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Necropsy examination of an adult Atlantic salmon (Salmo salar) from the Dalälven River in Sweden revealed numerous large, white nodules, with spherical cysts and granulomata in kidney and liver. Histopathology showed dark, septate, thin-walled hyphae. The aetiologic agent was found to be an Ochroconis species (Venturiales) that differed from known fish-associated species of the genus. Molecular phylogenetic studies of the culture (strain UIII09 = CBS 135766) demonstrated that Ochroconis globalis was concerned. The isolate proved to be susceptible to all investigated antifungals, as it is known for another Ochroconis species. The role of Ochroconis in opportunism of cold-blooded animals was discussed, and the diagnostic methods using DNA sequences for routine identification of the fungus were proposed., (© 2019 John Wiley & Sons Ltd.)

Multidrug resistance in V. cholerae has been increasing around the world including northeastern Thailand. The aquatic environment is a reservoir of V. cholerae and might be an important source of resistant strains. The aims of this study were to investigate the phylogenetic relationships of int SXT gene sequences from 31 clinical and 14 environmental V. cholerae O1 and non-O1/non-O139 isolates and 11 sequences amplified directly from environmental water samples. We also amplified class 1 integrons, the SXT elements (targeting the int SXT gene) and antimicrobial resistance genes directly from water samples. Phylogenetic analysis displayed two major distinct clusters (clusters 1 and 2). Most V. cholerae O1 (19/20, 95%) and non-O1/non-O139 isolates (8/11, 72.7%) from clinical sources, and all sequences obtained directly from water samples, belonged to cluster 1. Cluster 2 mostly comprised environmental non-O1/non-O139 isolates (10/12, 83.3%). We successfully amplified the SXT elements directly from17.5% of water samples. Associated resistance genes were also amplified as follows: sul2 (41.3% of water samples), dfrA1 (60%), dfr18 (33.8%), strB (70%) and tetA (2.5%). Class 1 integrons were not found in water samples, indicating that the SXT element was the major contributor of multidrug resistance determinants in this region. The SXT element and antimicrobial resistance genes could be transferred from clinical V. cholerae O1 to environmental V. cholerae non-O1/non-O139 was demonstrated by conjugation experiment. These findings indicate that there may have been cross dissemination and horizontal gene transfer (HGT) of the SXT element harbored by V. cholerae O1 and non-O1/non-O139 strains isolated from clinical and environmental water sources. Environmental water might be an important source of antimicrobial resistance genes in V. cholerae in this region. Direct detection of antimicrobial resistance genes in water samples can be used for monitoring the spread of such genes in the ecosystem., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Recently, the taxonomy of Ochroconis (Ascomycota, Pezizomycotina, Venturiales, Sympoventuriaceae) has been revised with the recognition of an additional genus, Verruconis. Ochroconis comprises mesophilic saprobes that occasionally infect vertebrates which mostly are cold-blooded, while Verruconis contains thermophilic species which is a neurotrope in humans and birds. On the basis of molecular data it is noted that only a single Ochroconis species regularly infects immunocompetent human hosts. Here we report a subcutaneous infection due to Ochroconis mirabilis in a 50-year-old immunocompetent female patient. In vitro antifungal susceptibility tests revealed that terbinafine was the most effective drug. The patient was successfully cured with oral administration of terbinafine 250 mg daily in combination with 3 times of topical ALA-photodynamic therapy for 9 months.

The genera Ochroconis and Verruconis (Sympoventuriaceae, Venturiales) have remarkably high molecular diversity despite relatively high degrees of phenotypic similarity. Tree topologies, inter-specific and intra-specific heterogeneities, barcoding gaps and reciprocal monophyly of all currently known species were analyzed. It was concluded that all currently used genes viz. SSU, ITS, LSU, ACT1, BT2, and TEF1 were unable to reach all 'gold standard' criteria of barcoding markers. They could nevertheless be used for reasonably reliable identification of species, because the markers, although variable, were associated with large inter-specific heterogeneity. Of the coding protein-genes, ACT1 revealed highest potentiality as barcoding marker in mostly all parts of the investigated sequence. SSU, LSU, ITS, and ACT1 yielded consistent monophyly in all investigated species, but only SSU and LSU generated clear barcoding gaps. For phylogeny, LSU was an informative marker, suitable to reconstruct gene-trees showing correct phylogenetic relationships. Cryptic species were revealed especially in complexes with very high intra-specific variability. When all these complexes will be taxonomically resolved, ACT1 will probably appear to be the most reliable barcoding gene for Ochroconis and Verruconis., (Copyright © 2015 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.)

