We submitted a panel of 416 isolates of Candida albicans from separate sources to multilocus sequence typing (MLST). The data generated determined a population structure in which four major clades of closely related isolates were delineated, together with eight minor clades comprising five or more isolates. By Fisher's exact test, a statistically significant association was found between particular clades and the anatomical source, geographical source, ABC genotype, decade of isolation, and homozygosity versus heterozygosity at the mating type-like locus (MTL) of the isolates in the clade. However, these associations may have been influenced by confounding variables, since in a univariate analysis of variance, only the clade associations with ABC type and anatomical source emerged as statistically significant, providing the first indication of possible differences between C. albicans strain type clades and their propensity to infect or colonize different anatomical locations. There were no significant differences between clades with respect to distributions of isolates resistant to fluconazole, itraconazole, or flucytosine. However, the majority of flucytosine-resistant isolates belonged to clade 1, and these isolates, but not flucytosine-resistant isolates in other clades, bore a unique mutation in the FUR1 gene that probably accounts for their resistance. A significantly higher proportion of isolates resistant to fluconazole, itraconazole, and flucytosine were homozygous at the MTL, suggesting that antifungal pressure may trigger a common mechanism that leads both to resistance and to MTL homozygosity. The utility of MLST for determining clade assignments of clinical isolates will form the basis for strain selection for future research into C. albicans virulence.Differentiation of microbial isolates by sequencing a small sample of unrelated housekeeping genes has become established as a reliable and effective method for typing strains of many bacteria (9, 50). Such multilocus sequence typing (MLST) is highly reproducible, and data can be archived in Web-based databases accessible to all users. For the fungal pathogen Candida albicans, an MLST system based on seven DNA fragments was developed as an optimal consensus for typing strains within the species (6) following two earlier proposed systems (5, 48). Because C. albicans is a diploid organism, sequence data contain heterozygous as well as homozygous sites, adding an extra discriminatory feature to MLST for this species.Among other approaches to strain typing of C. albicans, DNA fingerprinting based on the moderately repetitive sequence Ca3 has been widely used. By this method, C. albicans populations were shown to comprise five major clades of closely related strain types (46), including clades enriched in isolates from Europe (36) and South Africa (2, 3). Resistance of C. albicans to flucytosine in vitro was found to be a property restricted almost entirely to isolates from a single C. albicans clade as determined by Ca3 fingerprinting (37), and the sole me...

Amoebic gill disease (AGD) in farmed Atlantic salmon is caused by the amoeba Paramoeba perurans. The recent establishment of in vitro culture techniques for P. perurans has provided a valuable tool for studying the parasite in detail. In this study, flow cytometry was used to generate clonal cultures from single-sorted amoeba, and these were used to successfully establish AGD in experimental Atlantic salmon. The clonal cultures displayed differences in virulence, based on gill scores. The P. perurans load on gills, determined by qPCR analysis, showed a positive relationship with gill score, and with clonal virulence, indicating that the ability of amoebae to proliferate and/or remain attached on gills may play a role in virulence. Gill scores based on gross signs and histopathological analysis were in agreement. No association between level of gill score and specific gill arch was observed. It was found that for fish with lower gill scores based on histopathological examination, gross examination and qPCR analysis of gills from the same fish were less successful in detecting lesions and amoebae, respectively.

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Monitoring of planktonic salmon louse (Lepeophtheirus salmonis salmonis) abundance and parameterization of key life‐history traits has been hindered by labour‐intensive and error‐prone quantification using traditional light microscopy. Fluorescence illumination has been proposed as a means of improving visualization, but prior to this study adequate investigation of the relevant fluorescence profiles and measurement conditions has not been undertaken. We investigated the fluorescence profiles of L. salmonis and non‐target copepod spp. with excitation and emission matrices (200–600 nm) and identified unique fluorescence signals. Fluorescence microscopy using excitation wavelengths of 470 ± 40 nm, and emission wavelengths of 525 ± 50 nm, showed that after 90 days of formalin storage salmon lice have a mean fluorescence intensity that is 2.4 times greater than non‐target copepods (copepodid and adult stages). A 7‐day heat treatment of 42°C in formalin increased the difference between salmon louse copepodids and non‐target copepods to a factor of 3.6, eliminating the need for prolonged storage. Differences in the fluorescence signal and endogenous fluorophores were investigated with respect to variation in sea lice species, age, stage and host fish origin. Under the conditions outlined in this paper, the fluorescence signal was found to be a reliable means of visualizing and differentiating salmon lice from non‐target zooplankters. Adaptation of the fluorescence signal would greatly expedite traditional methods of enumerating salmon louse larvae in plankton samples and could provide a means of automated detection.

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