Please use this identifier to cite or link to this item: https://hdl.handle.net/10321/4829
Title: Investigation of antifungal compounds against Aspergillus species from respiratory infections of immunocompromised patients
Authors: Juglal, Sarla 
Issue Date: May-2023
Abstract: 
The rapid emergence of invasive fungal infections correlates with the increasing population
of immunocompromised individuals, with many cases leading to death. The progressive
increase in the incidence of Aspergillus isolates is even more severe due to clinical
challenges in treating invasive respiratory infections in immunocompromised patients.
Azoles are the drugs of choice for the prevention and treatment of Aspergillus infections
however, azole resistance in Aspergillus-related infections is an increasing concern,
especially against Aspergillus fumigatus. This situation is further complicated by the use of
azoles in agriculture, as increasing resistance has been associated with increased use in
agriculture. Rapid initiation in detection, diagnosis and appropriate antifungal therapy is
needed to reduce mortality among individuals with invasive Aspergillus-related infections.
Improvements in all aspects of azole resistance management, from identification and
antifungal therapy to the discovery of novel non-toxic drugs, impact clinical success. Hence,
this study assessed the effect of routine and salvage drug therapy on Aspergillus respiratory
infections and investigated the potential of new drugs in combination therapy in light of
increasing antifungal resistance in the management of invasive fungal infections. Aspergillus
species isolated from immunocompromised patients with respiratory infections at the Inkosi
Albert Luthuli Hospital in Kwa-Zulu Natal were investigated. Conventional morphology
identification methods were assessed for reliability and compared to molecular
identification. In the search for reliable and rapid alternatives in fungal identification to
impact diagnosis, phenotypic microarray (Biolog) for fungal species identification was
explored. Phenotypic microarray (Biolog) was also used to analyse the isolates' nutritional
patterns and drug sensitivity profiles to investigate the potential for new compounds in drug
therapy. In addition, gliotoxin expression, which has been linked to increased pathogenesis,
was quantified and analysed using high-performance liquid chromatography. This was
investigated in vitro to correlate gliotoxin production as a possible virulence factor in azole
resistance. The effect of pathogen-associated molecular patterns (PAMPS) in bacterial coinfection was also investigated, especially since the species were isolated from respiratory
infections where co-infection is common. Conventional morphological techniques were generally similar to 18S rRNA identification,
assigning twenty-six Aspergillus fumigatus species, eight A. niger and two A. flavus. Biolog
technology only identified isolates correctly up to genus level and had no significant
similarity matches at species level due to the Biolog fungal database not having adequate
reference clinical species however, an updated database makes this technology a good
alternative when reliability and speed in identification are considered. Antifungal profiles
showed that 6% of the 36 isolates were resistant to the routine azole voriconazole, with 61%
having moderate susceptibility. All isolates resistant to the salvage therapy drug,
posaconazole pose a serious concern. Significantly, A.niger was the only species resistant
(25%) to voriconazole and has recently been reported as the species isolated from patients
with COVID-19-associated pulmonary aspergillosis (CAPA). Mutations in the cpy51A gene
representing common mechanisms for azole resistance in clinical Aspergillus isolates were
highlighted in the azole susceptibility profile. Phenotypic microarray showed that 83% of
the isolates were susceptible to the 24 new compounds. Berberine and blasticidin
hydrochloride were selected and further investigated for combination drug therapy,
considering their non-toxicity upon oral administration currently with thirty of the thirty-six
isolates showing susceptibility to the new agents.
Carbon profiles demonstrated the consistent assimilation of alternate monosaccharides and
disaccharides by all three Aspergillus species, accomplished by secreted enzymes which
very likely contribute to nutrient acquisition during infection, impact persistent growth and
survival to influence infection maintenance and disease progression. Of the twenty-six A.
fumigatus isolates, 58% produced gliotoxin during the 48-hour incubation at 37℃. Forty per
cent of those gliotoxin-producing isolates showed increased gliotoxin production after
exposure to PAMPS, with the highest concentration of 2.6 mg/L and a mean of 0.6 mg/L.
Compared to similar investigations, the gliotoxin concentrations for invasive aspergillosis
were higher than for colonization in respiratory patients.
This study found that different identification strategies in azole resistance treatment
management are required for both developed and resource-challenged situations to influence
diagnosis. Drug resistance in azole monotherapy was demonstrated. Thus this study supports
the search for new agents in drug regimen strategy and found potential in novel compounds,
berberine and blasticidin hydrochloride, for combination therapy.
Description: 
Submitted in fulfilment of the requirement for the Degree of Doctorate of Philosophy in Biotechnology, Durban University of Technology, Durban, South Africa, 2023.
URI: https://hdl.handle.net/10321/4829
DOI: https://doi.org/10.51415/10321/4829
Appears in Collections:Theses and dissertations (Applied Sciences)

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