nominal species were proposed using these methods. Morphology, especially cyst shape and size, has proven to be plastic and dependent upon culture conditions. The DNA sequence of the nuclear small-subunit (18S) rRNA, the Rns gene, has become the most widely accepted method for rapid diagnosis and classification of Acanthamoeba. The Byers-Fuerst lab first proposed an Rns typing system in 1996. Subsequent refinements, with an increasing DNA database and analysis of diagnostic fragments within the gene, have become widely accepted by the Acanthamoeba research community. The development of the typing system, including its current state of implementation is illustrated by three cases: (i) the division between sequence types T13 and T16; (ii) the diversity within sequence supertype T2/T6, and (iii) verification of a new sequence type, designated T20. Molecular studies make clear the disconnection between phylogenetic relatedness and species names, as applied for the genus Acanthamoeba. Future reconciliation of genetic types with species names must become a priority, but the possible shortcomings of the use of a single gene when reconstructing the evolutionary history of the acanthamoebidae must also be resolved.

  13. Severe Amoebic Placentitis in a Horse Caused by an Acanthamoeba hatchetti Isolate Identified Using Next-Generation Sequencing

    PubMed Central

    Begg, Angela P.; Todhunter, Kristen; Donahoe, Shannon L.; Krockenberger, Mark

    2014-01-01

    A case of amoebic placentitis in a mare from eastern Australia was diagnosed postpartum by histopathological examination of the placenta. The identity of the etiological agent was confirmed as Acanthamoeba hatchetti by use of diversity profiling based on a next-generation sequencing approach. PMID:24829227

  14. Identification of Protein Arginine Methyltransferase 5 as a Regulator for Encystation of Acanthamoeba

    PubMed Central

    Moon, Eun-Kyung; Hong, Yeonchul; Chung, Dong-Il; Goo, Youn-Kyoung; Kong, Hyun-Hee

    2016-01-01

    Encystation is an essential process for Acanthamoeba survival under nutrient-limiting conditions and exposure to drugs. The expression of several genes has been observed to increase or decrease during encystation. Epigenetic processes involved in regulation of gene expression have been shown to play a role in several pathogenic parasites. In the present study, we identified the protein arginine methyltransferase 5 (PRMT5), a known epigenetic regulator, in Acanthamoeba castellanii. PRMT5 of A. castellanii (AcPRMT5) contained domains found in S-adenosylmethionine-dependent methyltransferases and in PRMT5 arginine-N-methyltransferase. Expression levels of AcPRMT5 were increased during encystation of A. castellanii. The EGFP-PRMT5 fusion protein was mainly localized in the nucleus of trophozoites. A. castellanii transfected with siRNA designed against AcPRMT5 failed to form mature cysts. The findings of this study lead to a better understanding of epigenetic mechanisms behind the regulation of encystation in cyst-forming pathogenic protozoa. PMID:27180570

  15. Investigating the role of Acanthamoeba polyphaga in protecting Human Adenovirus from water disinfection treatment.

    PubMed

    Verani, Marco; Di Giuseppe, Graziano; Tammaro, Carmine; Carducci, Annalaura

    2016-06-01

    Human adenoviruses are responsible for a wide range of clinical infections and are present in aquatic environments, including river, seawater, drinking-water and sewage. Free-living amoebae (Acanthamoeba) in the same environments may internalize them and other microorganisms can act as a reservoir for the internalized viruses. In this study, we studied the interaction between Acanthamoeba polyphaga and Human Adenovirus type 5 (HAdV 5) to determine whether the amoeba played a role in protecting the internalized viruses from chemical disinfection. The efficacy of sodium hypochlorite disinfection against A. polyphaga and HAdV 5 either singly or in combination was assessed at three different concentrations. Individually, the amoeba were more resistant to chemical disinfection than HAdV 5 and remained alive after exposure to 5mg/l of sodium hypochlorite. In contrast, HAdV 5 lost infectivity following exposure to 2.5mg/l of sodium hypochlorite. When the amoeba and HAdV 5 were co-cultured, infectious virus was found in the cytoplasm of the amoeba at 5mg/l disinfectant concentration. These findings suggest that the A. polyphaga is providing protection for the HAdV 5. PMID:26999560

  16. Purification from Acanthamoeba castellanii of proteins that induce gelation and syneresis of F-actin.

    PubMed

    Maruta, H; Korn, E D

    1977-01-10

    From Acanthamoeba castellanii, we have purified four proteins each of which alone causes a solution of F-actin to gel. The four active proteins have subunit molecular weights of about 23,000, 28,000, 32,000 and 38,000, respectively; the last three may be dimers in their native proteins. Together, these four proteins account for about 97% of the gelation activity of the whole extract; not more than about 3% of the total activity of the unfractionated extract can be due to a 250,000-dalton polypeptide. Another protein fraction, purified by agarose chromatography, induces shrinking (syneresis) of gels formed from F-actin and any of the gelation factors. That fraction contains a high Ca2+-, low (K+,EDTA)-ATPase and a major polypeptide of 170,000 daltons both of which bind to actin in the shrunken gel pellet. The active fraction does not contain the previously described Acanthamoeba myosin (Pollard, T. D., and Korn, E. D. (1973) J. Biol. Chem. 248, 4682-4690).

  17. Campylobacter jejuni actively invades the amoeba Acanthamoeba polyphaga and survives within non digestive vacuoles.

    PubMed

    Olofsson, Jenny; Axelsson-Olsson, Diana; Brudin, Lars; Olsen, Björn; Ellström, Patrik

    2013-01-01

    The Gram-negative bacterium Campylobacter jejuni is able to enter, survive and multiply within the free living amoeba Acanthamoeba polyphaga, but the molecular mechanisms behind these events are still unclear. We have studied the uptake and intracellular trafficking of viable and heat killed bacterial cells of the C. jejuni strain 81-176 in A. polyphaga. We found that viable bacteria associated with a substantially higher proportion of Acanthamoeba trophozoites than heat killed bacteria. Furthermore, the kinetics of internalization, the total number of internalized bacteria as well as the intracellular localization of internalized C. jejuni were dramatically influenced by bacterial viability. Viable bacteria were internalized at a high rate already after 1 h of co-incubation and were observed in small vacuoles tightly surrounding the bacteria. In contrast, internalization of heat killed C. jejuni was low at early time points and did not peak until 96 h. These cells were gathered in large spacious vacuoles that were part of the degradative pathway as determined by the uptake of fluorescently labeled dextran. The amount of heat killed bacteria internalized by A. polyphaga did never reach the maximal amount of internalized viable bacteria. These results suggest that the uptake and intracellular survival of C. jejuni in A. polyphaga is bacterially induced. PMID:24223169

  18. Behavior of Yersinia enterocolitica in the presence of the bacterivorous Acanthamoeba castellanii.

    PubMed

    Lambrecht, E; Baré, J; Van Damme, I; Bert, W; Sabbe, K; Houf, K

    2013-10-01

    Free-living protozoa play an important role in the ecology and epidemiology of human-pathogenic bacteria. In the present study, the interaction between Yersinia enterocolitica, an important food-borne pathogen, and the free-living amoeba Acanthamoeba castellanii was studied. Several cocultivation assays were set up to assess the resistance of Y. enterocolitica to A. castellanii predation and the impact of environmental factors and bacterial strain-specific characteristics. Results showed that all Y. enterocolitica strains persist in association with A. castellanii for at least 14 days, and associations with A. castellanii enhanced survival of Yersinia under nutrient-rich conditions at 25°C and under nutrient-poor conditions at 37°C. Amoebae cultivated in the supernatant of one Yersinia strain showed temperature- and time-dependent permeabilization. Intraprotozoan survival of Y. enterocolitica depended on nutrient availability and temperature, with up to 2.8 log CFU/ml bacteria displaying intracellular survival at 7°C for at least 4 days in nutrient-rich medium. Transmission electron microscopy was performed to locate the Yersinia cells inside the amoebae. As Yersinia and Acanthamoeba share similar ecological niches, this interaction identifies a role of free-living protozoa in the ecology and epidemiology of Y. enterocolitica. PMID:23934496

  19. Acanthamoeba castellanii metabolites increase the intracellular calcium level and cause cytotoxicity in wish cells.

    PubMed

    Mattana, A; Bennardini, F; Usai, S; Fiori, P L; Franconi, F; Cappuccinelli, P

    1997-08-01

    Previous studies have shown that trophozoites of the pathogenic free-living amoeba Acanthamoeba castellanii rapidly lyse a variety of cells in vitro. However, the role played by cytolitic molecules that may participate in Acanthamoebal cytopathogenicity has yet to be completely elucidated. The aim of this work was to study whether soluble molecules released by A. castellanii trophozoites could induce cytopathic effect in human epithelial cells in vitro. The results obtained indicate that A. castellanii trophozoites constitutively elaborate and release soluble factors that immediately elicit a cytosolic free-calcium increase in target cells. This phenomenon is induced by low molecular weight amoebic metabolites and depends on a transmembrane influx of extracellular calcium. Morphological changes, cytoskeletal damage, cell death and cytolysis followed the elevation of cytosolic free-calcium levels. Calcium ions are very important for cell homeostasis, in fact, they control the functions of a variety of cellular responses, including secretion, cell proliferation and apoptosis. Our results suggest that the substained elevation of the cytosolic free-calcium in response to A. castellanii metabolites might play a fundamental role in target cell damage during Acanthamoeba infections. PMID:9245619

  20. Acanthamoeba castellanii : growth on human cell layers reactivates attenuated properties after prolonged axenic culture

    PubMed Central

    Koehsler, Martina; Leitsch, David; Duchêne, Michael; Nagl, Markus; Walochnik, Julia

    2009-01-01

    The free-living, but potentially pathogenic, bacteriovorous amoebae of the genus Acanthamoeba can be easily grown axenically in a laboratory culture. This, however, often leads to considerable losses in virulence, and encystment capacity, and to changes in drug susceptibility. We evaluated potential options for a reactivation of a number of physiological properties, attenuated by prolonged axenic laboratory culture, including encystment potential, protease activity, heat resistance, growth rates and drug susceptibility against N-chlorotaurine (NCT). Toward this end, a strain that had been grown axenically for 10 years was repeatedly passaged on human HEp-2 cell monolayers or treated with 5′-azacytidine (AzaC), a methyltransferase inhibitor, and trichostatin A (TSA), a histone deacetylase inhibitor, in order to uplift epigenetic gene regulation. Culture on human cell monolayers resulted in significantly enhanced encystment potentials and protease activities, and higher susceptibility against NCT, whereas the resistance against heat shock was not altered. Treatment with AzaC/TSA resulted in increased encystment rates and protease activities, indicating the participation of epigenetic mechanisms. However, lowered resistances against heat shock indicate that possible stress responses to AzaC/TSA have to be taken into account. Repeated growth on human cell monolayers appears to be a potential method to reactivate attenuated characteristics in Acanthamoeba. PMID:19732153

