Report of rare genotypes of Acanthamoeba from soil source of the Payeh Maga Highland forest, North-eastern Sarawak, Malaysia.

Pathogenic Acanthamoeba species are the causative agents of Acanthamoeba keratitis and granulomatous amoebic encephalitis. Members of this amoeba genus are ubiquitous in the environments. In Malaysia, most environmental studies performed to date have targeted the detection and characterisation of Acanthamoeba sp. in different water sources, dust and soil samples collected near human habitats. However, no local study has yet to examine these amoebae in a forest, an isolated terrestrial environment, where human activity is relatively scarce. Further, there are also limited studies to investigate the same globally. The current study reported the isolation, morphological and genotypic characterisations of eleven Acanthamoeba sp. isolated from soils of the Payeh Maga Highland forest, Sarawak, Malaysia. Morphological analysis revealed that nine isolates belonged to Group II, whereas two isolates belonged to Group III as defined by the criteria of Pussard and Pons. The phylogenetic analysis based on complete 18S rRNA gene sequences showed that the isolates belonged to the rare T1 (six isolates), T6 (two isolates) and T13 (three isolates) genotypes. To the best of our knowledge, this is the first report about the detection of T6 Acanthamoeba sp. in this country. Overall, the current findings have enriched the knowledge pertaining to the occurrence, morphological and genotypic characteristics of Acanthamoeba sp. in an isolated terrestrial environment of Malaysia.

Imagenología diferencial por microscopia confocal in vivo entre la epiteliopatía por acanthamoeba y la queratitis por herpes simple / In vivo confocal microscopy imaging for the differential diagnosis of Acanthamoeba epitheliopathy and herpes simplex keratitis

La queratitis por Acanthamoeba es una infección corneal de baja incidencia, condicionada por varios factores, pero con manifestación clínica variada y sintomatología típica. En su fase epitelial debe ser diferenciada de otras queratitis, específicamente de la queratitis por herpes simple, por la similitud entre ambas en cuanto a las características de la lesión corneal. La microscopia confocal in vivo constituye una alternativa diagnóstica. Es una biopsia fotográfica en cuyas imágenes podemos describir los quistes y trofozoítos de Acanthamoeba desde etapas iniciales, que nos ayudan a diferenciarla de otros tipos de queratitis e iniciar el tratamiento precoz. Se realizó una búsqueda de artículos publicados, con el objetivo de mostrar las imágenes por microscopia confocal de la fase epitelial de la infección corneal por Acanthamoeba y herpes simple. Se utilizó la plataforma Infomed, específicamente la Biblioteca Virtual de Salud(AU)

Acanthamoeba keratitis is a low-incidence corneal infection caused by several factors and characterized by a variety of clinical manifestations and typical symptoms. In its epithelial phase, it should be differentiated from other keratitis, particularly from herpes simplex keratitis, due to the similar characteristics of the corneal lesion. In vivo confocal microscopy is a diagnostic alternative consisting in a photographic biopsy showing images of Acanthamoeba cysts and trophozoites since their initial stages, thus allowing differentiation from other types of keratitis and the initiation of early treatment. A search was conducted of published papers with the purpose of showing confocal microscopy images of the epithelial phase of Acanthamoeba and herpes simplex corneal infection. Use was made of the platform Infomed, specifically the Virtual Health Library(AU)

Delayed diagnoses of Acanthamoeba keratitis at a tertiary care medical centre.

PURPOSE: To determine the prevalence and reasons for delays in diagnosis in patients with Acanthamoeba keratitis (AK) presenting to Wilmer Eye Institute, Baltimore, Maryland. METHODS: This retrospective study analysed all patients with culture-positive AK seen between 2012 and 2019 at a tertiary referral centre. Patient demographic information, clinical history, risk factors, symptom duration, referral patterns, slit lamp examination findings, visual acuity and need for surgery were collected. RESULTS: The study included 45 eyes of 43 patients. On average, patients were symptomatic for 52.6 days before culture collection. Thirty-one percent of patients were diagnosed within 28 days of symptom onset while 69% were diagnosed after 28 days. Before presentation to a tertiary care centre, 69% of patients were evaluated by an ophthalmologist outside of this institution and 27% were evaluated by a provider other than an ophthalmologist. AK was most commonly misdiagnosed as herpetic keratitis, occurring in 38% of patients. The strongest risk factor for AK was contact lens use. Only 11% of patients presented with the classic ring infiltrate and 82% had pain. Patients with an early versus late diagnosis had a mean Snellen visual acuity (VA) of 20/224 versus 20/296 at presentation (p = 0.33) and a mean Snellen VA of 20/91 versus 20/240 at final visit (p = 0.07). 11% of patients required a therapeutic penetrating keratoplasty. CONCLUSION: Delayed diagnosis of AK in our cohort occurred due to a misdiagnosis as herpetic keratitis, non-specific clinical signs including the lack of pain in a number of patients, and a delay in referral to a tertiary care centre. Any contact lens wearer with an atypical keratitis should be referred promptly for Acanthamoeba cultures.

