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The ability to distinguish between viable and non-viable protozoan parasites is central to improved human and animal health management. While conceptually simple, methods to differentiate cell viability in situ remain challenging. Amoebic gill disease, caused by Neoparamoeba perurans is a parasitic disease impacting Atlantic salmon aquaculture globally. Although commercial freshwater treatments alleviate AGD, viable amoebae remain on gills or in used treatment water. Existing PCR-based assays are able to quantify N. perurans abundance but cannot discriminate amoeba viability. We investigated the use of propidium monoazide (PMA) application, prior to real-time PCR, to distinguish between alive and dead cells. We demonstrate that 200 μM PMA can significantly reduce amplification from non-viable (isopropanol treated) cultured amoebae across at least three logs of cell concentrations. Using a serial dilution of viable and non-viable cells, we show that non-PMA PCR amplifies both viable and non-viable amoebae, while PMA exposure suppresses (but does not completely inhibit) amplification from non-viable amoebae. The effect of freshwater treatment on N. perurans viability was assessed using the PMA-PCR. Following PMA exposure, amplification from freshwater treated amoebae was reduced by approximately 94–97 %. Taken together this study demonstrates that PMA combined with traditional real-time PCR can estimate amoeba viability. [ABSTRACT FROM AUTHOR]

Abstract Neoparamoeba perurans is the aetiological agent of amoebic gill disease (AGD) in salmonids, however multiple other amoeba species colonise the gills and their role in AGD is unknown. Taxonomic assessments of these accompanying amoebae on AGD-affected salmon have previously been based on gross morphology alone. The aim of the present study was to document the diversity of amoebae colonising the gills of AGD-affected farmed Atlantic salmon using a combination of morphological and sequence-based taxonomic methods. Amoebae were characterised morphologically via light microscopy and transmission electron microscopy, and by phylogenetic analyses based on the 18S rRNA gene and cytochrome oxidase subunit I (COI) gene. In addition to N. perurans , 11 other amoebozoans were isolated from the gills, and were classified within the genera Neoparamoeba , Paramoeba , Vexillifera , Pseudoparamoeba , Vannella and Nolandella. In some cases, such as Paramoeba eilhardi , this is the first time this species has been isolated from the gills of teleost fish. Furthermore, sequencing of both the 18S rRNA and COI gene revealed significant genetic variation within genera. We highlight that there is a far greater diversity of amoebae colonising AGD-affected gills than previously established. [ABSTRACT FROM AUTHOR]

The study was conducted to evaluate patient dose receiving from mammographic examinations using the Dose Management System at Vimut hospital, Bangkok, Thailand. The main purpose of this study was to compare the dose from conventional 2-dimension mammography and combined 2-dimension conventional with Tomosynthesis techniques with the published Diagnostic Reference Levels (DRLs) for mammography. Retrospective data were collected during June 2021 and June 2022 from Hologic 3Dimensions mammographic unit. The collected patient dose metrics include Average Glandular Dose (AGD) in mGy and Entrance Surface Air Kerma (ESAK) in mGy/mAs. Only acceptable quality images justified by the radiologists were included in this study. The patient dose data were collected using the Dose Management Software (Philips Dose Wise Portal). The AGD and ESAK from 2-dimensional mammographic 4 images in craniocaudal, mediolateral oblique and 4 combined conventional/tomosynthesis images in craniocaudal projection were recorded from a Hologic 3Dimensions mammographic unit. The median dose values (P50) were used to compare with the published standard dose and the Diagnostic Reference Levels (DRLs). The age of the patients in this study ranged from 35-60 years old. The kilovoltage peak (kVp) was found in the range of 27-31 kVp, with the milli-ampare-second between 60-160mAs. The breast compression thickness values of 2-dimensional mammographic from 4 images and 4 combined conventional/tomosynthesis images were between 2.9-6.9 cm and 7.0-7.9 cm, respectively. The median dose values (P50) from conventional technique and the combined technique, the AGD were found at 1.5 mGy and 1.48 mGy, respectively. The median values (P50) of the ESAK from conventional technique and the combined technique were found at 6.16 mGy and 5.92 mGy, respectively. Comparing our dose values with the International Atomic Energy Agency standard AGD of 3 mGy and ESAK of 11 mGy, we found that our dose data were below those international DRLs published values. [ABSTRACT FROM AUTHOR]