  1. Restriction fragment length polymorphisms of the DNA of selected Naegleria and Acanthamoeba amebae.

    PubMed

    McLaughlin, G L; Brandt, F H; Visvesvara, G S

    1988-09-01

    Fourteen strains of Naegleria fowleri, two strains of N. gruberi, and one strain each of N. australiensis, N. jadini, N. lovaniensis, Acanthamoeba sp., A. castellanii, A. polyphaga, and A. comandoni isolated from patients, soil, or water were characterized by restriction fragment length polymorphisms. Total cellular DNA (1 microgram) was digested with either HindIII, BglII, or EcoRI; separated on agarose gels; and stained with ethidium bromide. From 2 to 15 unusually prominent repetitive restriction fragment bands, totaling 15 to 50 kilobases in length and constituting probably more than 30% of the total DNA, were detected for all ameba strains. Each species displayed a characteristic pattern of repetitive restriction fragments. Digests of the four Acanthamoeba spp. displayed fewer, less intensely staining repetitive fragments than those of the Naegleria spp. All N. fowleri strains, whether isolated from the cerebrospinal fluid of patients from different parts of the world or from hot springs, had repetitive restriction fragment bands of similar total lengths (ca. 45 kilobases), and most repetitive bands displayed identical mobilities. However, polymorphic bands were useful in identifying particular isolates. Restriction fragment length polymorphism analysis generally was consistent with taxonomy based on studies of infectivity, morphology, isoenzyme patterns, and antibody reactivity and suggests that this technique may help classify amebae isolated from clinical specimens or from the environment.

  2. Morphological Features and In Vitro Cytopathic Effect of Acanthamoeba griffini Trophozoites Isolated from a Clinical Case

    PubMed Central

    González-Robles, Arturo; Salazar-Villatoro, Lizbeth; Omaña-Molina, Maritza; Reyes-Batlle, Maria; Martín-Navarro, Carmen M.; Lorenzo-Morales, Jacob

    2014-01-01

    Light and transmission electron microscopy observations are reported on the structure and in vitro cytopathic effect of Acanthamoeba griffini trophozoites isolated from a clinical case. Live trophozoites were moderately active with a remarkable pleomorphism which changed from ovoid to quite elongated shapes. When moving, amoebae formed cytoplasmic projections such as wide lamellae and acanthopodia of diverse size and thickness which contain a significant amount of actin. Ultrastructurally, the cytoplasm showed the main organelles found in other free-living amoebae. Coincubation of trophozoites with MDCK cell monolayers resulted in a local damage to target cells after 24 h of interaction, suggesting that the cytopathic effect is contact-dependent. By transmission electron microscopy, amoebae appeared to engulf small portions of the MDCK cells; however, the cells that were not in contact with trophozoites had an unaltered morphology. When epithelial monolayers were incubated with conditioned medium for 24 h, small areas of cell injury were also observed. The phylogenetical analysis as well as the sequencing of the acquired amplified product for the DF3 region of the amoebae isolate confirmed that it belongs to genotype T3, which includes other pathogenic amoebae; besides the activity of two drugs currently used against Acanthamoeba was tested on A. griffini. PMID:25313337

  3. Prophylactic Vancomycin Drops Reduce the Severity of Early Bacterial Keratitis in Keratoprosthesis

    PubMed Central

    Konstantopoulos, Aris; Tan, Xiao Wei; Goh, Gwendoline Tze Wei; Saraswathi, Padmanabhan; Chen, Liyan; Nyein, Chan Lwin; Zhou, Lei; Beuerman, Roger; Tan, Donald Tiang Hwee; Mehta, Jod

    2015-01-01

    Background Artificial cornea transplantation, keratoprosthesis, improves vision for patients at high risk of failure with human cadaveric cornea. However, post-operative infection can cause visual loss and implant extrusion in 3.2–17% of eyes. Long-term vancomycin drops are recommended following keratoprosthesis to prevent bacterial keratitis. Evidence, though, in support of this practice is poor. We investigated whether prophylactic vancomycin drops prevented bacterial keratitis in an animal keratoprosthesis model. Methodology Twenty-three rabbits were assigned either to a prophylactic group (n = 13) that received vancomycin 1.4% drops 5 times/day from keratoprosthesis implantation to sacrifice, or a non-prophylactic group (n = 10) that received no drops. All rabbits had Staphylococcus aureus inoculation into the cornea at 7–12 days post-implantation and were sacrificed at predetermined time-points. Prophylactic and non-prophylactic groups were compared with slit-lamp photography (SLP), anterior segment optical coherence tomography (AS-OCT), and histology, immunohistochemistry and bacterial quantification of excised corneas. Corneal vancomycin pharmacokinetics were studied in 8 additional rabbits. Results On day 1 post-inoculation, the median SLP score and mean±SEM AS-OCT corneal thickness (CT) were greater in the non-prophylactic than the prophylactic group (11 vs. 1, p = 0.049 and 486.9±61.2 vs. 327.4±37.1 μm, p = 0.029 respectively). On days 2 and 4, SLP scores and CT were not significantly different. Immunohistochemistry showed a greater CD11b+ve/non-CD11b+ve cell ratio in the non-prophylactic group (1.45 vs. 0.71) on day 2. Bacterial counts were not significantly different between the two groups. Corneal vancomycin concentration (2.835±0.383 μg/ml) exceeded minimum inhibitory concentration (MIC) for Staphylococcus aureus only after 16 days of vancomycin drops. Two of 3 rabbits still developed infection despite bacterial inoculation after 16 days of

  4. Interactions of Pseudomonas aeruginosa and Corynebacterium spp. with non-phagocytic brain microvascular endothelial cells and phagocytic Acanthamoeba castellanii.

    PubMed

    Siddiqui, Ruqaiyyah; Lakhundi, Sahreena; Khan, Naveed Ahmed

    2015-06-01

    Several lines of evidence suggest that Acanthamoeba interact with bacteria, which may aid in pathogenic bacterial transmission to susceptible hosts, and these interactions may have influenced evolution of bacterial pathogenicity. In this study, we tested if Gram-negative Pseudomonas aeruginosa and Gram-positive Corynebacterium spp. can associate/invade and survive inside Acanthamoeba castellanii trophozoites and cysts, as well as non-phagocytic human brain microvascular endothelial cells. The results revealed that both Corynebacterium spp. and P. aeruginosa were able to associate as well as invade and/or taken up by the phagocytic A. castellanii trophozoite. In contrast, P. aeruginosa exhibited higher association as well as invasion of non-phagocytic HBMEC compared with Corynebacterium spp. Notably, P. aeruginosa remained viable during the encystment process and exhibited higher levels of recovery from mature cysts (74.54 bacteria per amoebae) compared with Corynebacterium spp. (2.69 bacteria per amoeba) (P < 0.05). As Acanthamoeba cysts can be airborne, these findings suggest that Acanthamoeba is a potential vector in the transmission of P. aeruginosa to susceptible hosts. When bacterial-ridden amoebae were exposed to favourable (nutrient-rich) conditions, A. castellanii emerged as vegetative trophozoites and remained viable, and likewise viable P. aeruginosa were also observed but rarely any Corynebacterium spp. were observed. Correspondingly, P. aeruginosa but not Corynebacterium spp. exhibited higher cytotoxicity to non-phagocytic HBMEC, producing more than 75% cell death in 24 h, compared to 20% cell death observed with Corynebacterium spp. Additionally, it was observed that the bacterial conditioned medium had no negative effect on A. castellanii growth. Further characterization of amoebal and bacterial interactions will assist in identifying the role of Acanthamoeba in the transmission and evolution of pathogenic bacteria. PMID:25792227

  5. Mycotic Keratitis Due to Curvularia senegalensis and In Vitro Antifungal Susceptibilities of Curvularia spp.

    PubMed Central

    Guarro, Josep; Akiti, Tiyomi; Horta, Roberto Almada-; Morizot Leite-Filho, L. A.; Gené, Josepa; Ferreira-Gomes, Sueli; Aguilar, Carme; Ortoneda, Montserrat

    1999-01-01

    A case of mycotic keratitis due to Curvularia senegalensis is reported. This case represents the third known reported infection caused by this rare species. Fungal hyphae were detected in corneal scrapings, and repeated cultures were positive for this fungi. The patient was presumed cured after a corneal transplant and treatment with itraconazole, but the infection recurred and the patient is waiting for a keratoplasty. The in vitro antifungal susceptibilities of the case strain and another 24 strains belonging to seven species of Curvularia were tested for six antifungal agents. With the exception of flucytosine, and occasionally fluconazole, the other drugs assayed (amphotericin B, miconazole, itraconazole, and ketoconazole) were highly effective in vitro. PMID:10565956

  6. Investigation of the clinical features in filamentary keratitis in Hangzhou, east of China

    PubMed Central

    Chen, Siming; Ruan, Yimeng; Jin, Xiuming

    2016-01-01

    Abstract Filamentary keratitis (FK) is a chronic and recurrent disorder of the cornea. FK is reportedly associated with various kinds of ocular surface diseases or conditions. Until now, there have been lacks of studies based on quantitative sample analysis concerning FK incidence regularity and inducement characteristics at different ages. This was a retrospective study of 147 patients (162 eyes) with FK who had been continuously and completely recorded from August 2012 to August 2015 at the Second Affiliated Hospital of Zhejiang University in Hangzhou, east of China. Our results suggest that the causative factors of FK varied at different ages and the distribution of filaments on the corneal surface was also diverse with different inducements. By exploring the frequency and clinical features of FK, we believe that the findings from our research will be clinically significant and aid in the early prevention and treatment guidance of the disease. PMID:27583881

  7. Oral squamous cell carcinoma in a patient with keratitis-ichthyosis-deafness syndrome: a rare case.

    PubMed

    Homeida, L; Wiley, R T; Fatahzadeh, M

    2015-04-01

    Keratitis-ichthyosis-deafness (KID) syndrome is a rare form of ectodermal dysplasia with significant visual and auditory impairment. Pathogenesis involves a mutation in the GJB2 gene, which encodes connexin-26, a protein in the epithelial gap junctions thought to be involved in the differentiation of ectodermally derived tissues. Affected patients are also at increased risk for the epithelial malignancies. To our knowledge, nearly 100 cases of KID syndrome, including 19 with squamous cell carcinoma (SCC) complications, have been reported worldwide. We report here a patient with KID syndrome who developed an ulcerative oral lesion causing him significant discomfort; he was subsequently diagnosed with oral SCC. We review the clinical presentation and symptomatology, including those affecting the oral cavity for this syndrome and highlight the importance of multidisciplinary collaboration and life-long screening aimed at prevention of the evolving complications.