Infectious keratitis: an update on epidemiology, causative microorganisms, risk factors, and antimicrobial resistance.

Corneal opacity is the 5th leading cause of blindness and visual impairment globally, affecting ~6 million of the world population. In addition, it is responsible for 1.5-2.0 million new cases of monocular blindness per year, highlighting an ongoing uncurbed burden on human health. Among all aetiologies such as infection, trauma, inflammation, degeneration and nutritional deficiency, infectious keratitis (IK) represents the leading cause of corneal blindness in both developed and developing countries, with an estimated incidence ranging from 2.5 to 799 per 100,000 population-year. IK can be caused by a wide range of microorganisms, including bacteria, fungi, virus, parasites and polymicrobial infection. Subject to the geographical and temporal variations, bacteria and fungi have been shown to be the most common causative microorganisms for corneal infection. Although viral and Acanthamoeba keratitis are less common, they represent important causes for corneal blindness in the developed countries. Contact lens wear, trauma, ocular surface diseases, lid diseases, and post-ocular surgery have been shown to be the major risk factors for IK. Broad-spectrum topical antimicrobial treatment is the current mainstay of treatment for IK, though its effectiveness is being challenged by the emergence of antimicrobial resistance, including multidrug resistance, in some parts of the world. In this review, we aim to provide an updated review on IK, encompassing the epidemiology, causative microorganisms, major risk factors and the impact of antimicrobial resistance.

Acanthamoeba keratitis: a review of biology, pathophysiology and epidemiology.

Acanthamoeba keratitis is an infection caused by a unicellular protozoan of the genus Acanthamoeba that is universally widespread. Until now, most cases were reported in contact lens wearers, although it is also a reality for non-wearers, mostly connected to corneal trauma. There is also a variation in incidence regarding the aetiology of the disease between developed and developing countries. PURPOSE: This work is based on a literature review, and the main goal is to deepen the knowledge about Acanthamoeba keratitis, presenting the main risk factors and focusing on prevention actions for this type of corneal infection since the treatments are not always effective. It targets specialists in visual health to strengthen their knowledge in this area, as well as to allow them to better inform their patients about hygiene care, appropriate measures of disinfection and ways to minimise the risk of infection. At this stage, it is important to highlight the essential role that practitioners play in fitting, monitoring and following-up patients to minimise the danger of infection. RECENT FINDINGS: It is well recognised that corneal trauma facilitates invasion by leaving an open door for microorganisms to penetrate the cornea. In addition to trauma, risk factors are mostly associated with patients' behaviours, such as interaction of contact lenses with contaminated water in the shower, swimming pools and beaches, etc., lack of hygiene habits with contact lenses and respective cases, and the use of ineffective disinfecting solutions. The fact that a disinfecting solution is not completely effective against trophozoites and/or cysts, both forms of Acanthamoeba's lifecycle, can cause the infection since one cyst alone leads to the emergence of a whole new population of Acanthamoeba. SUMMARY: It is necessary to reduce the risk of infection and, beyond the need to promote patient education to encourage correct CL hygiene behaviours, it should also be highlighted that there is an urgent need to enhance the efficacy of CL disinfection systems against all strains and both stages of Acanthamoeba through the creation of standardised methods. The ease of purchasing CLs without any supervision must also be considered a concern, and, in the near future, it is also important to develop and implement effective diagnostic methods and treatments for Acanthamoeba keratitis.

