Aspergillus species are responsible for a variety of human diseases, ranging from allergic bronchopulmonary aspergillosis to invasive aspergillosis. Identification of Aspergillus species could facilitate the selection of antifungal regimens and epidemiological studies. Most of the clinical microbiology laboratories identify Aspergillus species by traditional phenotypic and/or antigen detection methods, which are laborious, time-consuming and inaccurate. In recent years, sequence analysis of β-tubulin and calmodulin genes becomes widely used for the identification of fungal species due to their relatively high resolving power, universality of usage, and high availability in the public databases. It can also be used for the taxonomic classification and the identification of rare and even novel fungal species. In this study, we aim to evaluate the effectiveness of analyzing the β-tubulin and calmodulin gene sequences for the identification of Aspergillus species and subsequently to determine the clinical spectrum of Aspergillus infections in Hong Kong during 2012-2014. In this study, 48 Aspergillus strains isolated from patients over a 3-year period were characterised to the species level using sequencing of β-tubulin and calmodulin genes and the clinical spectrum of the 48 patients was described. Sequencing of β-tubulin and calmodulin genes showed that all 48 strains were known Aspergillus species. Ten different Aspergillus species were identified, including A. fumigatus (n=16), Aspergillus species of the A. flavus clade (n=7), A. awamori (n=7), A. terreus (n=6), A. tubingensis (n=4), A. sydowii (n=3), A. pseudocaelatus (n=2), A. uniguis (n=1), Aspergillus species of the A. tamarii clade (n=1), and A. austroafricanus (n=1). These Aspergillus species were shown to be associated with respiratory infections, infections of nail, ear canal infections, invasive infections, and eye infection. This study also described the first reported cases of ear infection associated with A. pseudocaelatus. To conclude, sequence analysis of β-tubulin and calmodulin genes enables accurate and rapid Aspergillus species differentiation. / published_or_final_version / Microbiology / Master / Master of Medical Sciences

No description available.

Thesis (M.S.)--University of Puerto Rico, Mayagüez Campus, 2008. / Tables. Printout. Abstract in English and Spanish. Includes bibliographical references (leaves 82-87).

No description available.

Background: Invasive pulmonary aspergillosis (IPA) is a common cause of mortality in haemato-oncology patients and early diagnosis is vital for improving outcomes. Since lung biopsy in this acute setting is rarely performed due to the associated risks, an empirical strategy remains the standard of care in many haematology units, but leads to overtreatment with antifungal drugs, which have significant side-effects. This project has developed novel approaches for detecting IPA, allowing early and specific treatment of genuine fungal infection. Methods: A combination marker approach involving a new Aspergillus qPCR assay, an EORTC/MSG-endorsed GM ELISA and an Aspergillus LFD, was used to establish a robust diagnosis of IPA from clinical broncho-alveolar lavage (BAL) fluid samples. The inflammatory cytokine profile associated with IPA biomarker positive BAL fluid was also evaluated. Finally, antigen and qPCR detection were combined in a proximity ligation assay (PLA), to demonstrate proof-of-principle for a diagnostic assay for the earliest possible detection of fungal infections. Results: A dual testing approach involving a novel MIQE-compliant Aspergillus qPCR assay and an Aspergillus LFD showed a sensitivity and specificity of 100% and 94%, respectively in BAL fluid, unlike in blood where this approach was not sensitive. Results confirmed previously published concerns over the repeatability of GM in serum, whereas BAL GM results appear stable. Biomarker detection results in exhaled breath condensate did not correlate well with results in BAL fluid samples. Respiratory samples did not identify a distinct inflammatory marker profile in IPA. Finally, antibodies raised against JF5 mannoprotein were used to develop a PLA test to detect active growth of Aspergillus. Conclusions: The optimised qPCR is a very sensitive and highly specific aid in IPA diagnosis. A combination biomarker approach could be incorporated into a diagnostic-driven approach to patient management to direct antifungal treatment to patients with evidence of invasive fungal disease.

Thesis (M. Phil.)--University of Hong Kong, 2002. / Includes bibliographical references (leaves 101-135).

Thesis (Ph. D.)--Oregon State University, 2009. / Printout. Includes bibliographical references (leaves 312-323). Also available on the World Wide Web.

Aspergillus fumigatus is an opportunistic fungus known to cause a group of life-threatening infections collectively known as aspergillosis. In this thesis, multilocus sequence and microsatellite markers, among others, were used to study global and local A. fumigatus population structures. We examined the roles of sexual and asexual reproduction in the initiation of azole resistance globally. Furthermore, we investigated the origin of multi-triazole resistance in India and whether the use of fungicides on farms propagates resistance in environmental strains of clinical importance in Hamilton, Ontario. We characterized for the first time the A. fumigatus population in Cameroon while concomitantly screening for environmental resistance. Our results showed that sexual reproduction plays a key role in the development of triazole resistance globally. We found that multi-triazole resistance in India has multiple origins, which include mutational, recombinational and exotic origins. Our results provided little to no evidence that azole fungicides are the origin, or increase the frequency of triazole resistance in clinical A. fumigatus in Hamilton. Additionally, we identified a significantly unique A. fumigatus population in Cameroon. Our findings will potentially contribute towards developing effective long-term management strategies against aspergillosis. / Thesis / Doctor of Philosophy (PhD) / Aspergillus fumigatus is a mold capable of causing severe infection in humans. Infections caused by A. fumigatus can often be treated using antifungals. However, there have been several reported cases of treatment failure around the world over the last two decades. Generally speaking, treatment failure in patients is often associated with antifungal resistance in A. fumigatus. My thesis aims at better understanding the distribution and investigating the origin of resistance in A. fumigatus at both global and local levels. Here, we analyze A. fumigatus strains from 15 countries, including strains from Hamilton, Ontario. Our findings will potentially contribute towards establishing effective long-term management strategies against A. fumigatus infections locally and globally.

Aspergillus fumigatus is the major etiology of invasive aspergillosis, a leading cause of death in immunocompromised patients. Septins are conserved GTPases that could be mediating host-spore interactions in A. fumigatus. To test this hypothesis, we are using a combination of microscopy techniques and cell culture-based methods. Using Atomic Force Microscopy, we found that spores from the ∆aspB strain have a disorganized rodlet layer compared to the parent strain. Disorganized rodlet layer can lead to an increase in immune recognition of common pathogen associated molecular patterns (PAMPs), chitin and β-glucan. It is unknown whether the increase in immune recognition of ∆aspB spores is due to more exposure to the host or if they total quantity of PAMPs has increased. To test this, we used several staining methods to compare total and exposed levels of PAMPs in the conidia. Our work shows that there is an increase in exposed chitin and total levels of chitin and β-glucan. To evaluate how this plays a role in vitro, we first exposed macrophage-like cells (J744.1) to septin deletion strain’s spores and measured the TNF-a to determine spore immunogenicity using an ELISA. As expected, only the deletions that abolish all septin complexes had a significant increase in TNF-a. Then, we determine how effective macrophages were at killing the spores. We found that the deletion strains were more susceptible to killing by the macrophages. A murine study revealed that ∆aspB infection results in more lung inflammation than the wildtype. Taken together, these results suggest the septin cytoskeleton is involved in A. fumigatus spore cell wall organization and immune evasion.

published_or_final_version / Microbiology / Master / Master of Philosophy