  8. Nosocomial keratitis caused by methicillin-resistant Staphylococcus aureus: case report and preventative measures

    PubMed Central

    Braich, Puneet S.; Aggarwal, Shruti; Mukhtar, Sabrina; Almeida, David RP.

    2015-01-01

    A 47-year-old African-American woman was admitted to the intensive care unit of our community hospital for respiratory failure secondary to severe decompensated heart failure, requiring intubation. In the ensuing days, she developed a methicillin-resistant Staphylococcus aureus (MRSA) infection of the cornea, despite no growth of MRSA in multiple blood, sputum, and urine cultures. This unexpected corneal infection complicated her hospital stay, and increased morbidity and disease-related cost. Risk factors, warning signs, and preventative measures for MRSA keratitis secondary to lagophthalmos (inability to completely close one's eyelids) are outlined in this case report. Implementing simple precautions such as taping eyelids shut or using artificial lubrication may reduce patient morbidity and disease-related costs. These recommendations are directed to non-ophthalmic clinicians who provide care to patients in settings where MRSA colonization is widespread. PMID:26486112

  9. Investigation of the clinical features in filamentary keratitis in Hangzhou, east of China.

    PubMed

    Chen, Siming; Ruan, Yimeng; Jin, Xiuming

    2016-08-01

    Filamentary keratitis (FK) is a chronic and recurrent disorder of the cornea. FK is reportedly associated with various kinds of ocular surface diseases or conditions. Until now, there have been lacks of studies based on quantitative sample analysis concerning FK incidence regularity and inducement characteristics at different ages. This was a retrospective study of 147 patients (162 eyes) with FK who had been continuously and completely recorded from August 2012 to August 2015 at the Second Affiliated Hospital of Zhejiang University in Hangzhou, east of China. Our results suggest that the causative factors of FK varied at different ages and the distribution of filaments on the corneal surface was also diverse with different inducements.By exploring the frequency and clinical features of FK, we believe that the findings from our research will be clinically significant and aid in the early prevention and treatment guidance of the disease. PMID:27583881

  10. In the aftermath of the Fusarium keratitis outbreak: What have we learned?

    PubMed Central

    Epstein, Arthur B

    2007-01-01

    The 2005–2006 outbreak of Fusarium keratitis associated with soft hydrophilic contact lens wear was both unprecedented and unexpected. More than 250 cases have been reported worldwide that have primarily been associated with Bausch & Lomb ReNu with MoistureLoc and, more recently, with Bausch and Lomb ReNu MultiPlus multipurpose contact lens disinfecting solutions. This article documents the outbreak, presenting the time line and the historical and scientific basis for its occurrence. Underlying causes are explored including likely mechanisms of contamination and subsequent corneal infection. Thorough exploration of this unique occurrence affords the opportunity to examine contact lens and lens care actions and interactions and to develop greater understanding of possible associated risks and ways to minimize them. PMID:19668512

  11. Acyclovir-resistant corneal HSV-1 isolates from patients with herpetic keratitis.

    PubMed

    Duan, Rui; de Vries, Rory D; Osterhaus, Albert D M E; Remeijer, Lies; Verjans, Georges M G M

    2008-09-01

    The prevalence and molecular characteristics of isolates from 173 immunocompetent patients with herpetic keratitis (HK) who were infected with acyclovir (ACV)-resistant (ACV(R)) corneal herpes simplex virus (HSV)-1 was determined. Isolates from 11 (6.4%) of the patients were ACV(R), and 9 of these 11 patients were refractory to therapy with ACV; the ACV(R) isolates from 5 and 1 of these 9 patients were cross-resistant to gancyclovir and to both gancyclovir and foscarnet, respectively. Of the 11 ACV(R) isolates, 10 had, in the thymidine kinase gene, mutations that presumably conferred the ACV(R) phenotype. These data demonstrate a relatively high prevalence of corneal HSV-1 ACV(R) isolates in patients with HK, which emphasizes the need to monitor for ACV susceptibility in patients with HK who are refractory to therapy with ACV.

  12. Inhibition of Ataxia Telangiectasia Mutated (ATM) Kinase Suppresses Herpes Simplex Virus Type 1 (HSV-1) Keratitis

    PubMed Central

    Alekseev, Oleg; Donovan, Kelly; Azizkhan-Clifford, Jane

    2014-01-01

    Purpose. Herpes keratitis (HK) remains the leading cause of cornea-derived blindness in the developed world, despite the availability of effective antiviral drugs. Treatment toxicity and the emergence of drug resistance highlight the need for additional therapeutic approaches. This study examined ataxia telangiectasia mutated (ATM), an apical kinase in the host DNA damage response, as a potential new target for the treatment of HK. Methods. Small molecule inhibitor of ATM (KU-55933) was used to treat herpes simplex virus type 1 (HSV-1) infection in three experimental models: (1) in vitro—cultured human corneal epithelial cells, hTCEpi, (2) ex vivo—organotypically explanted human and rabbit corneas, and (3) in vivo—corneal infection in young C57BL/6J mice. Infection productivity was assayed by plaque assay, real-time PCR, Western blot, and disease scoring. Results. Robust ATM activation was detected in HSV-1-infected human corneal epithelial cells. Inhibition of ATM greatly suppressed viral replication in cultured cells and in explanted human and rabbit corneas, and reduced the severity of stromal keratitis in mice. The antiviral effect of KU-55933 in combination with acyclovir was additive, and KU-55933 suppressed replication of a drug-resistant HSV-1 strain. KU-55933 caused minimal toxicity, as monitored by clonogenic survival assay and fluorescein staining. Conclusions. This study identifies ATM as a potential target for the treatment of HK. ATM inhibition by KU-55933 reduces epithelial infection and stromal disease severity without producing appreciable toxicity. These findings warrant further investigations into the DNA damage response as an area for therapeutic intervention in herpetic ocular diseases. PMID:24370835

  13. The anterior segment disorder autosomal dominant keratitis is linked to the Aniridia/PAX-6 gene

    SciTech Connect

    Mirzayans, F.; Pearce, W.G.; Mah, T.S.

    1994-09-01

    Autosomal dominant keratitis (ADK) is an eye disease characterized by anterior stromal corneal opacification and vascularization in the peripheral cornea. Progression into the central cornea may compromise visual acuity. Other anterior segment features include minimal radial defects of the iris stroma. Posterior segment involvement is characterized by foveal hypoplasia with minimal effect on visual acuity. Aniridia is a second autosomal dominantly inherited ocular disorder defined by structural defects of the iris, frequently severe enough to cause an almost complete absence of iris. This may be accompanied by other anterior segment manifestations, including cataract and keratitis. Posterior segment involvement in aniridia is characterized by foveal hypoplasia resulting in a highly variable impairment of visual acuity, often with nystagmus. Aniridia is usually inherited as an autosomal dominant disease and occurs in 1 in 50,000 to 100,000 people. Aniridia has been shown to result from mutations in PAX-6, a gene thought to regulate fetal eye development. The similar clinical findings in ADK and aniridia, with the similar patterns of inheritance, compelled us to investigate if these two ocular disorders are variants of the same genetic disorder. We have tested for linkage between PAX-6 and ADK within an ADK family with 33 members over four generations, including 11 affected individuals. Linkage studies reveal that D11S914 (located within 3 cM of PAX-6) does not recombine with ADK (LOD score 3.61; {theta} = 0.00), consistent with PAX-6 mutations being responsible for ADK. Direct sequencing of PAX-6 RT-PCR products from ADK patients is underway to identify the mutation within the PAX-6 gene that results in ADK. The linkage of PAX-6 with ADK, along with a recent report that mutations in PAX-6 also underlie Peter`s anomaly, implicates PAX-6 widely in anterior segment malformations.

  14. Development and clinical evaluation of clotrimazole-β-cyclodextrin eyedrops for the treatment of fungal keratitis.

    PubMed

    Rasool, Bazigha K Abdul; Salmo, Hiba M

    2012-09-01

    Fungal keratitis is a serious corneal disease that may result in loss of vision. There are limited treatment options available in Iraqi eye hospitals which might be the main reason behind the poor prognosis of many cases. The purpose of this study was to prepare and pharmaceutically evaluate clotrimazole-β-cyclodextrin (CTZ-β-CD) eyedrops then clinically assess its therapeutic efficacy on fungal keratitis compared with extemporaneous amphotericin B eyedrops (0.5% w/v). A CTZ-β-CD ophthalmic solution was prepared and evaluated by various physicochemical, microbiological, and biological tests. The prepared formula was stable in 0.05 M phosphate buffer pH 7.0 at 40 ± 2°C and 75 ± 5% RH for a period of 6 months. Light has no significant effect on the formula's stability. The CTZ-β-CD eyedrops efficiently complied with the isotonicity, sterility, and antimicrobiological preservative effectiveness tests. Results of the clinical study revealed that 20 (80%) patients showed a favorable response to the CTZ-β-CD eyedrops, while 16 patients (64%) exhibited a favorable response to amphotericin B (P > 0.05). The mean course of treatment was significantly (P < 0.05) less in the CTZ treatment group than in the amphotericin group (21.5 ± 5.2 vs. 28.3 ± 6.4 days, respectively). The CTZ formulation was significantly (P < 0.05) more effective in the management of severe cases and also against Candida sp. than amphotericin B. There was no significant difference (P < 0.05) between both therapies against filamentous fungi. The CTZ-β-CD formulation can be used alternatively to other ophthalmic antimycotic treatment options in developing countries where stability, cost, or efficacy is a limiting factor.

  15. Topical levofloxacin 1.5% overcomes in vitro resistance in rabbit keratitis models

    PubMed Central

    Kowalski, Regis P.; Romanowski, Eric G.; Mah, Francis S.; Shanks, Robert M. Q.; Gordon, Y. J.