Infectious keratitis in southern Brazil: a comparison culture negative and culture positive patients / Ceratite infecciosa no sul do Brasil: comparação entre pacientes com cultura negativa e positiva

Abstract Purpose: To compare clinical-epidemiological profile and treatment outcome between culture negative and culture positive keratitis patients. Methods: Patients with suspected infectious keratitis seen at two ophthalmic hospitals in Curitiba, Brazil, between June 2014 and April 2016, were prospectively studied. Ophthalmological exam with corneal scraping and microbiological tests were performed. Data regarding follow up, surgical interventions and treatment outcome were collected after 12 weeks of the first visit trough medical chart review. From the results of the culture, two groups were formed: culture negative keratitis (CNK) and culture positive keratitis (CPK). Results: According to inclusion criteria 21 patients were classified as culture negative keratitis and 20 patients as culture positive keratitis. The number of patients on antibiotic drops at the first visit was greater in CNK group (90.5% versus 60%; p=0.032). Surgical procedures were necessary in 3 patients (15%) in CNK group and in 7 patients (36,8%) in CPK group (p=0.155). Treatment success was achieved by 85% (17/20) of the patients in CNK group and by 61% (11/18) of the patients in CPK group (p=0.144). There was no significant difference between groups regarding age, gender, place of residence, presence of comorbidities, risk factors for infectious keratitis, duration of symptoms and characteristics of corneal ulcer. Conclusions: Previous treatment with antibiotics correlates with negative culture results. There was no significant difference in treatment outcome between culture negative and culture positive keratitis patients.

Resumo Objetivo: Comparar os perfis clinico-epidemiológicos e os desfechos entre pacientes com ceratite com cultura positiva e pacientes com ceratite com cultura negativa. Métodos: Pacientes com ceratite infecciosa, atendidos em dois hospitais oftalmológicos em Curitiba, Brasil, entre junho de 2014 e abril de 2016, foram estudados prospectivamente. Exame oftalmológico, raspado de córnea e exames microbiológicos foram realizados no primeiro atendimento. Os dados quanto a seguimento e desfecho foram coletados após 12 semanas do primeiro atendimento através de revisão de prontuário. A partir dos resultados das culturas, dois grupos foram formados: ceratite com cultura negativa e ceratite com cultura positiva. Resultados: Vinte e um pacientes foram classificados como ceratite com cultura negativa e 20 como ceratite com cultura positiva. O número de pacientes em uso de colírio antibiótico no primeiro atendimento foi maior no grupo de cultura negativa (90,5% versus 60%; p=0,032). Sete pacientes (37%) no grupo cultura positiva precisaram de procedimentos cirúrgicos no manejo da ceratite, versus 3 pacientes (15%) do grupo cultura negativa (p=0,155). Oitenta e cinco por cento (17/20) dos pacientes do grupo cultura negativa alcançaram sucesso no tratamento, contra 61% (11/18) dos pacientes no grupo cultura positiva (p=0,144). Não houve diferença entre os grupos quanto a idade, gênero, local de procedência, presença de comorbidades, fatores de risco, duração dos sintomas e características da úlcera de córnea. Conclusão: Tratamento prévio com colírio de antibiótico correlaciona-se com resultados negativos de cultura. Não houve diferença no desfecho após tratamento entre os pacientes com cultura negativa e cultura positiva.

Parasitosis ocular por Acanthamoeba / Acanthamoeba parasitic eye infection

RESUMEN El parasitismo es uno de los fenómenos más sorprendentes de los observados en los animales. El número de organismos conocidos de vida parasitaria es muy elevado. Existe una gran cantidad de especies cuya supervivencia está relacionada con la de otras y dependen, en distinta medida, de ellas. Esta dependencia no implica que los animales parásitos sean organismos degenerados o deficientes; al contrario, a la vida parasitaria se ha llegado tras largo tiempo de evolución, en el que los parásitos han ido superando barreras y adaptándose a vivir en, o sobre sus hospedadores. Las queratitis por Acanthamoeba son infrecuentes; pero se describe un aumento a nivel mundial relacionado con el uso creciente de lentes de contacto. El diagnóstico precoz y el tratamiento adecuado deben realizarse para evitar la pérdida de la visión. Se realizó una búsqueda de artículos publicados, con el objetivo de conocer sobre la Acanthamoeba como parásito y su afectación ocular. Se utilizó la plataforma Infomed, específicamente la Biblioteca Virtual de Salud(AU)

ABSTRACT Parasitism is one of the most surprising phenomena among those occurring in animals. The number of known parasitic organisms is very high. There is a large number of species whose survival is related to that of others and depend on them to a greater or lesser degree. Such dependence does not imply that parasitic animals are either degenerate or deficient. On the contrary, parasitic life has been the result of a long process of evolution along which parasites have gradually overcome hurdles and have adapted to live in or on their hosts. Acanthamoeba keratitis is a rare disease, but a worldwide increase has been reported due to the growing use of contact lenses. Early diagnosis and appropriate treatment are required to prevent sight loss. A search was conducted for published papers with the purpose of learning about Acanthamoeba as a parasite and the related eye conditions. Use was made of the platform Infomed, specifically the Virtual Health Library(AU)

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.

