Chi-Ching Tsang

Oral yeasts and coliforms in HIV-infected individuals in Hong Kong.

The objective was to determine the oral carriage patterns of yeasts and coliforms and their relationships, if any, with age, risk group, CDC classification, CD4+ count and medications in a predominantly Chinese, HIV‐infected cohort in Hong Kong. A prospective longitudinal study was carried out over a 12‐month period, of 32 predominantly Chinese male HIV‐infected cohort in a hospital setting in Hong Kong. Oral carriage rates were determined by the concentrated rinse culture method and correlated with other clinical parameters using regression analysis. A total of 73 oral rinse samples were collected and the weighted mean carriage rates of oral yeasts and coliforms were 54.8% and 28.8%, respectively. The most common yeast and the Enterobacteriaceae isolated were Candida albicans and Enterobacter cloacae, respectively. An increased carriage rate of yeasts was associated with zidovudine usage and Centres for Disease Control (CDC) stage IV of the HIV infection whereas the opposite was associated with the usage of antiparasitics and multivitamins. Although the oral carriage rate of coliforms was significantly lower in individuals taking antibacterials and multivitamins, it was not significantly influenced by age, CD4+ lymphocyte count and the intake of antivirals, antifungals or folates. These data imply that oral yeast carriage in HIV infection is related to the severity of the disease as opposed to oral coliform carriage which appears to be unusually transient in the study cohort.

HIV protease inhibitors differentially inhibit adhesion of Candida albicans to acrylic surfaces.

Highly active antiretroviral therapy (HAART), using HIV protease inhibitors, is commonly used in the management of HIV infection. HIV protease inhibitors also have a direct effect on a key virulence factor of Candida albicans, its secreted aspartyl proteinase (Sap). Although protease inhibitors can attenuate Candida adhesion to human epithelial cells, their effects on adhesion to acrylic substances, which is a common component of oral appliances, is unknown. This study investigated whether protease inhibitors affect C. albicans adhesion to acrylic substances. C. albicans suspensions were pretreated with different concentrations of saquinavir, ritonavir or indinavir for 1 h and allowed to adhere on acrylic strips, which had been  pretreated with pooled human saliva for 30 min, for another hour in the presence of each drug. The test groups showed a significantly lower degree of adhesion than the controls. Adhesion was reduced by 50% at drug concentrations of 100, 100 and 20 μmol l−1 for saquinavir, ritonavir and indinavir respectively. In conclusion, protease inhibitors attenuated C. albicans adhesion to an acrylic surface in vitro in a dose‐dependent manner, and different protease inhibitors exhibited different degrees of inhibition.

Genetic diversity of Aspergillus species isolated from onychomycosis and Aspergillus hongkongensis sp. nov., with implications to antifungal susceptibility testing.

Thirteen Aspergillus isolates recovered from nails of 13 patients (fingernails, n=2; toenails, n=11) with onychomycosis were characterized. Twelve strains were identified by multilocus sequencing as Aspergillus spp. (Aspergillus sydowii [n=4], Aspergillus welwitschiae [n=3], Aspergillus terreus [n=2], Aspergillus flavus [n=1], Aspergillus tubingensis [n=1], and Aspergillus unguis [n=1]). Isolates of A. terreus, A. flavus, and A. unguis were also identifiable by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The 13th isolate (HKU49(T)) possessed unique morphological characteristics different from other Aspergillus spp. Molecular characterization also unambiguously showed that HKU49(T) was distinct from other Aspergillus spp. We propose the novel species Aspergillus hongkongensis to describe this previously unknown fungus. Antifungal susceptibility testing showed most Aspergillus isolates had low MICs against itraconazole and voriconazole, but all Aspergillus isolates had high MICs against fluconazole. A diverse spectrum of Aspergillus species is associated with onychomycosis. Itraconazole and voriconazole are probably better drug options for Aspergillus onychomycosis.