    2016-01-01

    Purpose To determine whether topical levofloxacin 1.5% will successfully treat both levofloxacin-resistant and susceptible Staphylococcus aureus (SA) and Pseudomonas aeruginosa (PA) in rabbit keratitis models. Methods For levofloxacin-resistant and susceptible SA, respectively, 32 New Zealand White (NZW) rabbits were intrastromally injected with 1000 colony-forming units (CFU). After 4 hr, the corneas of eight rabbits were homogenized to determine onset CFU/ml. Twenty-four rabbits were divided into three treatments: levofloxacin, vancomycin (cefazolin for levofloxacin-susceptible SA) and saline. Twenty-one drops were administered over 5 hr. One hour post-treatment, the corneas were homogenized for CFU/ml. For levofloxacin-resistant and susceptible PA, respectively, 32 NZW rabbits were intrastromally injected with 1000 CFU. After 16 hr, the corneas of eight rabbits were homogenized for CFU/ml. Twenty-four rabbits were divided into three treatments: levofloxacin, tobramycin (ciprofloxacin for levofloxacin-susceptible PA) and saline. Nineteen drops were administered over 8 hr. One hour post-treatment, the corneas were homogenized for CFU/ml. The CFU/ml data were analysed for sterilization and non-parametrically for reduction. Results Levofloxacin 1.5% significantly reduced more (p < 0.05) levofloxacin-resistant SA than vancomycin; was equivalent to cefazolin (p > 0.05) for levofloxacin-susceptible SA; was equivalent to tobramycin for levofloxacin-resistant PA; was equivalent to ciprofloxacin for levofloxacin-susceptible PA; and significantly reduced more SA and PA than saline and onset. Levofloxacin 1.5% sterilized the corneas in the levofloxacin-resistant and susceptible PA groups (32/32) and levofloxacin-susceptible SA group (16/16), but not the levofloxacin-resistant SA group (0/16). Conclusion Levofloxacin 1.5% was effective for reducing SA and PA in the rabbit keratitis models regardless of in vitro resistance. PMID:20456251

  16. Sterile keratitis after combined riboflavin-UVA corneal collagen cross-linking for keratoconus

    PubMed Central

    Lam, F C; Geourgoudis, P; Nanavaty, M A; Khan, S; Lake, D

    2014-01-01

    Purpose To investigate patient risk factors and to look for potential causes of sterile infiltrates following an unexpected cluster of sterile keratitis after a routine collagen cross-linking (CXL) list. Methods The records of all 148 cases of CXL were reviewed retrospectively. The equipment and solutions used and our clinic's standard operating procedure for CXL were reviewed. An in-vitro experiment to explore the variation in ultraviolet A (UVA) irradiance from fluctuations in the working distance of the UVA lamp was conducted. Results The four patients who developed sterile infiltrates had steeper maximum corneal curvatures (68.0±7.3 D) and thinner pachymetry (389.9±49.0 μm) than the 144 who did not (57.0±8.2 D, P=0.05; 454.6±45.4 μm, P=0.08). A corneal curvature of >60 Dand a pachymetry of <425 μm were significant risk factors. All four affected cases obtained a complete resolution with topical antibiotics and steroids. The unaided VA and the maximum K improved from their pre-operative levels in three out of four patients. A 2-mm reduction in distance of the VEGA C.B.M. X-Linker from a treated surface increased irradiance to 3.5–3.7 mW/cm2, which is above the threshold for endothelial toxicity. Conclusion Patients with thinner and steeper corneas are at an increased risk of developing sterile keratitis. The visual outcomes despite this complication are good. PMID:25104740

  17. Pseudomonas aeruginosa Infectious Keratitis in a High Oxygen Transmissible Rigid Contact Lens Rabbit Model

    PubMed Central

    Wei, Cynthia; Zhu, Meifang; Petroll, W. Matthew; Robertson, Danielle M.

    2014-01-01

    Purpose. To establish a rabbit model of infectious Pseudomonas aeruginosa keratitis using ultrahigh oxygen transmissible rigid lenses and characterize the frequency and severity of infection when compared to a non–oxygen transmissible lens material. Methods. Rabbits were fit with rigid lenses composed of ultrahigh and non–oxygen transmissible materials. Prior to wear, lenses were inoculated with an invasive corneal isolate of P. aeruginosa stably conjugated to green fluorescent protein (GFP). Corneas were examined before and after lens wear using a modified Heidelberg Rostock Tomograph in vivo confocal microscope. Viable bacteria adherent to unworn and worn lenses were assessed by standard plate counts. The presence of P. aeruginosa-GFP and myeloperoxidase-labeled neutrophils in infected corneal tissue was evaluated using laser scanning confocal microscopy. Results. The frequency and severity of infectious keratitis was significantly greater with inoculated ultrahigh oxygen transmissible lenses. Infection severity was associated with increasing neutrophil infiltration and in severe cases, corneal melting. In vivo confocal microscopic analysis of control corneas following lens wear confirmed that hypoxic lens wear was associated with mechanical surface damage, whereas no ocular surface damage was evident in the high-oxygen lens group. Conclusions. These data indicate that in the absence of adequate tear clearance, the presence of P. aeruginosa trapped under the lens overrides the protective effects of oxygen on surface epithelial cells. These findings also suggest that alternative pathophysiological mechanisms exist whereby changes under the lens in the absence of frank hypoxic damage result in P. aeruginosa infection in the otherwise healthy corneal epithelium. PMID:25125601

  18. Occurrence and molecular characterization of free-living amoeba species (Acanthamoeba, Hartmannella, and Saccamoeba limax) in various surface water resources of Iran.

    PubMed

    Mahmoudi, Mohammad Reza; Rahmati, Behnaz; Seyedpour, Seyed Hosssen; Karanis, Panagiotis

    2015-12-01

    This study was conducted to determine the presence and molecular identity of Acanthamoeba species in the surface water resources of four provinces in Iran, namely Guilan, Mazandaran (North of Iran), Alborz, and Tehran (capital city), using culture- and molecular-based methods. During March to November 2014, 49 surface water samples were collected from environmental water sources-the distinct surface waters of Guilan, Mazandaran, Alborz, and Tehran provinces, in Iran. For the isolation of Acanthamoeba species, approximately 500 ml of the water samples were filtered through a cellulose nitrate membrane with a pore size of 0.45 μ. The filter was transferred onto non-nutrient agar plates seeded with Gram-negative bacteria (Escherichia coli) as a food source. The presence of Acanthamoeba was confirmed by the genus-specific primer pair JDP1 and 2, and/or NA primers were used to identify Acanthamoeba and certain other free-living amoebae. In total, 38 out of 49 samples were positive by culture and/or PCR for Acanthamoeba and other free-living amoebae from all three provinces. By sequencing the positive isolates, the strains were shown to belong to Acanthamoeba (16 isolates belonged to T4 and 2 isolates belonged to T5), Hartmannella vermiformis (3/24), and Saccamoeba limax (2/24). The T4 and T5 genotypes were detected in Guilan and Mazandaran provinces. Two isolates from Guilan and Tehran provinces belonged to S. limax, and H. vermiformis was detected in Guilan province. The results of this study highlight the need to pay more attention to free-living amoebae, as human activity was observed in all of the localities from which these samples were taken. These surface waters can be potential sources for the distribution and transmission of pathogenic Acanthamoeba in the study areas, and free-living amoebas (FLA) (particularly the Acanthamoeba species) can serve as hosts for and vehicles of various microorganisms.

  19. AkF ¯ chiral gauge theories

    NASA Astrophysics Data System (ADS)

    Shi, Yan-Liang; Shrock, Robert

    2015-11-01

    We study asymptotically free chiral gauge theories with an SU (N ) gauge group and chiral fermions transforming according to the antisymmetric rank-k tensor representation, Ak≡[k ]N , and the requisite number, nF ¯, of copies of fermions in the conjugate fundamental representation, F ¯ ≡[1] ¯ N , to render the theories anomaly-free. We denote these as AkF ¯ theories. We take N ≥2 k +1 so that nF ¯≥1 . The A2F ¯ theories form an infinite family with N ≥5 , but we show that the A3F ¯ and A4F ¯ theories are only asymptotically free for N in the respective ranges 7 ≤N ≤17 and 9 ≤N ≤11 , and that there are no asymptotically free AkF ¯ theories with k ≥5 . We investigate the types of ultraviolet to infrared evolution for these AkF ¯ theories and find that, depending on k and N , they may lead to a non-Abelian Coulomb phase, or may involve confinement with massless gauge-singlet composite fermions, bilinear fermion condensation with dynamical gauge and global symmetry breaking, or formation of multifermion condensates that preserve the gauge symmetry. We also show that there are no asymptotically free, anomaly-free SU (N ) SkF ¯ chiral gauge theories with k ≥3 , where Sk denotes the rank-k symmetric representation.

  20. Fungal Keratitis

    MedlinePlus

    ... 2016 Over-the-Counter Costume Contacts May Contain Chemicals Harmful to Eyes Oct 05, 2015 Three Rising Stars Bullied for Childhood ... Academy Jobs at the Academy Financial Relationships with Industry Medical Disclaimer Privacy Policy Terms of Service For ...

  1. Herpes Keratitis

    MedlinePlus

    ... Related Over-the-Counter Costume Contacts May Contain Chemicals Harmful to Eyes Oct 05, 2015 Contact Lens Horror Stories: Poor Care Can Lead ... at the Academy Financial Relationships with Industry Medical Disclaimer Privacy Policy Terms of Service For ...

  2. 40 CFR Appendixes A-K to Part 51 - [Reserved

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 2 2011-07-01 2011-07-01 false A Appendixes A-K to Part 51 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR PROGRAMS REQUIREMENTS FOR PREPARATION, ADOPTION, AND SUBMITTAL OF IMPLEMENTATION PLANS Appendixes A-K to Part 51...

  3. 40 CFR Appendixes A-K to Part 51 - [Reserved

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 2 2014-07-01 2014-07-01 false A Appendixes A-K to Part 51 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR PROGRAMS REQUIREMENTS FOR PREPARATION, ADOPTION, AND SUBMITTAL OF IMPLEMENTATION PLANS Appendixes A-K to Part 51...

  4. 40 CFR Appendixes A-K to Part 51 - [Reserved

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 2 2010-07-01 2010-07-01 false A Appendixes A-K to Part 51 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR PROGRAMS REQUIREMENTS FOR PREPARATION, ADOPTION, AND SUBMITTAL OF IMPLEMENTATION PLANS Appendixes A-K to Part 51...

  5. 40 CFR Appendixes A-K to Part 51 - [Reserved

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 2 2013-07-01 2013-07-01 false A Appendixes A-K to Part 51 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR PROGRAMS REQUIREMENTS FOR PREPARATION, ADOPTION, AND SUBMITTAL OF IMPLEMENTATION PLANS Appendixes A-K to Part 51...

  6. 40 CFR Appendixes A-K to Part 51 - [Reserved

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 2 2012-07-01 2012-07-01 false A Appendixes A-K to Part 51 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR PROGRAMS REQUIREMENTS FOR PREPARATION, ADOPTION, AND SUBMITTAL OF IMPLEMENTATION PLANS Appendixes A-K to Part 51...

  7. 46 CFR 7.180 - Kotzebue Sound, AK.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 1 2010-10-01 2010-10-01 false Kotzebue Sound, AK. 7.180 Section 7.180 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY PROCEDURES APPLICABLE TO THE PUBLIC BOUNDARY LINES Alaska § 7.180 Kotzebue Sound, AK. A line drawn from Cape Espenberg Light to latitude 66°52′ N. longitude 163°28′ W.;...