Virulent T4 Acanthamoeba causing keratitis in a patient after swimming while wearing contact lenses in Southern Brazil.

Several strains of free-living amoebae belonging to the genus Acanthamoeba can cause a painful sight-threatening disease of the cornea known as Acanthamoeba keratitis (AK). The numbers of AK cases keep rising worldwide mainly due to an increase in contact lens wearers and lack of hygiene in the maintenance of contact lenses and their cases. We report a case of AK in a healthy young woman admitted to the Hospital de Clinicas in Porto Alegre, southern Brazil. Corneal scrapings were examined for the presence of Acanthamoeba strains. The initial isolate was characterized by morphological and genotypic properties. The isolate belonged to group III according to Pussard and Pons' cyst morphology. Analysis of its 18S rDNA sequence identified the isolate as genotype T4. The T4 genotype is the most commonly reported among keratitis isolates and the most common in environmental samples.

Presence of Acanthamoeba in the ocular surface in a Spanish population of contact lens wearers.

Pathogenic strains of Acanthamoeba are causative agents of a sight-threatening infection of the cornea known as Acanthamoeba keratitis (AK) which mainly affects contact lens wearers and it is commonly related to poor hygiene of contact lenses and their cases. Moreover, treatment of AK is complex due to the existence of a highly resistant cyst stage and if not diagnosed early has poor prognosis, leading to blindness and/or keratoplasty. Even though AK is increasing worldwide as well as awareness among patients and clinicians, it is still a poorly studied pathogen. Additionally, a remaining question to be answered is whether these opportunistic pathogens are present in the ocular surface of healthy contact lens wearers since they are the main group at risk.In order to carry out this study, sterile Schirmer strip tests were collected from a group of individuals all of them contact lens wearers who were attending a local ophthalmology clinic in Tenerife, Canary Islands, Spain. The collected samples (100 eyes of 50 patients) were cultured in 2% Non-Nutrient Agar (NNA) plates and positive plates (16) were then cultured in axenic conditions for further analyses. Molecular analysis classified all isolated strains belonged to Acanthamoeba genotype T4 and osmotolerance and thermotolerance assays revealed that all strains were potentially pathogenic. In conclusion, the ocular surface of contact lens wearers included in this study was colonized by potentially pathogenic strains of Acanthamoeba and should be considered as a risk for AK infection in this region and worldwide.

Detection and molecular characterization of Acanthamoeba spp. in stray cats from Madrid, Spain.

Acanthamoeba spp. is a widespread protozoan that has been isolated from air, dust, soil, water and biological samples. An opportunistic pathogen of humans and animals, it may cause ocular keratitis, encephalitis, and even multisystem disease. The frequency of Acanthamoeba in animals is unknown. The aim of present study was determine the presence of Acanthamoeba spp. in immunocompromised stray cats - animals possibly more likely to harbour the infection given their immunocompromised status and frequenting of contaminated environments. Of 307 cats examined, 55 were positive for feline immunodeficiency virus and/or feline leukaemia virus and therefore included in the study. Corneal scrapings were obtained to isolate Acanthamoeba spp. by culture and molecular detection by conventional and real time PCR. None of the samples examined directly by molecular methods were positive for Acanthamoeba spp. However, two (3.6%) cases of the cultured samples provided positive results, which were confirmed by subsequent molecular analysis. Sequencing assigned one isolate to genotype T4 and the other to T2. Since Acanthamoeba spp. may also infect animals and humans, the present findings may raise some public health and veterinary concerns.

The rising tide of Acanthamoeba keratitis in Auckland, New Zealand: a 7-year review of presentation, diagnosis and outcomes (2009-2016).

