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Background: Linezolid-resistant Enterococcus faecium (LRE) is a global priority pathogen. Thirteen LRE were reported from clinical specimens between November 2021 and April 2023 at two laboratories in Karachi, Pakistan. We aimed to investigate the strain types and genes associated with linezolid resistance among these isolates. Whole genome sequencing (WGS) was performed and analyzed by multilocus sequence typing (MLST). The presence of linezolid resistance genes was identified using ResFinder v4.1.11 and the LRE-finder tool. Results: Twelve isolates belonged to clonal complex 17 (CC17); ST80 (n = 10), ST612 (n = 1) and ST1380 (n = 1). Six isolates showed the presence of optrA gene and G2576T mutations in the 23S rRNA gene, while six showed poxtA and cfr(D) genes. One isolate showed the combination of optrA, cfr(D) and poxtA genes. Conclusion: Our findings show the circulation of CC17 sequence types with a known outbreak potential and we identified molecular mechanisms of resistance that were not previously reported from Pakistan. [ABSTRACT FROM AUTHOR]

Chronic pulmonary aspergillosis (CPA) is a recognized complication of pulmonary tuberculosis (TB). In 2015, the World Health Organization reported 2.2 million new cases of nonbacteriologically confirmed pulmonary TB; some of these patients probably had undiagnosed CPA. In October 2016, the Global Action Fund for Fungal Infections convened an international expert panel to develop a case definition of CPA for resource-constrained settings. This panel defined CPA as illness for >3 months and all of the following: 1) weight loss, persistent cough, and/or hemoptysis; 2) chest images showing progressive cavitary infiltrates and/or a fungal ball and/or pericavitary fibrosis or infiltrates or pleural thickening; and 3) a positive Aspergillus IgG assay result or other evidence of Aspergillus infection. The proposed definition will facilitate advancements in research, practice, and policy in lower- and middle-income countries as well as in resource-constrained settings.

Background: Recent reports of the emergence of fluconazole resistance in Candida parapsilosis species complex poses a challenge, more specifically in settings where echinocandin-based treatment regime is not feasible. Objective: This study reported emergence of fluconazole resistance in C. parapsilosis species complex strains isolated from blood cultures. Materials and Methods: This retrospective observational study was conducted from 2018 to 2020 at a tertiary care laboratory from Pakistan. Fluconazole-resistant C. parapsilosis species complex fungemia cases were identified from laboratory database and clinical details were collected. Identification of C. parapsilosis species complex was done using API 20C AUX and Cornmeal Tween80 agar morphology. Minimum inhibitory concentrations (MICs) were determined using Sensititre YeastONE and interpretation was done with CLSI M60 ED1:2017. ERG11 gene region was amplified and sequenced by Sanger sequencing and analysed by MEGA 11 Software. Results: A total of 13 (8.5%) fluconazole-resistant isolates were identified from 152 C. parapsilosis species complex candidemia cases. Fluconazole MICs of resistant isolates ranged between 8 and 256 µg/mL. Analysis of ERG11 gene revealed nonsynonymous mutations at position Y132F in 86% of the fluconazole-resistant isolates. Diabetes and hospitalization were important risk factors for candidemia with fluconazole-resistant C. parapsilosis complex. Conclusion: This is the first report of the emergence and molecular mechanisms of fluconazole resistance in C. parapsilosis species complex from Pakistan. Y132F mutation in the ERG11 gene was the most common mutation in fluconazole-resistant strains. These findings are concerning and necessitate better diagnostics, newer antifungals, ongoing surveillance and further insights on resistance mechanisms in the country. [ABSTRACT FROM AUTHOR]

Introduction: The gradual increase in caspofungin usage in Pakistan raises a concern of emergence of echinocandin resistance in local Candida glabrata strains. We sequenced and determined mutations in fks1 and fks2 genes in invasive Candida glabrata strains from Pakistan. Material and Methods: Thirty‐six invasive C. glabrata strains were selected with median (min‐max) minimum inhibitory concentrations (MICs) of 0.06 (0.015–0.25) mg/L for caspofungin, 0.015 (0.008–0.06) mg/L for micafungin and 0.06 (0.015–0.12) mg/L for anidulafungin. fks1 and fks2 gene fragments were sequenced using Sanger methodology. Sequences were analysed with MEGA‐6 software to identify specific single‐nucleotide polymorphisms (SNP) against wild‐type sequences of C. glabrata. Results: In fks1 gene, non‐synonymous mutation D632H was observed in one isolate with caspofungin MIC of 0.25 mg/L. Synonymous mutation at position A742 was observed in 26/36 (72%) of the isolates. 34/36 (94.5%) isolates analysed for fks2 gene were observed as wild type. A novel non‐synonymous mutation at I661T was observed in fks2 gene in one isolate with caspofungin MIC of 0.12 mg/L and anidulafungin and micafungin MIC of 0.06 and 0.015 mg/L, respectively. Novel fks2 synonymous mutations at position T647, K652 and I706 were observed in 16/36 (44%), 25/36 (69%) and 23/36 (63%) isolates, respectively. Conclusion: Low frequencies of both non‐synonymous and synonymous polymorphisms were observed in invasive C. glabrata strains. Since S663P in fks2 gene is associated with caspofungin resistance, a novel mutation at 661 codon identified in our study needs correlation with treatment outcome data and mandates periodic genomic surveillance. [ABSTRACT FROM AUTHOR]

Background: Early identification of COVID‐19‐associated pulmonary aspergillosis (CAPA) is particularly challenging in low‐ middle‐income countries where diagnostic capabilities are limited, and risk factors for CAPA have not been identified. It is also essential to recognise CAPA patients who are likely to have a poorer outcome to decide on aggressive management approaches. Therefore, this study aimed to identify risk factors and outcomes for CAPA among admitted moderate to critical COVID‐19 patients at our centre in Pakistan. Methods: An unmatched case–control study with ratio of 1:2 was conducted on hospitalised adult patients with COVID‐19 from March 2020–July 2021. Cases were defined according to European Confederation of Medical Mycology and the International Society for Human and Animal Mycology consensus criteria. Controls were defined as patients hospitalised with moderate, severe or critical COVID‐19 without CAPA. Results: A total of 100 CAPA cases (27 probable CAPA; 73 possible CAPA) were compared with 237 controls. Critical disease at presentation (aOR 5.04; 95% CI 2.18–11.63), age ≥ 60 years (aOR 2.00; 95% CI 1.20–3.35) and underlying co‐morbid of chronic kidney disease (CKD) (aOR 3.78; 95% CI 1.57–9.08) were identified as risk factors for CAPA. Patients with CAPA had a significantly greater proportion of complications and longer length of hospital stay (p‐value <.001). Mortality was higher in patients with CAPA (48%) as compared to those without CAPA (13.5%) [OR = 6.36(95% CI 3.6–11)]. Conclusions: CAPA was significantly associated with advanced age, CKD and critical illness at presentation, along with a greater frequency of complications and higher mortality. [ABSTRACT FROM AUTHOR]