Subcutaneous phaeohyphomycosis in a patient with IgG4-related sclerosing disease caused by a novel ascomycete, Hongkongmyces pedis gen. et sp. nov.: first report of human infection associated with the family Lindgomycetaceae

No members of the freshwater ascomycetes family Lindgomycetaceae have been associated with human infections. We isolated a mould (HKU35(T)) from the biopsy specimen of a patient with invasive foot infection and underlying immunoglobulin G4-related sclerosing disease. Histology showed florid, suppurative, granulomatous inflammation in the dermis, with central microabscess formation surrounded by epithelioid histiocytes, scattered giant cells, and a small number of lymphocytes. A Grocott stain revealed fungal elements in the center of the lesion. On Sabouraud glucose agar, HKU35(T) grew as gray and velvety colonies. Among the members of the family Lindgomycetaceae, HKU35(T) was the only strain that grew at 37°C. Microscopically, only sterile mycelia, but no fruiting bodies, were observed. HKU35(T) was susceptible to itrazonazole, voriconazole, and posaconazole, which was in line with the patients clinical response to itraconazole treatment. Internal transcribed spacer and partial 18S nuclear rDNA (nrDNA), 28S nrDNA, β-tubulin gene, and EF1α gene sequencing showed that HKU35(T) occupied a unique phylogenetic position, most closely related to but distinct from members of the genera Clohesyomyces and Lindgomyces. We propose a new genus and species, Hongkongmyces pedis gen. et sp. nov., to describe this fungus, which belongs to the family Lindgomycetaceae in the orderPleosporales of class Dothideomycetes. This case also represents the first report of human infection associated with the family Lindgomycetaceae.

Tsukamurella hongkongensis sp. nov. and Tsukamurella sinensis sp. nov., isolated from patients with keratitis, catheter-related bacteraemia and conjunctivitis

Three bacterial strains, HKU51T, HKU52T and HKU53, were isolated from a conjunctival swab, corneal scraping and blood culture of three patients in Hong Kong with conjunctivitis, keratitis and catheter-related bacteraemia, respectively. Cells were Gram-stain-positive, aerobic, catalase-positive, non-sporulating and non-motile bacilli. The three strains had unique biochemical profiles that were distinguishable from those of closely related species of the genus Tsukamurella. Fatty acids, mycolic acids, cell-wall sugars and peptidoglycan analyses showed that they were typical of members of Tsukamurella. 16S rRNA gene sequence analysis revealed 100 % sequence identity between HKU52T and HKU53, and the two strains shared 99.5 % sequence identity with Tsukamurella sunchonensis JCM 15929T and Tsukamurella pseudospumae JCM 13375T; HKU51T shared 99.6 % sequence identity with Tsukamurella pulmonis CCUG 35732T. The DNA G+C contents of strains HKU51T, HKU52T and HKU53 were 70.9 ± 2.2, 71.3 ± 2.1 and 71.2 ± 2.3 mol% (mean ± sd; n = 3), respectively. DNA-DNA hybridization confirmed that the novel strains were distinct from other known species of the genus Tsukamurella ( ≤ 50.1 ± 3.7 % DNA-DNA relatedness); two of the isolates, HKU52T and HKU53, represented the same species ( ≥ 94.6 ± 5.6 % DNA-DNA relatedness), while the third isolate, HKU51T, represented another species. The novel species Tsukamurella hongkongensis sp. nov. is proposed to accommodate strains HKU52T and HKU53, with HKU52T ( = JCM 30715T = DSM 100208T) as the type strain; whilst another novel species, Tsukamurella sinensis sp. nov., is proposed to accommodate the third isolate, HKU51T ( = JCM 30714T = DSM 100207T), which is designated the type strain.

Intra-Genomic Internal Transcribed Spacer Region Sequence Heterogeneity and Molecular Diagnosis in Clinical Microbiology

Internal transcribed spacer region (ITS) sequencing is the most extensively used technology for accurate molecular identification of fungal pathogens in clinical microbiology laboratories. Intra-genomic ITS sequence heterogeneity, which makes fungal identification based on direct sequencing of PCR products difficult, has rarely been reported in pathogenic fungi. During the process of performing ITS sequencing on 71 yeast strains isolated from various clinical specimens, direct sequencing of the PCR products showed ambiguous sequences in six of them. After cloning the PCR products into plasmids for sequencing, interpretable sequencing electropherograms could be obtained. For each of the six isolates, 10–49 clones were selected for sequencing and two to seven intra-genomic ITS copies were detected. The identities of these six isolates were confirmed to be Candida glabrata (n = 2), Pichia (Candida) norvegensis (n = 2), Candida tropicalis (n = 1) and Saccharomyces cerevisiae (n = 1). Multiple sequence alignment revealed that one to four intra-genomic ITS polymorphic sites were present in the six isolates, and all these polymorphic sites were located in the ITS1 and/or ITS2 regions. We report and describe the first evidence of intra-genomic ITS sequence heterogeneity in four different pathogenic yeasts, which occurred exclusively in the ITS1 and ITS2 spacer regions for the six isolates in this study.