IMPORTANCE: Acanthamoeba is an increasingly prevalent cause of vision-threatening microbial keratitis. BACKGROUND: To assess the incidence, clinical presentation, diagnosis and outcomes of patients with Acanthamoeba keratitis (AK) in Auckland, New Zealand over a 7-year period. DESIGN: Retrospective observational consecutive case series. PARTICIPANTS: Fifty-eight eyes of 52 patients diagnosed with AK. METHODS: All cases of AK were identified using a cross-referenced search of clinical, laboratory and pharmacy records from March 2009 to May 2016. MAIN OUTCOME MEASURES: Demographic and clinical data were collected including age, gender, risk factors, clinical manifestations, initial diagnosis, diagnostic investigations, treatment, presenting and final visual acuity and surgical interventions. RESULTS: Contact lens (CL) use was noted in 96% of unilateral and 100% of bilateral cases. The mean duration of symptoms at presentation was 21 days and the mean duration from presentation to definitive diagnosis was 14 days. Initial diagnosis was recorded as CL-related keratitis in 70.6%, viral keratitis in 15.5% and AK in 12.0%. The diagnosis was confirmed with In vivo confocal microscopy (IVCM) in 67.2%, corneal scrape in 22.4%, corneal biopsy in 1.7% and clinically in 8.6%. IVCM sensitivity was 83.0%. Surgical intervention was required in four patients, all with delayed diagnosis (range 63-125 days). The incidence of AK has more than doubled when compared with the preceding 7-year period. CONCLUSIONS AND RELEVANCE: AK is a rare vision-threatening protozoal infection with rapidly-increasing incidence in New Zealand, predominantly affecting CL users. Diagnosis is often challenging and when delayed is associated with worse outcomes. IVCM offers rapid diagnosis with high sensitivity.

Acanthamoeba keratitis in 194 patients: risk factors for bad outcomes and severe inflammatory complications.

BACKGROUND/AIMS: To determine demographic and clinical features of patients with Acanthamoeba keratitis (AK) that are independent risk factors both for bad outcomes and for severe inflammatory complications (SIC). METHODS: A retrospective audit of medical records of AK cases at Moorfields Eye Hospital from July 2000 to April 2012, including 12 earlier surgical cases. Cases with a bad outcome were defined as those having one or more of the following: corneal perforation, keratoplasty, other surgery (except biopsy), duration of antiamoebic therapy (AAT) ≥10.5 months (the 75th percentile of the whole cohort) and final visual acuity ≤20/80. SICs were defined as having scleritis and/or a stromal ring infiltrate. Multivariable analysis was used to identify independent risk factors for both bad outcomes and SICs. RESULTS: Records of 194 eyes (194 patients) were included, having bad outcomes in 93 (48%). Bad outcomes were associated with the presence of SIC, aged >34 years, corticosteroids used before giving AAT and symptom duration >37 days before AAT. The development of SIC was independently associated with aged >34 years, corticosteroids used before giving AAT and herpes simplex virus (HSV) keratitis treatment before AAT. CONCLUSIONS: The prompt diagnosis of AK, avoidance of a misdiagnosis of HSV keratitis and corticosteroid use before the exclusion of AK as a potential cause of keratitis are essential to the provision of a good outcome for patients and for the avoidance of SIC. Older age is an unmodifiable risk factor that may reflect differences in the immune response to AK in this patient subset.

Occurrence of Acanthamoeba genotypes in Central West Malaysian environments.

Acanthamoeba species are ubiquitous free-living protozoa that can be found worldwide. Occasionally, it can become parasitic and the causative agent of acanthamoebic keratitis (AK) and Granulomatous Amoebic Encephalitis (GAE) in man. A total of 160 environmental samples and 225 naturally-infected animal corneal swabs were collected for Acanthamoeba cultivation. Acanthamoeba was found to be high in samples collected from environments (85%, 136/160) compared to infected animal corneas (24.89%, 56/225) by microscopic examination. Analysis of nucleotide sequence of 18S rRNA gene of all the 192 cultivable Acanthamoeba isolates revealed 4 genotypes (T3, T4. T5 and T15) with T4 as the most prevalent (69.27%, 133/192) followed by T5 (20.31%), T15 (9.90%) and T3 (0.52%). Genotype T4 was from the strain of A. castellanii U07401 (44.27%), A. castellanii U07409 (20.83%) and A. polyphagaAY026243 (4.17%), but interestingly, only A. castellanii U07401 was detected in naturally infected corneal samples. In environmental samples, T4 was commonly detected in all samples including dry soil, dust, wet debris, wet soil and water. Among the T4, A. castellanii (U07409) strains were detected high occurrence in dry (45%) followed by aquatic (32.50%) and moist (22.50%) samples but however A. castellanii (U07401) strains were dominant in dry samples of soil and dust (93.10%). Subsequently, genotype T5 of A. lenticulata (U94741) strains were dominant in samples collected from aquatic environments (58.97%). In summary, A. castellanii (U07401) strains were found dominant in both environmental and corneal swab samples. Therefore, these strains are possibly the most virulent and dry soil or dusts are the most possible source of Acanthamoeba infection in cats and dogs corneas.

