Acanthamoeba in Southeast Asia - Overview and Challenges.

Acanthamoeba, one of free-living amoebae (FLA), remains a high risk of direct contact with this protozoan parasite which is ubiquitous in nature and man-made environment. This pathogenic FLA can cause sight-threatening amoebic keratitis (AK) and fatal granulomatous amoebic encephalitis (GAE) though these cases may not commonly be reported in our clinical settings. Acanthamoeba has been detected from different environmental sources namely; soil, water, hot-spring, swimming pool, air-conditioner, or contact lens storage cases. The identification of Acanthamoeba is based on morphological appearance and molecular techniques using PCR and DNA sequencing for clinico-epidemiological purposes. Recent treatments have long been ineffective against Acanthamoeba cyst, novel anti-Acanthamoeba agents have therefore been extensively investigated. There are efforts to utilize synthetic chemicals, lead compounds from medicinal plant extracts, and animal products to combat Acanthamoeba infection. Applied nanotechnology, an advanced technology, has shown to enhance the anti-Acanthamoeba activity in the encapsulated nanoparticles leading to new therapeutic options. This review attempts to provide an overview of the available data and studies on the occurrence of pathogenic Acanthamoeba among the Association of Southeast Asian Nations (ASEAN) members with the aim of identifying some potential contributing factors such as distribution, demographic profile of the patients, possible source of the parasite, mode of transmission and treatment. Further, this review attempts to provide future direction for prevention and control of the Acanthamoeba infection.

Natural Honey-Induced Acanthamoeba keratitis.

Natural honey in spite of its usefulness is known to contain certain microorganisms. In the present study, we describe a case of Acanthamoeba keratitis after using topical honey administered by a traditional medicine therapist. A 32-year-old male came with red eye and blurred vision. The pain and other symptoms became more severe after the 1st week, with appearance of radial perineuritis at the cornea. A repeated interview revealed that 1 week before appearance of ocular symptoms, the patient had instilled a drop of natural honey in his left eye. Confocal microscopic cornea imaging demonstrated cyst and trophozoite of Acanthamoeba in the corneal stroma.

Effect of povidone iodine, chlorhexidine digluconate and toyocamycin on amphizoic amoebic strains, infectious agents of Acanthamoeba keratitis - a growing threat to human health worldwide.

INTRODUCTION: Free-living amoebae, ubiquitous in outer environments, in predisposing circumstances may exist as parasites, infectious agents of Acanthamoeba keratitis. In recent decades, the vision-threatening corneal infection is a growing human health threat worldwide, including Poland. The applied therapy is often ineffective due to diagnostic mistakes, various pathogenicity of Acanthamoeba strains and high resistance of cysts to drugs; many agents with possible anti-amoebic activity are still being tested. In the presented study, selected chemicals are investigated in terms of their in vitro effect on corneal and environmental Acanthamoeba strains. MATERIAL AND METHODS: Samples of a corneal isolate from a patient with severe Acanthamoeba keratitis,of assessed on the basis of genotype associations of 18S rRNA and the type strain, Acanthamoeba castellanii Neff cultivated in bacteria-free condition, were exposed to povidone iodine, chlorhexidine digluconate or toyocamycin. In vitro population dynamics of the strains were monitored and compared to those of control cultures. RESULTS: All chemicals showed anti-amoebic effects with different degrees of effectiveness. Significant differences were observed in the in vitro population dynamics, and the morpho-physiological status of A. castellanii Neff T4 and corneal strains determined as A. polyphaga T4 genotype, exposed to povidone iodine or toyocamycin, in comparison with chlorhexidine taken as reference. CONCLUSIONS: Time-dependent amoebstatic in vitro effects were demonstrated for all agents, in particular, the results of assays with povidone iodine are promising. No significant stimulation of encystation appeared; however, as cysticidal efficacy of chemicals is expected, complementary research is needed on different Acanthamoeba strains with modified agent concentrations and method application.

Development of nanoparticle-assisted PCR assay in the rapid detection of brain-eating amoebae.