Lung specimens (n = 216) from six wildlife species were examined for occurrence of Pneumocystis species in pulmonary tissues. Among small mammals the shrew Sorex antinorii (80 %) were most frequently colonized. In contrast, foxes and badgers did not yield positive amplification. Host-specificity was noted, at least at the level of the host genus. Phylogenetic trees based on partial mtLSU and mtSSU showed high diversity of species corresponding to animal host diversity. Nuclear rDNA ITS data confirmed unambiguous separation of species. In conclusion, ITS is an excellent marker to distinguish species of the genus Pneumocystis., (Copyright © 2015 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.)

Purpose: To describe typical clinical and laboratory characteristics of severe fungal keratitis caused by Pythium insidiosum during the rainy season in Northeast Thailand and to report the efficacy of P. insidiosum vaccine in the treatment of Pythium keratitis., Methods: A series of hospital-based consecutive cases of Pythium keratitis were diagnosed and treated at Srinagarind Hospital (Khon Kaen University, Khon Kaen, Thailand). The clinical presentations, diagnostic tests, and management are described., Results: Severe fungal keratitis caused by P. insidiosum was diagnosed in 5 eyes of 4 patients between May 2009 and July 2009. All cases had a history of fungal keratitis after being exposed to contaminated water. Upon slit-lamp examination, subepithelial and superficial stromal opacities were observed in a reticular pattern in all cases. Pythium insidiosum was identified and confirmed by both microbiological culture and polymerase chain reaction. Clinical worsening was detected after conventional treatment with antifungal agents. Therapeutic penetrating keratoplasty with either donor cornea or scleral graft was performed together with topical antifungal administration and P. insidiosum vaccination. Subsequent evisceration was performed in 1 eye., Conclusions: An outbreak of Pythium keratitis in Northeast Thailand was reported. Distinctive clinical features are a suggestive clue for early diagnosis. Combination treatment including topical antifungal agents, radical surgery, and P. insidiosum vaccine may be considered for the management of Pythium keratitis.

We report the first case and clinical course of a case of human subcutaneous phaeohyphomycosis caused by Ochroconis tshawytschae, a rare fish pathogen. The diagnosis was based upon histopathological and mycological examinations of clinical samples. Identification of the etiologic agent was assessed on its phenotypic characteristics and subsequently, confirmed by molecular data. In vitro antifungal susceptibility of the isolate was investigated and a comparison was prepared of all of its features to those of its two most relevant related species, O. gallopava and O. humicola.

Two strains resembling the neurotropic fungus Ochroconis gallopava were isolated from hot spring river water (IFM 54738 and IFM 54739). The isolates showed optimal growth at 42 degrees C, while the maximum growth temperature was 49 degrees C, thus having temperature relationships similar to those of O. gallopava. Colonies were light olive green, with a color change to dark reddish brown after several passages, which was also observed in O. gallopava. Conidia were indistinguishable from those of O. gallopava. The antifungal susceptibility profile of the isolates was also similar to that of O. gallopava, except for a lower susceptibility to micafungin. The two isolates had 100% homologous rRNA genes including the internal transcribed spacer (ITS) region and the D1/D2 region of the large subunit. The gene fragments, as O. gallopava, could be amplified with species-specific rDNA primers, and loop-mediated isothermal amplification designed for O. gallopava yielded positive results in the two isolates. However, homologies with O. gallopava in ITS and D1/D2 regions were 79.2 and 95.9%, respectively, widely exceeding generally accepted species boundaries. These differences were corroborated in virulence tested in experimental infection. The two isolates did not kill a mouse even until 28 days. However, mortalities of four O. gallopava strains ranged from 40 to 100%. The new isolates mainly affected the kidneys; whereas O. gallopava had a strong preference for the brain. We therefore propose a new species, Ochroconis calidifluminalis, for the two isolates.

A rapid, inexpensive, simple, and accurate multiplex polymerase chain reaction (PCR) was developed in a single tube for identification of Mycobacterium tuberculosis. Assessment of sensitivity and specificity of simple PCR was performed with 116 strains of M. tuberculosis complex (MTC) and 144 strains of nontuberculous mycobacteria (NTM) compared with the biochemical method. Specific amplification of KS4, MTC-specific DNA fragment, was found in 98% (114/116) of MTC and not detected in 99% (143/144) of NTM. Amplification of the mtp40 gene revealed 95% sensitivity (100/105 strains of M. tuberculosis) and 77% specificity (not found in 119/155 mycobacterial strains). A multiplex PCR method based on the combination of KS4- and mtp40-derived primers was used for identification of M. tuberculosis. Crude DNA from slow growing mycobacteria with cream rough colonies that showed both 768-bp amplified product for KS4 and 396-bp for mtp40 was identified as M. tuberculosis whereas that from MTC gave only the 768-bp product.