Non-contact lens related Acanthamoeba keratitis.

The purpose of the study is to describe epidemiology, clinical features, diagnosis, and treatment of Acanthamoeba keratitis (AK) with special focus on the disease in nonusers of contact lenses (CLs). This study was a perspective based on authors' experience and review of published literature. AK accounts for 2% of microbiology-proven cases of keratitis. Trauma and exposure to contaminated water are the main predisposing factors for the disease. Association with CLs is seen only in small fraction of cases. Contrary to classical description experience in India suggests that out of proportion pain, ring infiltrate, and radial keratoneuritis are seen in less than a third of cases. Majority of cases present with diffuse infiltrate, mimicking herpes simplex or fungal keratitis. The diagnosis can be confirmed by microscopic examination of corneal scraping material and culture on nonnutrient agar with an overlay of Escherichia coli. Confocal microscopy can help diagnosis in patients with deep infiltrate; however, experience with technique and interpretation of images influences its true value. Primary treatment of the infection is biguanides with or without diamidines. Most patients respond to medical treatment. Corticosteroids play an important role in the management and can be used when indicated after due consideration to established protocols. Surgery is rarely needed in patients where definitive management is initiated within 3 weeks of onset of symptoms. Lamellar keratoplasty has been shown to have good outcome in cases needing surgery. Since the clinical features of AK in nonusers of CL are different, it will be important for ophthalmologists to be aware of the scenario wherein to suspect this infection. Medical treatment is successful if the disease is diagnosed early and management is initiated soon.

Molecular and morphological characterization of Acanthamoeba isolated from corneal scrapes and contact lens wearers in Argentina.

In this study, we describe the frequency of Acanthamoeba keratitis (AK) in patients that assisted in the Ophthalmology Department and determine the species/genotypes of free living amoebas (FLA) isolates. FLA from Corneal scrapes (CS) and contact lens (CL) wearers were studied by morphological and molecular characterization. A database was constructed with sociodemographic, clinical findings and history of use of CL variables. During January 2000 and September 2016 patients with corneal pathology admitted to the Ophthalmology Service of the University Hospital in Córdoba city, Argentina were included in the study. FLA were detected in 1.5% (11/739) and in 17% (11/65) of CS and CL analyzed respectively. FLA isolates from CL users evidenced an 80.9% of inappropriate lens maintenance, 4.8% (1/21) were not CL users that have been in contact with waters in outdoor environment and 14,3% (3/21) with no data about CL users. Acanthamoeba was confirmed in 100% and 82% of CS and LC respectively. The most frequent symptom associated with AK was red eye and photophobia. FLA from CS belonged to group II but 82% (9/11) and 18% (2/11) from CL belonged to group II and III respectively. T4 genotype and A. polyphaga species were detected in 100% of Acanthamoeba isolates. Poor CL hygiene practices, highlights the need for improved education about the severity of AK and consequences of improper CL hygiene. Genotype T4 detected in 100% of both CS and CL samples, consistently with previous findings indicating that this genotype is by far the most prevalent isolated from ocular infection.

Acanthamoeba Keratitis among Rigid Gas Permeable Contact Lens Wearers in the United States, 2005 through 2011.

