1,721,633 research outputs found
A molecular epidemiology study on conjunctivitis using conventional nucleic acid amplification technologies and resequencing microarray
No full textpublished_or_final_versionMicrobiologyMasterMaster of Philosoph
Molecular epidemiology of erythromycin resistance in Streptococcus bovis and lancefield group G beta-hemolytic streptococci andhorizontal gene transfer of antibiotic resistance genes
No full textpublished_or_final_versionabstracttocMicrobiologyMasterMaster of Philosoph
Multilocus sequence typing of laribacter hongkongensis and other bacteria
No full textpublished_or_final_versionMicrobiologyDoctoralDoctor of Philosoph
Antifungal susceptibilities of emerging and novel filamentous fungal pathogens
No full textThe incidence of fungal diseases is increasing with the inevitable aging population and the expansion of immunocompromised patients. Through the advancement of molecular technologies, it has been revealed that many rare/cryptic and novel fungal species were associated with human diseases. However, knowledge of their antifungal susceptibility profiles and clinical spectrum is very limited, especially in Hong Kong. In the present study, it was hypothesized that the antifungal susceptibility profiles and clinical spectrum of these emerging and novel filamentous fungi are different compared to those of the commonly encountered species. The present project aimed to study the diversity of emerging and novel filamentous fungi and characterize their in vitro antifungal susceptibilities.
A total of 362 fungal isolates were included in this study. This study is divided into three parts. In part 1, 24 phenotypically non-dermatophyte, non-Aspergillus isolates were recovered from 24 independent cases of onychomycosis. In part 2, seven Talaromyces marneffei isolates were recovered from seven independent cases of systemic talaromycosis. In part 3, 331 non-duplicated Aspergillus isolates were recovered from cases of pulmonary and extrapulmonary aspergillosis. These fungi were studied by molecular phylogenetics (based on the ITS and other secondary DNA barcodes), or matrix-assisted laser desorption/ionization–time-of-flight mass spectrometry (MALDI–TOF MS), and antifungal susceptibility testing (AFST).
Among the 24 phenotypically non-dermatophyte, non-Aspergillus mold isolates, AFST showed that 33.3%, 41.7% and 95.8% of the 24 isolates exhibited minimum inhibitory concentrations (MICs) of >1 mg/L to terbinafine, itraconazole and fluconazole, respectively, which are the first-line treatment of onychomycosis. All seven T. marneffei isolates possessed low MICs to amphotericin B, itraconazole, voriconazole, posaconazole, and the newest approved isavuconazole, but high MICs to anidulafungin. As for the 331 Aspergillus isolates, AFST showed that the rare/cryptic species possessed very different susceptibility profiles when compared to the classical pathogenic Aspergillus species. Notably, most isolates of the cryptic species in series Nigri of section Nigri and series Versicolores of section Nidulantes were less susceptible to the triazole antifungals. Fortunately, majority of the cryptic species in section Nigri were not resistant and wild type to amphotericin B. They also exhibited low minimum effective concentrations (MECs) to all three echinocandins. On the other hand, amphotericin B is not useful for species in section Flavi, Terrei, Nidulantes and Circumdati. Overall, the geometric mean, MIC50 and MIC90 for posaconazole were the lowest amongst the four triazoles tested across all Aspergillus sections. Regarding to clinical spectrum, the rare/cryptic Aspergillus species had quite similar spectrum of infections when compared to the common/classical Aspergillus species.