Brain-eating amoebae (Acanthamoeba spp., Balamuthia mandrillaris, Naegleria fowleri) have gained increasing attention owing to their capacity to produce severe human and animal infections involving the brain. Early detection is a pre-requisite in successful prognosis. Here, we developed a nanoPCR assay for the rapid detection of brain-eating amoebae using various nanoparticles. Graphene oxide, copper and alumina nanoparticles used in this study were characterized using Raman spectroscopy measurements through excitation with a He-Ne laser, while powder X-ray diffraction patterns were taken on a PANanalytical, X'Pert HighScore diffractometer and the morphology of the materials was confirmed using high-resolution transmission electron microscopy (HRTEM). Using nanoparticle-assisted PCR, the results revealed that graphene oxide, copper oxide and alumina nanoparticles significantly enhanced PCR efficiency in the detection of pathogenic free-living amoebae using genus-specific probes. The optimal concentration of graphene oxide, copper oxide and alumina nanoparticles for Acanthamoeba spp. was determined at 0.4, 0.04 and 0.4 µg per mL respectively. For B. mandrillaris, the optimal concentration was determined at 0.4 µg per mL for graphene oxide, copper oxide and alumina nanoparticles, and for Naegleria, the optimal concentration was 0.04, 4.0 and 0.04 µg per mL respectively. Moreover, combinations of these nanoparticles proved to further enhance PCR efficiency. The addition of metal oxide nanoparticles leads to excellent surface effect, while thermal conductivity property of the nanoparticles enhances PCR productivity. These findings suggest that nanoPCR assay has tremendous potential in the clinical diagnosis of parasitic infections as well as for studying epidemiology and pathology and environmental monitoring of other microbes.

Twelve-year analysis of microbial keratitis trends at a UK tertiary hospital.

PurposeTo investigate the frequencies, trends, and in vitro drug susceptibilities of the causative pathogens in microbial keratitis in Manchester Royal Eye Hospital.Patients and methodsCorneal scrape results recorded by the microbiology service between 2004 and 2015 were extracted from an established database. A total of 4229 corneal scrape specimens were identified from an established database. First-line antibiotic treatment in our centre during the study period was ofloxacin and second line was cefuroxime and gentamicin.ResultsMean age was 45.9±21.0. A total of 1379 samples (32.6%) were culture positive. One hundred forty-eight (10.7%) specimens cultured multiple organisms. Of the 1539 organisms identified, 63.3% were Gram-positive bacteria, 27.3% Gram-negative bacteria, 7.1% fungi, and 2.3% Acanthamoebae. A decreasing trend in Gram-positive isolates was found together with a stable trend in Gram negatives and an increasing trend in Acanthamoeba and fungi. There appeared to be a significant increasing trend of Moraxella infection (P=0.001). In all, 83.1 and 90.8% of Gram-positive and -negative isolates tested were susceptible to ofloxacin, respectively. Cefuroxime covered 86.6% of Gram-positive and 61.4% of Gram-negative isolates, whereas gentamicin covered 88.8 and 96.5% of Gram-positive and -negative isolates, respectively.ConclusionWe found a change in the type of Gram-negative organisms isolated over time, with the Moraxella species on the rise. Reassuringly, no significant increase in resistance was observed in vitro for any of the commonly used antibiotics. Ofloxacin remains a good first-line antibiotic treatment but duo-therapy does have broader coverage and should be considered in non-responsive cases.

Granulomatous amebic encephalitis in a child with acute lymphoblastic leukemia successfully treated with multimodal antimicrobial therapy and hyperbaric oxygen.

Acanthamoeba is the causative agent of granulomatous amebic encephalitis, a rare and usually fatal disease. We report a child with acute lymphoblastic leukemia who developed brain abscesses caused by Acanthamoeba during induction therapy. Multimodal antimicrobial chemotherapy and hyperbaric oxygen therapy resulted in complete resolution of symptoms and of pathology as seen by magnetic resonance imaging.

Impact of prior therapy on the recovery and frequency of corneal pathogens.

OBJECTIVE: To document the impact of prior antibiotic therapy on the recovery of corneal pathogens. METHODS: Medical records and laboratory reports of 334 consecutive microbial keratitis patients examined from January to December 2000 were reviewed. Comparisons of pathogens, culture positive rate, recovery time, antibiotic sensitivity profile, delay in presentation, and final visual acuity were analyzed for patients treated before presentation and those who were not. The chi square test was used to determine statistical significance. RESULTS: Of the 334 patients, 56% were exposed to at least one course of topical antimicrobials before culture. Patients on therapy were only slightly more likely to be culture negative (P = 0.317) but significantly more likely to have a delay in pathogen recovery (P = 0.002). Patients given prior antibiotics took significantly longer to heal (P = 0.003). Gram-negative organisms (47.5%) were the most frequent pathogens isolated from all culture-positive patients, followed by gram positives (28.7%), fungi (15.8%), and parasites (2%). An increase and significant difference in the frequency of fungi (P = 0.000) and acanthamoeba was reserved for the pretreated group. Gram negative organisms were more often isolated from patients who had not been pretreated (P = 0.002). Pretreated patients were more like to have a pathogen resistant to 1 or more of the commonly prescribed ocular antibiotics (P = 0.02). CONCLUSIONS: There is a delay in starting microbiologic-guided antibiotic treatment in patients who have received empiric therapy. Nonbacterial corneal pathogens may be associated more frequently with patients on prior therapy.