PURPOSE: To describe the clinical presentation and outcomes of Acanthamoeba keratitis (AK) in rigid gas permeable (RGP) contact lens wearers and to identify modifiable risk factors. DESIGN: Case-control investigation. PARTICIPANTS: Patients were RGP contact lens-wearing United States residents with a diagnosis of AK from 2005 through 2011. Controls were RGP contact lens wearers with no history of AK who were at least 12 years of age. METHODS: Patients were identified during 2 multistate AK outbreak investigations. Controls from the first investigation in 2007 were identified using a reverse address directory. In the second investigation, controls were recruited from participating ophthalmology and optometry practices. Patients and controls were interviewed by phone using a standardized questionnaire. Odds ratios (ORs) and Fisher exact P values were calculated to assess risk factors associated with infection. MAIN OUTCOME MEASURES: Acanthamoeba keratitis, a rare eye disease primarily affecting contact lens wearers, is caused by free-living amebae, Acanthamoeba species. RESULTS: We identified 37 patients in the 2 investigations, 10 (27%) from the 2007 investigation and 27 (73%) from 2011. There were 17 healthy controls, 9 (53%) from 2007 and 8 (47%) from 2011. Among patients, 9 (24%) wore RGP lenses for orthokeratology or therapeutic indication; no controls wore RGP lenses for these indications. Significant risk factors for AK were wearing lenses for orthokeratology (OR, undefined; P = 0.02), sleeping while wearing lenses (OR, 8.00; P = 0.04), storing lenses in tap water (OR, 16.00; P = 0.001), and topping off contact lens solution in the case (OR, 4.80; P = 0.01). After stratifying by use of RGP lenses for orthokeratology, storing lenses in tap water and topping off remained significant exposures. CONCLUSIONS: Nearly one quarter of patients were orthokeratology wearers. Using tap water to store RGP lenses and topping off solution in the lens case were modifiable risk behaviors identified in RGP wearers who wore lenses for both orthokeratology and nonorthokeratology indications. Rigid gas permeable wearers should avoid exposing their lenses to tap water and should empty their cases and use fresh lens solution each time they take out their lenses.

Genotypic, physiological, and biochemical characterization of potentially pathogenic Acanthamoeba isolated from the environment in Cairo, Egypt.

Acanthamoebae are the most common opportunistic amphizoic protozoa that cause life-threatening granulomatous amoebic encephalitis in immunocompromised individuals and sight-threatening amoebic keratitis (AK) in contact lens wearers. The present work aimed to determine the presence of Acanthamoeba isolates in different environmental sources: water, soil, and dust in Cairo, Egypt and to characterize the pathogenic potential of the isolated Acanthamoeba using physiological and biochemical assays as well as determination of the genotypes in an attempt to correlate pathogenicity with certain genotypes. The study included the collection of 22 corneal scrapings from patients complaining of symptoms and signs indicative of acanthamoeba keratitis (AK) and 75 environmental samples followed by cultivation on non-nutrient agar plates preseeded with E. coli. Positive samples for Acanthamoeba were subjected to osmo- and thermo-tolerance assays and zymography analysis. Potentially pathogenic isolates were subjected to PCR amplification using genus-specific primer pair. Isolates were classified at the genotype level based on the sequence analysis of Acanthamoeba 18S rRNA gene (diagnostic fragment 3). The total detection rate for Acanthamoeba in environmental samples was 33.3 %, 31.4 % in water, 40 % in soil, and 20 % in dust samples. Three and two Acanthamoeba isolates from water and soil sources, respectively, had the potential for pathogenicity as they exhibited full range of pathogenic traits. Other 12 isolates were designated as weak potential pathogens. Only ten of the environmental isolates were positive in PCR and were classified by genotype analysis into T4 genotype (70 %), T3 (10 %) and T5 (20 %). Potential pathogens belonged to genotypes T4 (from water) and T5 (from soil) while weak potential pathogens belonged to genotypes T3 (from water) and T4 (from water and soil). Additionally, T7 genotype was isolated from keratitis patients. There is a considerable variation in the response of Acanthamoeba members of the same genotype to pathogenicity indicator assays making correlation of pathogenicity with certain genotypes difficult. Presence of potentially pathogenic Acanthamoeba isolates in habitats related directly to human populations represent a risk for human health. Isolation of Acanthamoeba genotype T7 from AK cases, which is commonly considered as nonpathogenic, might draw the attention to other Acanthamoeba genotypes considered as non pathogenic and reevaluate their role in production of human infections. To our knowledge, this is the first study on the presence and distribution of Acanthamoeba genotypes in the environment, Cairo, Egypt.

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

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).