  8. 46 CFR 7.180 - Kotzebue Sound, AK.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 1 2013-10-01 2013-10-01 false Kotzebue Sound, AK. 7.180 Section 7.180 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY PROCEDURES APPLICABLE TO THE PUBLIC BOUNDARY LINES Alaska § 7.180 Kotzebue Sound, AK. A line drawn from Cape Espenberg Light to latitude 66°52′ N. longitude 163°28′ W.;...

  9. 46 CFR 7.180 - Kotzebue Sound, AK.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 1 2012-10-01 2012-10-01 false Kotzebue Sound, AK. 7.180 Section 7.180 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY PROCEDURES APPLICABLE TO THE PUBLIC BOUNDARY LINES Alaska § 7.180 Kotzebue Sound, AK. A line drawn from Cape Espenberg Light to latitude 66°52′ N. longitude 163°28′ W.;...

  10. 46 CFR 7.180 - Kotzebue Sound, AK.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 1 2014-10-01 2014-10-01 false Kotzebue Sound, AK. 7.180 Section 7.180 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY PROCEDURES APPLICABLE TO THE PUBLIC BOUNDARY LINES Alaska § 7.180 Kotzebue Sound, AK. A line drawn from Cape Espenberg Light to latitude 66°52′ N. longitude 163°28′ W.;...

  11. 46 CFR 7.180 - Kotzebue Sound, AK.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 1 2011-10-01 2011-10-01 false Kotzebue Sound, AK. 7.180 Section 7.180 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY PROCEDURES APPLICABLE TO THE PUBLIC BOUNDARY LINES Alaska § 7.180 Kotzebue Sound, AK. A line drawn from Cape Espenberg Light to latitude 66°52′ N. longitude 163°28′ W.;...

  12. Raman spectroscopic study on the excystation process in a single unicellular organism amoeba (Acanthamoeba polyphaga).

    PubMed

    Lin, Yu-Chung; Perevedentseva, Elena; Cheng, Chia-Liang

    2015-05-01

    An in vivo Raman spectroscopic study of amoeba (Acanthamoeba polyphaga) is presented. The changes of the spectra during the amoeba cyst activation and excystation are analyzed. The spectra show the changes of the relative intensities of bands corresponding to protein, lipid, and carotenoid components during cyst activation. The presence of carotenoids in the amoeba is observed via characteristic Raman bands. These signals in the Raman spectra are intense in cysts but decrease in intensity with cyst activation and exhibit a correlation with the life cycle of amoeba. This work demonstrates the feasibility of using Raman spectroscopy for the detection of single amoeba microorganisms in vivo and for the analysis of the amoeba life activity. The information obtained may have implications for the estimation of epidemiological situations and for the diagnostics and prognosis of the development of amoebic inflammations.

  13. An intracellular replication niche for Vibrio cholerae in the amoeba Acanthamoeba castellanii.

    PubMed

    Van der Henst, Charles; Scrignari, Tiziana; Maclachlan, Catherine; Blokesch, Melanie

    2016-04-01

    Vibrio cholerae is a human pathogen and the causative agent of cholera. The persistence of this bacterium in aquatic environments is a key epidemiological concern, as cholera is transmitted through contaminated water. Predatory protists, such as amoebae, are major regulators of bacterial populations in such environments. Therefore, we investigated the interaction between V. cholerae and the amoeba Acanthamoeba castellanii at the single-cell level. We observed that V. cholerae can resist intracellular killing. The non-digested bacteria were either released or, alternatively, established a replication niche within the contractile vacuole of A. castellanii. V. cholerae was maintained within this compartment even upon encystment. The pathogen ultimately returned to its aquatic habitat through lysis of A. castellanii, a process that was dependent on the production of extracellular polysaccharide by the pathogen. This study reinforces the concept that V. cholerae is a facultative intracellular bacterium and describes a new host-pathogen interaction.

  14. Raman spectroscopic study on the excystation process in a single unicellular organism amoeba (Acanthamoeba polyphaga)

    NASA Astrophysics Data System (ADS)

    Lin, Yu-Chung; Perevedentseva, Elena; Cheng, Chia-Liang

    2015-05-01

    An in vivo Raman spectroscopic study of amoeba (Acanthamoeba polyphaga) is presented. The changes of the spectra during the amoeba cyst activation and excystation are analyzed. The spectra show the changes of the relative intensities of bands corresponding to protein, lipid, and carotenoid components during cyst activation. The presence of carotenoids in the amoeba is observed via characteristic Raman bands. These signals in the Raman spectra are intense in cysts but decrease in intensity with cyst activation and exhibit a correlation with the life cycle of amoeba. This work demonstrates the feasibility of using Raman spectroscopy for the detection of single amoeba microorganisms in vivo and for the analysis of the amoeba life activity. The information obtained may have implications for the estimation of epidemiological situations and for the diagnostics and prognosis of the development of amoebic inflammations.

  15. An intracellular replication niche for Vibrio cholerae in the amoeba Acanthamoeba castellanii

    PubMed Central

    Van der Henst, Charles; Scrignari, Tiziana; Maclachlan, Catherine; Blokesch, Melanie

    2016-01-01

    Vibrio cholerae is a human pathogen and the causative agent of cholera. The persistence of this bacterium in aquatic environments is a key epidemiological concern, as cholera is transmitted through contaminated water. Predatory protists, such as amoebae, are major regulators of bacterial populations in such environments. Therefore, we investigated the interaction between V. cholerae and the amoeba Acanthamoeba castellanii at the single-cell level. We observed that V. cholerae can resist intracellular killing. The non-digested bacteria were either released or, alternatively, established a replication niche within the contractile vacuole of A. castellanii. V. cholerae was maintained within this compartment even upon encystment. The pathogen ultimately returned to its aquatic habitat through lysis of A. castellanii, a process that was dependent on the production of extracellular polysaccharide by the pathogen. This study reinforces the concept that V. cholerae is a facultative intracellular bacterium and describes a new host–pathogen interaction. PMID:26394005

  16. Isolation of two strains of Acanthamoeba castellanii from human tissue and their pathogenicity and isoenzyme profiles.

    PubMed Central

    Visvesvara, G S; Mirra, S S; Brandt, F H; Moss, D M; Mathews, H M; Martinez, A J

    1983-01-01

    Two strains of amoebae, one (CDC:0180:1) from the lung tissue of a patient who died of granulomatous amoebic encephalitis and the other (CDC:0179:1) from the debrided tissue of a mandibular autograft, were isolated and identified as Acanthamoeba castellanii based on the morphological and immunofluorescent staining characteristics of the trophozoites and cysts. Both strains of amoebae caused cytopathic effects in mammalian cell cultures and destroyed the cell sheet. However, only the CDC:0180:1 strain, on intranasal instillation into mice, produced the disease manifested by ruffled fur and aimless wandering, followed by coma and death within 30 days. The CDC:0180:1 strain also differed consistently from CDC:0179:1 and another nonpathogenic A. castellanii strain (ATCC 30,011) in isoenzyme makeup, a dissimilarity which probably reflects its pathogenic potential. Images PMID:6655045

  17. A Combination of Intrastromal and Intracameral Injections of Amphotericin B in the Treatment of Severe Fungal Keratitis

    PubMed Central

    Hu, Jianzhang; Zhang, Jingjin; Li, Yanling; Han, Xiaoli; Zheng, Weidong; Yang, Juan

    2016-01-01

    Purpose. To evaluate the efficacy of a combination of intrastromal and intracameral injections of amphotericin B in the treatment of severe recalcitrant fungal keratitis. Methods. Patients with severe fungal keratitis who were resistant to conventional antifungal medical treatments and needed potential surgical intervention were recruited at the First Affiliated Hospital of Fujian Medical University between January 2012 and July 2013. The patients were treated with a combination of intrastromal and intracameral injections of amphotericin B (25 μg/mL and 50 μg/mL, resp.). Selectively repeated injections were performed as necessary. The efficacy, complications, and outcome were evaluated. Results. Nine patients (9 eyes) were involved in this study. All 9 cases responded favorably, and the clinical appearance of serious corneal damage and intraocular extension was resolved after the treatment. Four eyes required only 1 injection, and 5 eyes required repeated injections. Seven corneal ulcers healed with leucoma, and 2 healed with adherent leucoma. All of our cases had a marked increase in the anterior chamber reaction and pain immediately after the injection. There was no obvious clinical evidence of corneal or lenticular toxicity in any patient. Conclusions. A combination of intrastromal and intracameral injections of amphotericin B may be safe and effective for the treatment of severe fungal keratitis that is resistant to conventional therapy. PMID:27721986

  18. Long-term Outcomes of Amniotic Membrane Transplantation in Contact Lens-Induced Pseudomonas Keratitis with Impending Corneal Perforation

    PubMed Central

    Mohammadpour, Mehrdad; Sabet, Fatemeh Alsadat

    2016-01-01

    Purpose: To report the anatomical and visual outcomes of double layered amniotic membrane transplantation (AMT) in eyes with advanced Pseudomonas keratitis leading to Descemetocele formation. Methods: This prospective interventional case series included 6 eyes of 6 female patients with pseudomonas keratitis caused by contact lens-induced infection who underwent double layered AMT. Surgery was performed after the ulcers were found to be poorly responsive to antibiotics, and severe thinning or Descemetocele had developed. All patients underwent a complete examination pre- and postoperatively, as well as anterior segment optical coherence tomography (OCT) and pachymetry or Orbscan after the procedure. Results: Mean follow-up period was 24 months. There was neither frank corneal perforation nor a need for emergent corneal transplantation in any of the eyes. All patients had visual acuity of hand motions before the procedure which improved to 20/50 to 20/30 three months after surgery. No surgical or postoperative complication occurred in this series. Conclusion: Double layered AMT may result in acceptable anatomical outcomes in patients with advanced Pseudomonas keratitis with Descemetocele formation and can eliminate the need for emergent corneal transplantation. PMID:27195083

  19. Public health implications of Acanthamoeba and multiple potential opportunistic pathogens in roof-harvested rainwater tanks.