In conclusion, a great diversity of less well-known fungi has been recognized as emerging pathogens in humans. Both accurate identifications to species level as well as AFST should be performed in clinical microbiology laboratories to guide proper patient management.published_or_final_versionMicrobiologyDoctoralDoctor of Philosoph
Modulation of Middle East respiratory syndrome coronavirus infection by human USP13 protein
No full textTwo major outbreaks of highly pathogenic coronaviruses, Severe Acute Respiratory Syndrome Coronavirus 1 (SARS-CoV-1) and Middle East Respiratory Syndrome Coronavirus (MERS-CoV), have been reported in 2003 and 2012 respectively. Most recently, a newly identified coronavirus, SARS-CoV-2, has emerged and causes more than 100 million human infection cases and two million deaths globally (as of March 2021). However, no clinically proven effective antiviral drug is available for the treatment of highly pathogenic coronavirus infections. There are two classes of antivirals: one targets the viral components such as polymerases or helicases, while another targets the host cellular factors that support the virus infection. To discover the host-targeting antivirals, the first step is to identify the key host factors or cellular pathways that support virus replication. In this study, the host factors or pathways were identified by the host-MERS-CoV interactomes study. More than 700 host factors that associate 25 MERS-CoV proteins were identified. Six of them (USP13, SIRT1, GSK3B, PTGES2, TTK and TPP2) were selected and further confirmed by immunoprecipitation. Commercially available small chemical inhibitors targeting these six factors or their corresponding pathways were used for further evaluation of antiviral efficacy against MERS-CoV. As expected, all these inhibitors potently suppressed MERS-CoV in human lung epithelial Calu-3 cells and human embryonic kidney 293 cells stably expressing DPP4, a receptor for MERS-CoV entry. It found that USP13 inhibitor showed the strongest inhibitory effect against MERS-CoV infection. USP13 is known as a deubiquitinase that can stabilize the host protein such as STAT1, a key transcription factor for type I IFN signaling. It was demonstrated that USP13 directly bound to MERS-CoV NSP16, and hence augment the NSP16 protein expression. This effect requires the ubiquitin-binding domain of USP13, but not depends on its catalytic domain. Lysine mapping showed that NSP16 lysine 249 and 253 were required for this enhancement of protein expression by USP13. Since the non-structural genes are conserved among different coronaviruses, it showed that USP13 also strongly enhanced the expression of NSP16 proteins derived from other five human coronaviruses (SARS-CoV-2, SARS-CoV-1, HCoV-229E, HCoV-NL63 and HCoV-OC43). Besides, USP13 specific inhibitor, Spautin-1, potently suppressed the three highly pathogenic human coronaviruses (MERS-CoV, SARS-CoV-1 and SARS-CoV-2) infection in vitro, suggested that USP13 is a potential host dependency factor for human coronavirus infection. Encouragingly, Spautin-1 also exhibited the same antiviral activity in SARS-CoV-2 infected human ex-vivo lung tissues. USP13 is the upstream deubiquitinase of Beclin-1/Vps34 complex that plays a key role on the omegasome formation. Therefore, USP13 may promote coronavirus infection on top of stabilizing the viral NSP16 protein. Indeed, it found that the Vps34 inhibitor (VPS34-IN1) strongly suppressed SARS-CoV-2 infection in both cultured cells and human ex-vivo lung tissues. Taken together, USP13 acts on both viral and host targets to promote human coronavirus infection. Thus, USP13 and its signaling pathways will be a good druggable target for the development of antivirals against highly pathogenic human coronaviruses including the recently emerged SARS-CoV-2.
(An abstract of 481 words)published_or_final_versionMicrobiologyDoctoralDoctor of Philosoph
Characterisation of defective interfering RNAs in avian influenza A (H7N9) virus
No full textDefective interfering (DI) virus is a virus particle that consists of functional structural viral proteins but has lost its essential genetic information. The presence of DI-RNA has been reported in different subtypes of influenza virus, including H1N1, H3N8, H7N7 and influenza B viruses. However, the mechanism of DI-RNA generation in influenza virus remains unclear. Since avian influenza A (H7N9) can cause severe illness in human, the existence of DI-RNA in H7N9-infected individuals is still unknown. By performing RT-PCR and Sanger sequencing, abundant DI-RNA production was observed in the nasopharyngeal aspirate (NPA) of H7N9-infected patients. To demonstrate the robustness of DI-RNA production in avian influenza A (H7N9) virus, the H7N9 reference strain, A/Anhui/1/2013 (AH1), was used to infect tissue-culture cells and mice. When comparing with a H1N1 laboratory strain (A/WSN/1933), the production of DI-RNA is higher in the AH1-infected NHBE cells. Single molecular real-time (SMRT) sequencing