Sensibilidad antibiótica de bacterias obtenidas de queratitis e infecciones intraoculares en la Fundación Oftalmológica de Santander (FOSCAL), Floridablanca, Colombia / Antibiotic susceptibility patterns of bacteria isolated from keratitis and intraocular infections at Fundación Oftalmológica de Santander (FOSCAL), Floridablanca, Colombia

Introducción. La resistencia bacteriana es crítica para la selección de los antibióticos en el tratamiento de las infecciones, por ello es vital conocer su estado actual en nuestro medio. Objetivo. Determinar la sensibilidad antibiótica bacteriana in vitro obtenida de los cultivos de queratitis e infecciones intraoculares. Materiales y métodos. Se llevó a cabo un estudio retrospectivo en la Fundación Oftalmológica de Santander (FOSCAL), entre junio de 2011 y enero de 2012. Resultados. Se examinaron 92 muestras. Se identificaron 110 bacterias, 27 hongos y 12 amebas de vida libre. Del total de bacterias Gram positivas, 1,1 %, 0 %, 1,1 %, 16,9 %, 29,3 % y 85 % fue resistente a imipenem, moxifloxacina, gatifloxacina, levofloxacina, ciprofloxacina y tobramicina, respectivamente, mientras que la resistencia a estos mismos fármacos se presentó, respectivamente, en 0 %, 8,3 %, 0 %, 0 %, 18,2 % y 27,3 % de las bacterias Gram negativas. Los porcentajes de resistencia de los estafilococos positivos para coagulasa resistentes a la meticilina fueron 0 %, 0 %, 0 %, 7 %, 17 % y 100 %, respectivamente, y los porcentajes de los estafilococos negativos para coagulasa resistentes a la meticilina fueron 3 %, 0 %, 0 %, 24 %, 44 % y 100 %, respectivamente. Los porcentajes de resistencia bacteriana globales (tanto para bacterias Gram positivas como para Gram negativas) a imipenem, moxifloxacina, gatifloxacina, levofloxacina, ciprofloxacina y tobramicina fueron 1 %, 1 %, 1 %, 15,1 %, 28 % y 64,5 %, respectivamente. Conclusiones. Los niveles de resistencia bacteriana para imipenem, moxifloxacina y gatifloxacina fueron menores que para levofloxacina, ciprofloxacina y tobramicina. Los niveles de resistencia para la tobramicina fueron muy altos, lo que pone en duda su utilidad clínica en las infecciones oculares en nuestro medio.

Introduction: Bacterial resistance is critical for the selection of antibiotics in the treatment of infections, so it is vital to know its current status in our geographical area. Objective: To determine in vitro antibiotic susceptibility of bacterial isolates obtained from keratitis and intraocular infections. Materials and methods: A retrospective study of microbiological tests in Fundación Oftalmológica de Santander (FOSCAL) was carried out between June, 2011, and January, 2012. Results: A total of 92 samples were examined and 110 bacteria, 27 fungi and 12 free-living amoebae were identified. Polymicrobial infections constituted 50% of the total; 1.1%, 0%, 1.1%, 16.9%, 29.3% and 85% of Gram-positive bacteria were resistant to imipenem, moxifloxacin, gatifloxacin, levofloxacin, ciprofloxacin and tobramycin, respectively, while 0%, 8.3%, 0%, 0%, 18.2% and 27.3% of Gram-negative bacteria were resistant to imipenem, moxifloxacin, gatifloxacin, levofloxacin, ciprofloxacin and tobramycin, respectively. For methicillin-resistant coagulase-positive staphylococci, resistance percentages to imipenem, moxifloxacin, gatifloxacin, levofloxacin, ciprofloxacin and tobramycin were 0%, 0%, 0%, 7%, 17% and 100%, respectively. For methicillin-resistant coagulase-negative staphylococci, resistance percentages to imipenem, moxifloxacin, gatifloxacin, levofloxacin, ciprofloxacin and tobramycin were 3%, 0%, 0%, 24%, 44% and 100%, respectively. Overall bacterial resistance to imipenem, moxifloxacin, gatifloxacin, levofloxacin, ciprofloxacin and tobramycin, for both Gram-positive and Gram-negative, was 1%, 1%, 1%, 15.1%, 28% and 64.5%, respectively. Conclusions: The levels of bacterial resistance to imipenem, moxifloxacin and gatifloxacin were lower than for levofloxacin, ciprofloxacin and tobramycin. The levels of resistance to tobramycin were very high, which calls into question its usefulness in this region of our country.