    PubMed

    Hamilton, K A; Ahmed, W; Palmer, A; Sidhu, J P S; Hodgers, L; Toze, S; Haas, C N

    2016-10-01

    A study of six potential opportunistic pathogens (Acanthamoeba spp., Legionella spp., Legionella longbeachae, Pseudomonas aeruginosa, Mycobacterium avium and Mycobacterium intracellulare) and an accidental human pathogen (Legionella pneumophila) in 134 roof-harvested rainwater (RHRW) tank samples was conducted using quantitative PCR (qPCR). All five opportunistic pathogens and accidental pathogen L. pneumophila were detected in rainwater tanks except Legionella longbeachae. Concentrations ranged up to 3.1×10(6) gene copies per L rainwater for Legionella spp., 9.6×10(5) gene copies per L for P. aeruginosa, 6.8×10(5) gene copies per L for M. intracellulare, 6.6×10(5) gene copies per L for Acanthamoeba spp., 1.1×10(5) gene copies per L for M. avium, and 9.8×10(3) gene copies per L for L. pneumophila. Among the organisms tested, Legionella spp. (99% tanks) were the most prevalent followed by M. intracellulare (78%). A survey of tank-owners provided data on rainwater end-uses. Fecal indicator bacteria (FIB) Escherichia coli and Enterococcus spp. were enumerated using culture-based methods, and assessed for correlations with opportunistic pathogens and L. pneumophila tested in this study. Opportunistic pathogens did not correlate well with FIB except E. coli vs. Legionella spp. (tau=0.151, P=0.009) and E. coli vs. M. intracellulare (tau=0.14, P=0.015). However, M. avium weakly correlated with both L. pneumophila (Kendall's tau=0.017, P=0.006) and M. intracellulare (tau=0.088, P=0.027), and Legionella spp. also weakly correlated with M. intracellulare (tau=0.128, P=0.028). The presence of these potential opportunistic pathogens in tank water may present health risks from both the potable and non-potable uses documented from the current survey data. PMID:27336236

  20. Amino Acid Uptake and Metabolism of Legionella pneumophila Hosted by Acanthamoeba castellanii*

    PubMed Central

    Schunder, Eva; Gillmaier, Nadine; Kutzner, Erika; Herrmann, Vroni; Lautner, Monika; Heuner, Klaus; Eisenreich, Wolfgang

    2014-01-01

    Legionella pneumophila survives and replicates within a Legionella-containing vacuole (LCV) of amoebae and macrophages. Less is known about the carbon metabolism of the bacteria within the LCV. We have now analyzed the transfer and usage of amino acids from the natural host organism Acanthamoeba castellanii to Legionella pneumophila under in vivo (LCV) conditions. For this purpose, A. castellanii was 13C-labeled by incubation in buffer containing [U-13C6]glucose. Subsequently, these 13C-prelabeled amoebae were infected with L. pneumophila wild type or some mutants defective in putative key enzymes or regulators of carbon metabolism. 13C-Isotopologue compositions of amino acids from bacterial and amoebal proteins were then determined by mass spectrometry. In a comparative approach, the profiles documented the efficient uptake of Acanthamoeba amino acids into the LCV and further into L. pneumophila where they served as precursors for bacterial protein biosynthesis. More specifically, A. castellanii synthesized from exogenous [U-13C6]glucose unique isotopologue mixtures of several amino acids including Phe and Tyr, which were also observed in the same amino acids from LCV-grown L. pneumophila. Minor but significant differences were only detected in the isotopologue profiles of Ala, Asp, and Glu from the amoebal or bacterial protein fractions, respectively, indicating partial de novo synthesis of these amino acids by L. pneumophila. The similar isotopologue patterns in amino acids from L. pneumophila wild type and the mutants under study reflected the robustness of amino acid usage in the LCV of A. castellannii. PMID:24904060

  1. Resistance of Acanthamoeba Cysts to Disinfection Treatments Used in Health Care Settings▿

    PubMed Central

    Coulon, Céline; Collignon, Anne; McDonnell, Gerald; Thomas, Vincent

    2010-01-01

    Free-living amoebae that belong to the genus Acanthamoeba are widespread in the environment, including water. They are responsible for human infections and can host pathogenic microorganisms. Under unfavorable conditions, they form cysts with high levels of resistance to disinfection methods, thus potentially representing a threat to public health. In the present study we evaluated the efficacies of various biocides against trophozoites and cysts of several Acanthamoeba strains. We demonstrated that disinfectant efficacy varied depending on the strains tested, with environmental strains demonstrating greater resistance than collection strains. Trophozoites were inactivated by all treatments except those using glutaraldehyde as an active compound: for these treatments, we observed resistance even after 30 min exposure. Cysts resisted many treatments, including certain conditions with glutaraldehyde and other biocides. Moist heat at 55°C was not efficient against cysts, whereas exposure at 65°C was. Several chemical formulations containing peracetic acid, hydrogen peroxide, or ortho-phthalaldehyde presented greater efficacy than glutaraldehyde, as did ethanol and sodium hypochlorite; however, some of these treatments required relatively long incubation times to achieve cyst inactivation. Amoebal cysts can be highly resistant to some high-level disinfectants, which has implications for clinical practice. These results highlight the need to consider the effective disinfection of protozoa in their vegetative and resistant forms due to their intrinsic resistance. This is important not only to prevent the transmission of protozoa themselves but also due to the risks associated with a range of microbial pathogens that are found to be associated intracellularly with these microorganisms. PMID:20519477

  2. Public health implications of Acanthamoeba and multiple potential opportunistic pathogens in roof-harvested rainwater tanks.

    PubMed

    Hamilton, K A; Ahmed, W; Palmer, A; Sidhu, J P S; Hodgers, L; Toze, S; Haas, C N

    2016-10-01

    A study of six potential opportunistic pathogens (Acanthamoeba spp., Legionella spp., Legionella longbeachae, Pseudomonas aeruginosa, Mycobacterium avium and Mycobacterium intracellulare) and an accidental human pathogen (Legionella pneumophila) in 134 roof-harvested rainwater (RHRW) tank samples was conducted using quantitative PCR (qPCR). All five opportunistic pathogens and accidental pathogen L. pneumophila were detected in rainwater tanks except Legionella longbeachae. Concentrations ranged up to 3.1×10(6) gene copies per L rainwater for Legionella spp., 9.6×10(5) gene copies per L for P. aeruginosa, 6.8×10(5) gene copies per L for M. intracellulare, 6.6×10(5) gene copies per L for Acanthamoeba spp., 1.1×10(5) gene copies per L for M. avium, and 9.8×10(3) gene copies per L for L. pneumophila. Among the organisms tested, Legionella spp. (99% tanks) were the most prevalent followed by M. intracellulare (78%). A survey of tank-owners provided data on rainwater end-uses. Fecal indicator bacteria (FIB) Escherichia coli and Enterococcus spp. were enumerated using culture-based methods, and assessed for correlations with opportunistic pathogens and L. pneumophila tested in this study. Opportunistic pathogens did not correlate well with FIB except E. coli vs. Legionella spp. (tau=0.151, P=0.009) and E. coli vs. M. intracellulare (tau=0.14, P=0.015). However, M. avium weakly correlated with both L. pneumophila (Kendall's tau=0.017, P=0.006) and M. intracellulare (tau=0.088, P=0.027), and Legionella spp. also weakly correlated with M. intracellulare (tau=0.128, P=0.028). The presence of these potential opportunistic pathogens in tank water may present health risks from both the potable and non-potable uses documented from the current survey data.

  3. The use of antimicrobial peptides in ophthalmology: an experimental study in corneal preservation and the management of bacterial keratitis.

    PubMed Central

    Mannis, Mark J

    2002-01-01

    PURPOSE: Bacterial keratitis is an ocular infection with the potential to cause significant visual impairment. Increasing patterns of antibiotic resistance have necessitated the development of new antimicrobial agents for use in bacterial keratitis and other serious ocular infections. With a view to exploring the use of novel antimicrobial peptides in the management of ocular infection, we performed a series of experiments using synthetic antimicrobial peptides designed for the eradication of common and serious ophthalmic pathogens. METHODS: Experiments were performed with three clinical ocular isolates--Pseudomonas aeruginosa, Staphylococcus aureus, and Staphylococcus epidermidis--in three experimental settings: (1) in vitro in a controlled system of 10 mM sodium phosphate buffer, (2) in vitro in modified chondroitin sulfate-based corneal preservation media (Optisol), and (3) in an in vivo animal model (rabbit) simulating bacterial keratitis. In all cases, outcomes were measured by quantitative microbiological techniques. RESULTS: The candidate peptides (CCI A, B, and C and COL-1) produced a total reduction of the test pathogens in phosphate buffered saline. In modified Optisol, the peptides were effective against S epidermidis at all temperatures, demonstrated augmented activity at 23 degrees C against the gram-positive organisms, but were ineffective against P aeruginosa. The addition of EDTA to the medium augmented the killing of P aeruginosa but made no difference in the reduction of gram-positive organisms. In an in vivo rabbit model of Pseudomonas keratitis, COL-1 demonstrated neither clinical nor microbicidal efficacy and appeared to have a very narrow dosage range, outside of which it appeared to be toxic to the ocular surface. CONCLUSION: Our data indicate that the antimicrobial peptides we tested were effective in vitro but not in vivo. In an age of increasing antibiotic resistance, antimicrobial peptides, developed over millions of years as innate

  4. Influence of temperature, oxygen and bacterial strain identity on the association of Campylobacter jejuni with Acanthamoeba castellanii.

    PubMed

    Baré, Julie; Sabbe, Koen; Huws, Sharon; Vercauteren, Dries; Braeckmans, Kevin; van Gremberghe, Ineke; Favoreel, Herman; Houf, Kurt

    2010-11-01

    Campylobacteriosis is the most frequently reported foodborne disease in the industrialized world, mainly through consumption of contaminated chicken meat. To date, no information is available on the primary infection sources of poultry. In this study, the ability of five Campylobacter jejuni strains with different invasion potential towards Caco-2 cells to survive and replicate in the protozoan Acanthamoeba castellanii was tested under simulated in situ conditions (i.e. chicken broiler houses). Results indicate that environmental conditions play a crucial role in C. jejuni-A. castellanii interactions. Co-culture in general did not result in an increase of either bacteria or amoebae. However, co-culture with Acanthamoeba did result in a delayed decline and an increased long-term survival of Campylobacter. Bacterial strain-specific effects were observed, with higher survival rates for low-invasive strains. The presence of C. jejuni in general did not affect A. castellanii viability, except at 37 °C under microaerobic conditions, where the presence of the reference and low-invasive Campylobacter strains resulted in a significant decline in amoebal viability. Confocal laser scanning microscopy revealed that intra-amoebal campylobacters were not always colocated with acidic organelles, suggesting potential bacterial interference with digestive processes. As Acanthamoeba enhances the persistence of C. jejuni, the presence of the amoeba in broiler house environments may have important implications for the ecology and epidemiology of this food pathogen. PMID:20722733

  5. Direct photoaffinity labeling by nucleotides of the apparent catalytic site on the heavy chains of smooth muscle and Acanthamoeba myosins

    SciTech Connect

    Maruta, H.; Korn, E.D.