Chile is one of the world's most prolific producers of farmed salmon, but has suffered great economic losses due to fish diseases. In this study, the interactions of treatments for the caligidosis agent Caligus rogercresseyi and major diseases in salmon aquaculture caused by Piscirickettsia salmonis , Infectious Salmon Anaemia Virus and Neoparamoeba perurans were determined. This information is essential to develop an integrated management strategy in Chile. Immunosuppression due to fish stress, increased by frequent chemical baths to treat C. rogercresseyi, was considered to be the main source of chronic caligidosis and high prevalence of other diseases. P. salmonis and pathogenic ISAV were detected using PCR analysis at all farms sampled, but N. perurans at one farm in April only. In 2011, individuals of Oncorhynchus mykiss with gill pathology not caused by N. perurans and receiving chemical baths at Farm 1 were affected by 100% prevalence of sea lice. Similar prevalence was observed on Salmo salar at Farm 2, which did not receive bath treatments. Both farms were under similar oceanographic influence. A prevalence of 80% of a pathogenic strain of ISAV, but without clinical symptoms, was detected at Farm 2, combined with low sea lice presence and almost complete absence of ovigerous females (OF) even though no treatment had been applied. S. salar sampled at Farm 3 in 2013 were also chronically infested with sea lice despite routine chemical treatment baths. P. salmonis was correlated with the increase in OF and chalimii. It was concluded that chemical baths to treat sea lice triggered fish mortality concomitantly with AGD due to prior gill damage. Fish stress conditions elicited by fish-gill disease, fish maturation and P. salmonis were related to an increase of ovigerous sea lice females. The decrease in recurrent and routine chemical baths allowed a reduction of sea lice reproduction and, most likely, of ISAV outbreaks. [ABSTRACT FROM AUTHOR]

The article evaluates the in vitro toxicity of bithionol and bithionol sulphoxide to Neoparamoeba spp., the causative agent of amoebic gill disease in farmed fishes. Materials and methods used in isolating and collecting samples of Neoparamoeba spp. are described. The experiment showed that the number of amoebae declined rapidly when exposed to freshwater when compared to the seawater controls with bithionol and bithionol sulphoxide. The experiment also demonstrated the effectiveness of bithionol in reducing aoeba numbers.

• High throughput method to screen treatments effective against amoebic gill disease. • Hydrogen peroxide is amoebicidal 24 h post-treatment on return to seawater. • Freshwater treatment survivors exhibit reduced metabolic activity up to 24 h later. • Dual assays differentiate a novel treatments mechanism of action. The marine protozoan parasite Neoparamoeba perurans has been established as the causative agent for amoebic gill disease (AGD) in Atlantic salmon (Salmo salar). Freshwater bathing is the only routinely used treatment for AGD in Australia while hydrogen peroxide (H 2 O 2) is used in countries with cooler water temperatures. The identification of new treatments that do not rely on either freshwater or H 2 O 2 bathing is highly sought. However, in vitro based methods for high throughput screening of antiparasitic compounds have not been established for this parasite. To this end the present study evaluated two in vitro bioassays based on metabolic energy production and cellular membrane integrity to distinguish between amoebistatic (crenated or pseudocyst forms with recovery possible) and amoebicidal (death) activity. Amoebae were subject to either freshwater, H 2 O 2 or chloramine-T for 4 h treatment and assessed 24 h after recovery. Visualization by microscopy and bioassay assessment 24 h post-treatment confirmed that H 2 O 2 and freshwater are 95% amoebicidal albeit due to different mechanisms of action. These data are consistent with other studies where amoebae have been observed to recover following exposure to these compounds and provide evidence for the inclusion of a recovery component to differentiate between the mechanism of action of amoebicidal and amoebistatic treatments. Together these bioassays are a critical tool for high throughput screening of novel and more effective treatments against AGD. [ABSTRACT FROM AUTHOR]