was applied to identify different DI-RNA species in H7N9-infected individuals. Identical PB1 DI-RNA species were observed in the NPAs of H7N9-infected patients. Moreover, long overlapping sequences (up to 23bp) were observed in the clinical specimens, indicating that similar sequences at the breakpoint sites may be a cause of DI-RNA production in H7N9 virus. Since the preliminary data demonstrated that the recombinant virus consisted of inconsistent influenza polymerase and NP segments (H5-3P) resulted in the abundant DI-RNA production. In this study, the H7-3P recombinant virus confirmed that inconsistent influenza polymerase and NP segments lead to the production of DI-RNA. Eight H7N9 strains were selected for future studies to further investigate the mechanism of DI-RNA generation in influenza virus. By performing amino acid alignments and constructing maximum likelihood phylogenetic trees, matching for the PB1 and NP segments was observed in most H7N9 viruses in the first three waves, and about half of the H7N9 viruses in the fifth wave. The eight selected strains were selected under the following criteria; (1) in the same wave and region with (2) similar PB2 and PA segment if possible. The production of DI-RNA in these eight selected strains would be investigated in the future studies.published_or_final_versionMicrobiologyMasterMaster of Philosoph
Genome-informed studies on Penicillium marneffei: horizontal gene transfer survey and differentialsecretomics
No full textpublished_or_final_versionMicrobiologyMasterMaster of Philosoph
A novel gene target for serodiagnosis and immunization of A. fumigatusaspergillosis
No full textpublished_or_final_versionMicrobiologyMasterMaster of Philosoph
Comparative analysis of Zika viral proteins in the inhibition of interferon signaling
No full textZika virus belongs to the family of Flaviridae. The virus was first isolated in 1947 in monkey of the Zika forest in Uganda. Human infection by the Zika virus was first reported in 1952. Since then, only about twenty additional human cases have been documented until 2006, and yet no or mild symptoms were observed in these patients. The virus was initially confined in Africa and Asia until the first outbreak in Yap Islands in the southwestern Pacific Ocean in 2007, where patients infected by the Asia-lineage Zika virus showed symptoms including conjunctivitis and rashes. In 2015-2016, the Zika virus of the same lineage caused a major epidemic in the Americas. An estimate of 1.5 million people in Brazil was infected by the Zika virus. More importantly, previously un-observed neurological symptoms including severe birth defects, such as microcephaly in new-born babies, and Guillain-Barré syndrome in adults were displayed by infected patients. In the past three years, investigation on the genetic differences between the original African strains and the current circulating Asian strains has been performed by many groups to study their causation to distinctive pathophysiological outcomes and enhanced human-to-human transmission. Type I interferon response of the host is the first-line defense mechanism against infections. Nevertheless, the differential dysregulation of this host signaling pathway by the two Zika virus lineages has not been fully characterized. In this study, we will compare the ability of individual Zika virus proteins from the two lineages in antagonizing the host Type I interferon response.
In this study, we first performed phylogenetic analysis and amino acid alignment to determine the amino acid differences of more than 600 African and non-African Zika virus sequences. 20 representative sequences of African and Asian strains were selected and aligned for the identification of the conserved substitutions. It encodes a single polyprotein that is proteolytically cleaved into three structural proteins (C, E, M) and seven non-structural proteins (NS1, 2A, 2B, 3, 4A, 4B, 5). The alignment results showed that NS5 has the largest number of amino acid substitutions. In addition, to study the interferon antagonistic properties of structural and non-structural proteins of the African and Asian Zika virus, we generated expression vectors for individual viral proteins of MR766 (African strain/Uganda/1947) and PRVABC-59 (Asian strain/Puerto Rico/2015) strains. Using dual luciferase reporter system, we showed that structural proteins had a less suppressive effect on type I interferon signaling, while non-structural proteins inhibited IFNβ, ISRE, IRF3, and NFκB to different extends. Moreover, non-structural proteins mainly suppressed the RIG-I induced IFNβ activity. On the other hand, NS5 has the greatest inhibitory effect on RIG-I induced type I interferon signaling among all Zika virus-encoded proteins (439 words).published_or_final_versionMicrobiologyMasterMaster of Philosoph
Antibiotic resistance in laribacter hongkongensis
No full textpublished_or_final_versionMicrobiologyDoctoralDoctor of Philosoph
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