    1981-01-10

    The heavy chains of Acanthamoeba myosins, IA, IB and II, turkey gizzard myosin, and rabbit skeletal muscle myosin subfragment-1 were specifically labeled by radioactive ATP, ADP, and UTP, each of which is a substrate or product of myosin ATPase activity, when irradiated with uv light at 0/sup 0/C. With UTP, as much as 0.45 mol/mol of Acanthamoeba myosin IA heavy chain and 1 mol/mol of turkey gizzard myosin heavy chain was incorporated. Evidence that the ligands were associated with the catalytic site included the observations that reaction occurred only with nucleotides that are substrates or products of the ATPase activity; that the reaction was blocked by pyrophosphate which is an inhibitor of the ATPase activity; that ATP was bound as ADP; and that label was probably restricted to a single peptide following limited subtilisin proteolysis of labeled Acanthamoeba myosin IA heavy chain and extensive cleavage with CNBr and trypsin of labeled turkey gizzard myosin heavy chain.

  6. A fatal case of meningoencephalitis due to a free-living amoeba of uncertain identity--probably acanthamoeba sp.

    PubMed

    Carter, R F; Cullity, G J; Ojeda, V J; Silberstein, P; Willaert, E

    1981-01-01

    There are 2 main types of meningoencephalitis caused by free-living amoebae. The first is a well-defined acutely fatal disease resembling fulminating bacterial meningitis. It is caused by the single species Naegleria fowleri. The second is a more poorly defined disease that runs a subacute or chronic course and is characterized by focal granulomatous lesions in the brain. The causative organisms are probably Acanthamoeba sp. in most cases, but it is possible that other genera may be involved. The first case of the subacute form of the disease to be recognized in Australia is described. A 2 1/2-yr-old, previously well girl presented with ataxia and lower motor neurone paralyses. The cerebrospinal fluid was pleocytic and she was thought to be suffering from a relatively minor viral brain-stem encephalitis. Her symptoms persisted in a peculiarly fluctuating way for 30 d when she suddenly collapsed and died from an intracranial haemorrhage. Necropsy showed focal granulomatous lesions associated with necrotizing vasculitis in the basal regions of the brain. The lesions contained well preserved free-living amoebae which were morphologically different from N. fowleri and most closely resembled Acanthamoeba sp. The ultrastructure of the organisms was particularly well preserved and is described in some detail. Immunohistological studies also excluded N. fowleri but were inconclusive for Acanthamoeba or other genera of free-living amoebae. Difficulties with the diagnosis and treatment of this disease are discussed and some practical suggestions are made. PMID:6261208

  7. Conditioned medium from stimulated mononuclear leukocytes augments human neutrophil-mediated killing of a virulent Acanthamoeba sp.

    PubMed Central

    Ferrante, A; Abell, T J

    1986-01-01

    Human neutrophils in the presence of serum containing anti-amoeba antibody either lacked amoebicidal activity or were poorly amoebicidal for Acanthamoeba culbertsoni. In contrast, neutrophils preexposed for 1 h to supernatants from human peripheral blood mononuclear leukocytes (MNLs) stimulated with phytohemagglutinin demonstrated significant amoeba killing in the presence of serum containing anti-acanthamoeba antibodies. Supernatant from MNL cultured in the absence of phytohemagglutinin were not effective in stimulating significant activity in the neutrophils. Serum containing antibody promoted the adherence of many neutrophils to one amoeba. There was no significant difference between the ability of neutrophils treated with supernatants from stimulated MNLs (stimulated conditioned medium [sCM]) and supernatants from nonstimulated MNLs (nonstimulated conditioned medium [nsCM]) in their binding to acanthamoeba. The effects of sCM on neutrophils was a general phenomenon. For example, the sCM but not the nsCM enhanced the antibody-dependent neutrophil-mediated cytotoxicity against three tumor targets (K562 erythroid myeloid leukemia cell line, B16 melanoma, and P815 (DBA/2 mastocytoma). Furthermore, the sCM but not the nsCM increased the bactericidal (against Staphylococcus aureus and Streptococcus pneumoniae) and fungicidal (against Torulopsis glabrata) activity of the neutrophil. The sCM but not the nsCM contained activities which inhibited neutrophil migration and stimulated a respiratory burst in these leukocytes. These results suggest that the neutrophil antimicrobial power can be increased by exposing the leukocytes to MNL mediators. Images PMID:3943902

  8. [Acanthamoeba, naturally intracellularly infected with Pseudomonas aeruginosa, after their isolation from a microbiologically contaminated drinking water system in a hospital].

    PubMed

    Michel, R; Burghardt, H; Bergmann, H

    1995-03-01

    The drinking water system of a new hospital building that was highly contaminated with bacteria before opening was investigated too for the prevalence of small free living amoebae. Germ counts resulted in > 100 CFU/ml in 100% of the cold water samples, that showed also growth of P. aeruginosa, whereas E. coli and coliforme bacteria could not be identified. The investigation of 37 water samples for protozoa revealed growth of small freeliving amoebae in 20 samples (54%) belonging to 10 species of the genus Acanthamoeba, Naegleria, Hartmannella, Echinamoeba among others. In addition 2 Ciliate- and 2 Microflagellate-species could be observed. While all Naegleria strains isolated belonged to the N. gruberi-complex two of 16 Acanthamoeba-isolates proved to be pathogenic for laboratory mice. From 7 watersamples positive with P. aeruginosa 5 Acanthamoeba- and 2 Echinamoeba strains could be isolated which revealed intracellular multiplication of P. aeruginosa. Because of their well known resistances against chlorine, the amoebae and their cysts are considered to be vectors for these intracellular bacteria. A complete sanitation of the incriminated drinking water system was accomplished by combined chemical and thermic disinfection measures.

  9. Sequences, structural models, and cellular localization of the actin- related proteins Arp2 and Arp3 from Acanthamoeba

    PubMed Central

    1995-01-01

    We cloned and sequenced the two actin-related proteins (Arps) present in the profilin-binding complex of Acanthamoeba (Machesky, L. M., S. J. Atkinson, C. Ampe, J. Vandekerckhove, and T. D. Pollard. 1994, J. Cell Biol. 127:107-115). The sequence of Arp2 is more similar to other Arp2s than to actin, while the sequence of Arp3 is more similar to other Arp3s than to actin. Phylogenetic analysis of all known Arps demonstrates that most group into three major families, which are likely to be shared across all eukaryotic phyla. Together with conventional actins, the Arps form a larger family distinct from structurally related ATPases such as Hsp70's and sugar kinases. Atomic models of the Arps based on their sequences and the structure of actin provide some clues about function. Both Arps have atoms appropriately placed to bind ATP and divalent cation. Arp2, but not Arp3, has a conserved profilin-binding site. Neither Arp has the residues required to copolymerize with actin, but an Arp heterodimer present in the profilin-binding complex might serve as a pointed end nucleus for actin polymerization. Both Acanthamoeba Arps are soluble in cell homogenates, and both are concentrated in the cortex of Acanthamoeba. The cellular concentrations are 1.9 microM Arp2 and 5.1 microM Arp3, substoichiometric to actin (200 microM) but comparable to many actin- binding proteins. PMID:7593166

  10. Age and therapeutic outcome of experimental Pseudomonas aeruginosa keratitis treated with ciprofloxacin, prednisolone, and flurbiprofen.

    PubMed Central

    Hobden, J A; Hill, J M; Engel, L S; O'Callaghan, R J

    1993-01-01

    This study was conducted to determine whether the age of the host influences the pathogenesis and therapeutic outcome of drug-treated Pseudomonas aeruginosa keratitis. Young (3- to 5-month-old) and old (1.5- to 3-year-old) rabbits were intrastromally infected with P. aeruginosa ATCC 27853. Sixteen hours later, rabbits in both age subpopulations were divided into three groups and treated topically as follows: group 1, phosphate-buffered saline; group 2, 0.3% ciprofloxacin; and group 3, 0.3% ciprofloxacin, 1.0% prednisolone, and 0.03% flurbiprofen. Drops were given every 15 min for 1 h and then every 30 min for 9 h. At 27 h postinfection, ocular pathology was graded with a slit lamp examination (SLE) scoring system. Aqueous humor was collected for ciprofloxacin quantitation, and corneas were harvested for bacterial enumeration and estimation of polymorphonuclear leukocytes. Young rabbits had more severe inflammation and pathology than old rabbits. At 27 h postinfection, SLE scores and polymorphonuclear leukocyte numbers were significantly higher for young rabbits than old rabbits (P < 0.02), regardless of treatment. Prednisolone and flurbiprofen significantly reduced SLE scores in both age groups (P < 0.03) without affecting the antimicrobial efficacy of ciprofloxacin. PMID:8239596

  11. Mutation of the PAX6 gene in patients with autosomal dominant keratitis

    SciTech Connect

    Mirzayans, F.; Pearce, W.G.; MacDonald, I.M.; Walter, M.A.

    1995-09-01

    Autosomal dominant keratitis (ADK) is an eye disorder chiefly characterized by corneal opacification and vascularization and by foveal hypoplasia. Aniridia (shown recently to result from mutations in the PAX6 gene) has overlapping clinical findings and a similar pattern of inheritance with ADK. On the basis of these similarities, we used a candidate-gene approach to investigate whether mutations in the PAX6 gene also result in ADK. Significant linkage was found between two polymorphic loci in the PAX6 region and ADK in a family with 15 affected members in four generations (peak LOD score = 4.45; {theta} = .00 with D11S914), consistent with PAX6 mutations being responsible for ADK. SSCP analysis and direct sequencing revealed a mutation in the PAX6 exon 11 splice-acceptor site. The predicted consequent incorrect splicing results in truncation of the PAX6 proline-serine-threonine activation domain. The Sey{sup Neu} mouse results from a mutation in the Pax-6 exon 10 splice-donor site that produces a PAX6 protein truncated from the same point as occurs in our family with ADK. Therefore, the Sey{sup Neu} mouse is an excellent animal model of ADK. The finding that mutations in PAX6 also underlie Peters anomaly implicates PAX6 broadly in human anterior segment malformations. 42 refs., 5 figs., 3 tabs.

