Reliable and specific detection of Acanthamoeba spp. in dishcloths using quantitative real-time PCR assay.

Acanthamoeba spp., are ubiquitous protist which belongs to Free-Living Amoeba (FLA) group, is considered as causal agent of side-threatening keratitis or fatal encephalitis among other human infections. Besides, this parasite has been reported as host for other microorganisms important to human health such as Campylobacter spp. or Vibrio spp. among others. This role of Acanthamoeba as pathogen and environmental phagocyte has increased the reports confirming its presence in human related environments, acting as a water quality indicator. Considering the tide relationship between water and kitchen environments, and the high prevalence of Acanthamoeba in water sources, the present study aims to establish a quick and accurate protocol based on DNA extraction and a real time qPCR assay to detect Acanthamoeba spp. in dishcloths. The procedure has been validated by processing 17 used dishcloths. Our findings demonstrated the high sensitivity of the qPCR assay used which was capable of detecting up to one Acanthamoeba from an in vitro contaminated dishcloth. The protocol accurately detected 64.7% of positive samples for Acanthamoeba spp, (in 4 samples DNA concentrations corresponded to 1-102 amoebae). Our findings demonstrate the importance of FLA surveillance by efficient and sensitive methods since one amoeba is capable of colonizing human related food environments such as kitchens sinks and could be a potential source of infection.

In vitro amoebicidal effects of arabinogalactan-based ophthalmic solution.

The main corneal infections reported worldwide are caused by bacteria and viruses but, recently, the number of Acanthamoeba keratitis (AK) cases has increased. Acanthamoeba genus is an opportunistic free living protozoa widely distributed in environmental and clinical sources, with two life-cycle stages: the trophozoite and the cyst. AK presents as primary symptoms eye redness, epithelial defects, photophobia and intense pain. An early diagnosis and an effective treatment are crucial to avoid blindness or eye removal but, so far, there is no established treatment to this corneal infection. Diverse research studies have reported the efficacy of commercialized eye drops and ophthalmic solutions against the two life cycle stages of Acanthamoeba strains, that usually present preservatives such as Propylene Glycol of Benzalkonium chloride (BAK). These compounds present toxic effects in corneal cells, favouring the inflammatory response in the so sensitive eye tissue. In the present work we have evaluated the efficacy of nine proprietary ophthalmic solutions with and without preservatives (ASDA Dry Eyes Eyedrops, Miren®, ODM5®, Ectodol®, Systane® Complete, Ocudox®, Matrix Ocular®, Alins® and Coqun®) against the two life cycle stages of three Acanthamoeba strains. Our work has demonstrated the high anti-Acanthamoeba activity of Matrix Ocular®, which induces the programmed cell death mechanisms in Acanthamoeba spp. trophozoites. The high efficacy and the absence of ocular toxic effects of Matrix Ocular®, evidences the use of the Arabinogalactan derivatives as a new source of anti-AK compounds.

Evaluation of unenhanced post-mortem computed tomography to detect chest injuries in violent death.

PURPOSE: The aim of this study was to assess the performances of unenhanced post-mortem computed tomography (CT) to detect thoracic injuries in violent death. MATERIALS AND METHODS: Retrospectively, we conducted a review of unenhanced CT scans of 67 victims of violent deaths with thoracic injuries and compared CT findings with the results of clinical autopsy. Our gold standard was a comparison of CT scans with autopsy discussed in a monthly forensic radiology multidisciplinary team meeting (MDTM). The data were collected by organ system: heart, pericardium, aorta, lungs, pleura, bone, and diaphragm and performance indices (sensitivity, specificity, accuracy) were calculated. RESULTS: Pleural (59/67) and bone (55/67) injuries detected on CT were also found at autopsy and confirmed by the MDTM (sensitivity and specificity 100%). Seventeen out of 67 diaphragmatic lesions were visible on CT. Eighteen out of 67 were confirmed during MDTM after autopsy, yielding overall sensitivity of 94% and specificity of 98%. Forty out of 67 lung contusions were found on CT with two false positives and one false negative yielding 95% sensitivity for CT with a specificity of 96%, and accuracy of 95%. Fourteen out of 67 aortic injuries were found on CT compared to 19 confirmed during MDTM (sensitivity 74%, specificity 85%, accuracy 82%). In terms of pericardial lesions, 19/67 were found on CT and 20 on autopsy and confirmed during MDTM (sensitivity 80%, specificity 94%, accuracy 85%). Ten out of 10/67 cardiac lesions were visible on CT imaging and 15 found on autopsy and confirmed during MDTM (sensitivity 57%, specificity 94%, accuracy 81%). CONCLUSION: Unenhanced post-mortem CT performs well to detect pleural, pulmonary, bone and diaphragmatic injuries but less well to identify cardiac and aortic injuries, for which the use of indirect signs is essential.

Isolation of Naegleria fowleri from a domestic water tank associated with a fatal encephalitis in a 4 month-old Venezuelan child.

This study describes the association of household water system contamination with the pathogenic Free-Living Amoeba (FLA) Naegleria fowleri and a case of fatal Primary Amoebic Meningoencephalitis (PAM) in a child from the state of Monagas in Venezuela. Amoebae were initially identified by microscopy from a sample of cerebrospinal fluid (CSF) from the child. Direct DNA extraction and specific PCR/sequencing for N. fowleri was also carried out from the same CSF sample. In order to determine a possible environmental source of infection, water samples from the water tank of the child's home and also water bodies recently visited by the child and his family, were examined for the presence of N. fowleri by culture and PCR/sequencing. The results obtained from the collected water samples revealed that only the water tank of the house was positive for N. fowleri. PCR/sequencing showed that the strains isolated from the patient and the water tanks were 100 % identical. Therefore, the house water tank was confirmed as the source of infection in this case, possibly as a result of the occasional immersion of the child´s head under the water while bathing. This case highlights a novel source of thermally polluted water and another threat of N. fowleri infection.