Polyketides, Toxins and Pigments in Penicillium marneffei

Penicillium marneffei (synonym: Talaromyces marneffei) is the most important pathogenic thermally dimorphic fungus in China and Southeastern Asia. The HIV/AIDS pandemic, particularly in China and other Southeast Asian countries, has led to the emergence of P. marneffei infection as an important AIDS-defining condition. Recently, we published the genome sequence of P. marneffei. In the P. marneffei genome, 23 polyketide synthase genes and two polyketide synthase-non-ribosomal peptide synthase hybrid genes were identified. This number is much higher than those of Coccidioides immitis and Histoplasma capsulatum, important pathogenic thermally dimorphic fungi in the Western world. Phylogenetically, these polyketide synthase genes were distributed evenly with their counterparts found in Aspergillus species and other fungi, suggesting that polyketide synthases in P. marneffei did not diverge from lineage-specific gene duplication through a recent expansion. Gene knockdown experiments and ultra-high performance liquid chromatography-photodiode array detector/electrospray ionization-quadruple time of flight-mass spectrometry analysis confirmed that at least four of the polyketide synthase genes were involved in the biosynthesis of various pigments in P. marneffei, including melanin, mitorubrinic acid, mitorubrinol, monascorubrin, rubropunctatin, citrinin and ankaflavin, some of which were mycotoxins and virulence factors of the fungus.

Gordonia Species as Emerging Causes of Continuous-Ambulatory-Peritoneal-Dialysis-Related Peritonitis Identified by 16S rRNA and secA1 Gene Sequencing and Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS)

ABSTRACT We report here four cases of continuous ambulatory peritoneal dialysis-related peritonitis caused by three different species of Gordonia. The portal of entry was likely through Tenckhoff catheters. 16S rRNA and secA1 gene sequencing are so far the most reliable methods for the accurate identification of Gordonia species.

Subcutaneous Phaeohyphomycotic Nodule Due to Phialemoniopsis hongkongensis sp. nov.

ABSTRACT Phialemoniopsis species are ubiquitous dematiaceous molds associated with a wide variety of superficial and systemic infections in human. In this study, we isolated a mold from the forearm nodule biopsy specimen from a patient with underlying liver cirrhosis, ankylosing spondylosis, and tuberculosis. He was treated with itraconazole, but unfortunately, he succumbed as a result of disseminated tuberculosis with multiorgan failure. The histology results of the skin biopsy showed necrotizing granulomas in which numerous fungal elements were found. On Sabouraud dextrose agar, the fungal isolate grew as white-to-cream and smooth-to-velvety colonies. Microscopically, oval-to-cylindrical conidia were observed from abundant adelophialides, which possessed barely visible parallel collarettes but no basal septa. The azole drugs voriconazole, itraconazole, and posaconazole, as well as amphotericin B, showed high activities against this fungus. Internal transcribed spacer, 28S nuclear ribosomal DNA (nrDNA), and β-actin and β-tubulin gene sequencing showed that this fungus is most closely related to but distinct from Phialemonium curvata. Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) and hierarchical cluster analysis showed that the MALDI-TOF MS spectrum of this fungus is most closely related to that of Phialemonium pluriloculosa. We propose a new species, Phialemoniopsis hongkongensis sp. nov., to describe this fungus.

Fatal Fusarium solani species complex infections in elasmobranchs: the first case report for black spotted stingray (Taeniura melanopsila) and a literature review

Fusarium species are environmental saprophytic fungi. Among the many Fusarium species, members of the Fusarium solani species complex (FSSC) are the most prevalent and virulent in causing human and animal infections. In this study, we describe the first case of fatal FSSC infection in a black spotted stingray and three concomitant infections in scalloped hammerhead sharks. In the stingray, cutaneous lesions were characterised by ulcers and haemorrhage of the ventral pectoral fin, or ‘ray’, especially around the head; while cutaneous lesions in the sharks were characterised by ulcers, haemorrhage, as well as white and purulent exudates at the cephalic canals of the cephalofoil and lateral line. Histological sections of the cutaneous lesions revealed slender (1–4 μm in diameter), branching, septate fungal hyphae. Internal transcribed spacer region and 28S nrDNA sequencing of the fungal isolates from the fish showed two isolates were F. keratoplasticum (FSSC 2) and the other two were FSSC 12. Environmental investigation revealed the FSSC strains isolated from water and biofilms in tanks that housed the elasmobranchs were also F. keratoplasticum and FSSC 12. Fusarium is associated with major infections in elasmobranchs and FSSC 12 is an emerging cause of infections in marine animals. DNA sequencing is so far the most reliable method for accurate identification of Fusarium species.