  12. Antiviral treatment and other therapeutic interventions for herpes simplex virus epithelial keratitis

    PubMed Central

    Wilhelmus, Kirk R

    2015-01-01

    Background Eye disease due to herpes simplex virus (HSV) commonly presents as epithelial keratitis which, though usually self-limiting, may persist or progress without treatment. Objectives To compare the relative effectiveness of antiviral agents, interferon, and corneal debridement in the treatment of HSV epithelial keratitis. Search methods We searched CENTRAL (which contains the Cochrane Eyes and Vision Group Trials Register) (2014, Issue 12), PubMed (January 1946 to 31 December 2014), EMBASE (January 1980 to 31 December 2014), Latin American and Caribbean Health Sciences Literature Database (LILACS) (January 1982 to 31 December 2014), System for Information on Grey Literature in Europe (OpenGrey) (January 1995 to 31 December 2014), BIOSIS (January 1926 to 5 May 2014), Scopus (January 1966 to 31 December 2014), Japan Science and Technology Institute (J-Global) (January 1975 to 31 December 2014), China National Knowledge Infrastructure (CNKI) (January 1979 to 31 December 2014), British Library’s Electronic Table of Contents (Zetoc) (January 1993 to 7 May 2014). We looked for trials listed on the the metaRegister of Controlled Trials (www.controlled-trials.com), ClinicalTrials.gov (www.clinicaltrials.gov), the World Health Organization (WHO) International Clinical Trials Registry Platform (ICTRP) (www.who.int/ictrp/search/en), Chinese Clinical Trial Registry, the U.S. Food and Drug Administration (FDA) (www.fda.gov/), National Institute for Health and Clinical Excellence (NICE) (www.evidence.nhs.uk) and the European Medicines Agency (EMA) (www.ema.europa.eu/ema/) as of 31 December 2014. There were no language or date restrictions in the search for trials. We also culled literature digests and conference proceedings as of 15 April 2014. There were no language or date restrictions in the search for trials. Selection criteria Randomised and quasi-randomised trials of HSV dendritic or geographic epithelial keratitis were included that reported the proportion of

  13. Mapping of dendritic lesions in patients with herpes simplex keratitis using in vivo confocal microscopy

    PubMed Central

    Yokogawa, Hideaki; Kobayashi, Akira; Mori, Natsuko; Sugiyama, Kazuhisa

    2015-01-01

    Purpose To produce a two-dimensional reconstruction map of dendritic lesions in patients with herpes simplex keratitis (HSK) using in vivo confocal microscopy. Methods Four eyes of four patients (mean 65.8 years) with HSK presenting with a dendritic lesion were enrolled. Slit-lamp biomicroscopy and in vivo laser confocal microscopy were performed. Acquired confocal images at the level of the epithelium were arranged and mapped into subconfluent montages. Changes in the shape and degree of light reflection of abnormal cells and deposits around dendritic lesions as well as other corneal layers were qualitatively evaluated. Results Mapping of dendritic lesion was successful in all cases, and the subconfluent montages clearly showed the larger image of dendritic lesion. In all cases, the dendritic lesion consisted of hyperreflective irregular epithelial cells, and was surrounded by distorted and elongated epithelial cells. In three cases, hyperreflective deposits were noted at the midline of the lesion. The corneal stroma showed a hyperreflective honeycomb pattern. In two cases, inflammatory cells were observed at the level of endothelial cell layer. Conclusion Mapping of dendritic lesions in patients with HSK was successful in all patients using in vivo confocal microscopy. Cellular level observation of dendritic lesion at a relatively larger magnification may help understand the in vivo morphological change of HSK. Further study in more patients with HSK and nonherpetic dendritic lesion is needed to utilize confocal microscopy images in differential diagnosis and follow-up of the epithelial lesions with dendrite. PMID:26445524

  14. 75 FR 3641 - Television Broadcasting Services; Anchorage, AK

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-01-22

    ... From the Federal Register Online via the Government Publishing Office FEDERAL COMMUNICATIONS COMMISSION 47 CFR Part 73 Television Broadcasting Services; Anchorage, AK AGENCY: Federal Communications... broadcasting. 0 For the reasons discussed in the preamble, the Federal Communications Commission amends 47...

  15. 78 FR 55772 - Alaska Disaster Number AK-00028

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-09-11

    ... From the Federal Register Online via the Government Publishing Office ] SMALL BUSINESS ADMINISTRATION Alaska Disaster Number AK-00028 AGENCY: U.S. Small Business Administration. ACTION: Amendment 1... information in the original declaration remains unchanged. (Catalog of Federal Domestic Assistance...

  16. Root Cause Analysis of the Fusarium Keratitis Epidemic of 2004–2006 and Prescriptions for Preventing Future Epidemics

    PubMed Central

    Bullock, John D.

    2009-01-01

    Purpose: A root cause analysis of the Fusarium keratitis epidemic of 2004–2006 was performed. Methods: Three US Food and Drug Administration (FDA) documents were analyzed. Poisson and case-control studies were performed on outbreak data from Singapore. Irreversible thermochromic labels were applied to cartons of contact lens solution bottles, which were then subjected to elevated temperatures. Results: The 1997 FDA guidance document concerning storage temperatures of contact lens care products predicted temperature-related solution instability. Bausch & Lomb (B&L) requested FDA approval for ReNu with MoistureLoc, claiming that it was substantially equivalent to other products. FDA Form 483 stated that cases of ReNu-related Fusarium keratitis from Asia had not been reported, the removal of the product from the Asian markets was unreported, and B&L had not performed biocidal testing on samples associated with Asian cases. The outbreak in Singapore could have been recognized after only 3 cases (Pr = .0067). The cause of the Singapore outbreak could have been determined after the recognition of only 3 (P = .0429), 5 (95% confidence interval [CI], 1.15–126.0), or 15 cases (95% CI, 1.60–14.1). Thermochromic labels can irreversibly change color when exposed to elevated temperatures, thus warning of potential antimicrobial failure. Conclusions: The worldwide Fusarium keratitis epidemic of 2004–2006 could, theoretically, have been prevented entirely, recognized much earlier, or mitigated by much more rigorous oversight by the FDA, by strict adherence by B&L to FDA guidelines and requirements, by the application of basic statistical methods, and/or by the use of temperature indication technology. The lessons learned from a root cause analysis of this pharmacologic catastrophy may help avert or mitigate future epidemics. PMID:20126495

  17. Atomic force microscopic imaging of Acanthamoeba castellanii and Balamuthia mandrillaris trophozoites and cysts.

    PubMed

    Aqeel, Yousuf; Siddiqui, Ruqaiyyah; Ateeq, Muhammad; Raza Shah, Muhammad; Kulsoom, Huma; Khan, Naveed Ahmed

    2015-01-01

    Light microscopy and electron microscopy have been successfully used in the study of microbes, as well as free-living protists. Unlike light microscopy, which enables us to observe living organisms or the electron microscope which provides a two-dimensional image, atomic force microscopy provides a three-dimensional surface profile. Here, we observed two free-living amoebae, Acanthamoeba castellanii and Balamuthia mandrillaris under the phase contrast inverted microscope, transmission electron microscope and atomic force microscope. Although light microscopy was of lower magnification, it revealed functional biology of live amoebae such as motility and osmoregulation using contractile vacuoles of the trophozoite stage, but it is of limited value in defining the cyst stage. In contrast, transmission electron microscopy showed significantly greater magnification and resolution to reveal the ultra-structural features of trophozoites and cysts including intracellular organelles and cyst wall characteristics but it only produced a snapshot in time of a dead amoeba cell. Atomic force microscopy produced three-dimensional images providing detailed topographic description of shape and surface, phase imaging measuring boundary stiffness, and amplitude measurements including width, height and length of A. castellanii and B. mandrillaris trophozoites and cysts. These results demonstrate the importance of the application of various microscopic methods in the biological and structural characterization of the whole cell, ultra-structural features, as well as surface components and cytoskeleton of protist pathogens.

  18. Co-incubation of Acanthamoeba castellanii with strains of Pseudomonas aeruginosa alters the survival of amoeba.

    PubMed

    Cengiz, A M; Harmis, N; Stapleton, F

    2000-06-01

    Enhanced survival of Acanthamoeba castellanii has previously been reported following co-incubation with a single strain of Pseudomonas aeruginosa. The aim of this study was to evaluate the impact of different strains of P. aeruginosa on amoebae survival. Four contact lens solutions were challenged with A. castellanii for between 6 and 24 h, and survival rates of amoeba were calculated. Subsequently, A. castellanii was co-incubated with different strains of P. aeruginosa (strain 6294, an invasive isolate; 6206, a cytotoxic isolate; and Paer 001, a null isolate). Differences in amoeba survival over time between solutions for each bacterial strain were analysed. Non-neutralized hydrogen peroxide was the most effective system against A. castellani at all time points (P<0.05). Survival rates were not different between multipurpose solutions and neutralized hydrogen peroxide. Co-incubation with P. aeruginosa altered amoeba survival, and maximum survival occurred in the presence of the invasive strain of P. aeruginosa. Enhanced amoeba survival may occur in the presence of certain strains of Gram-negative bacteria, and with certain types of contact lens disinfection systems.

  19. Acanthamoeba and other free-living amoebae in bat guano, an extreme habitat.

    PubMed

    Mulec, Janez; Dietersdorfer, Elisabeth; Üstüntürk-Onan, Miray; Walochnik, Julia

    2016-04-01

    Several representatives of the so-called free-living amoebae (FLA) are of medical relevance, not only as facultative pathogens but also as vehicles for pathogenic bacteria. Some FLA can survive and even grow under extreme environmental conditions. Bat guano is an exceptional habitat, the conditions becoming gradually more extreme with aging. In the current study, samples of bat guano of different ages from five caves in Slovenia were screened for the presence of FLA. FLA were isolated from almost all guano samples, including guano with a pH of 3.5. Only the two samples that had been drawn from >20-year-old guano were negative for FLA. Generally, FLA diversity correlated to high concentrations of cultivable bacteria (∼10(8) CFU/g) and fungi (∼10(5) CFU/g). Interestingly, the absence of FLA in seasoned guanos was mirrored by the presence of dictyostelid slime moulds. The isolated amoebae were identified as belonging to the genera Acanthamoeba, Copromyxa, Naegleria, Sappinia, Tetramitus, Thecamoeba, Vahlkampfia, Vannella and Vermamoeba. To the best of our knowledge, this is the first study on the diversity of FLA in guano.

  20. Transcription and processing of mitochondrial RNA in the human pathogen Acanthamoeba castellanii.

    PubMed

    Accari, Jessica; Barth, Christian

    2015-07-01

    The size, structure, gene content and organisation of mitochondrial genomes can be highly diverse especially amongst the protists. We investigated the transcription and processing of the mitochondrial genome of the opportunistic pathogen Acanthamoeba castellanii and here we present a detailed transcription map of the 41.6 kb genome that encodes 33 proteins, 16 tRNAs and 2 rRNAs. Northern hybridisation studies identified six major polycistronic transcripts, most of which are co-transcriptionally processed into smaller mono-, di- and tricistronic RNAs. The maturation of the polycistronic transcripts is likely to involve endonucleolytic cleavage where tRNAs serve as processing signals. Reverse transcription polymerase chain reactions across the intervening regions between the six major polycistronic transcripts suggest that these transcripts were once part of an even larger transcript. Our findings indicate that the mitochondrial genome of A. castellanii is transcribed from only one or two promoters, very similar to the mode of transcription in the mitochondria of its close relative Dictyostelium discoideum, where transcription is known to occur from only a single transcription initiation site. Transcription initiation from a minimal number of promoters despite a large genome size may be an emerging trend in the mitochondria of protists.