We investigated the molecular mechanisms underlying azole resistance in seven Candida tropicalis isolates that caused candidemia and candiduria in Paraná, Brazil (2016–2022). Biofilm production, antifungal susceptibility testing, multilocus sequence typing, amplification and sequencing of ERG11, and quantification of ERG11, MDR1, and CDR1 expression levels were performed. Notably, five isolates (71.4%) were from urine samples and two (28.6%) were from blood samples. All strains were biofilm producers, with levels ranging from moderate to strong. The minimum inhibitory concentration (MIC) values ranged from 8–>64 mg/L for fluconazole and 0.25–1 mg/L for voriconazole. All isolates had mutations in ERG11; Y132F and Y257N were predominant (71.4%), followed by Y132F and S154F (14.3%) and Y257H (14.3%). No differences in ERG11 expression were found between the susceptible and resistant groups, but MDR1 and CDR1 were more highly expressed in the susceptible isolates. All the isolates contained previously unassigned diploid sequence types. The emergence of C. tropicalis azole resistance has been previously described in Brazil; however, the presence of resistant isolates in urine highlights the need for surveillance resistant strains in both urinary and invasive contexts. In our study mutations in ERG11 were the main resistance mechanism identified in C. tropicalis.

Invasive candidiasis (IC) contributes to the morbidity and mortality of hospitalized patients and represents a significant burden to the healthcare system. Previous Brazilian studies have reported the presence of endemic Candida parapsilosis sensu stricto genotypes causing candidemia and clonal transmission involving fluconazole-resistant isolates. We performed a 5-year retrospective analysis of IC cases in a Brazilian tertiary pediatric hospital and conducted a molecular investigation of C. parapsilosis sensu stricto. Non-duplicate C. parapsilosis sensu stricto genotyping was performed by microsatellite analysis. Antifungal susceptibility and biofilm formation were also evaluated. A total of 123 IC episodes were identified, with an IC incidence of 1.24 cases per 1000 hospital admissions and an overall mortality of 34%. The main species were the C. parapsilosis complex (35.8%), Candida albicans (29.2%), and Candida tropicalis (21.9%). All C. parapsilosis sensu stricto were recovered from blood cultures, and 97.5% were biofilm producers. Microsatellite typing identified high genotypic diversity among the isolates. We observed that all isolates were sensitive to amphotericin B, and although one isolate was non-sensitive to fluconazole, only a silent mutation on ERG11 gene was identified. No clear evidence of clonal outbreak or emergence of fluconazole-resistant isolates was found, suggesting that multiple sources may be involved in the epidemiology of IC in children.

We aimed to investigate an outbreak of vancomycin-resistant Enterococcus faecium (VREfm) in paediatric patients from Hospital Pequeno Príncipe. The susceptibility profile was determined, and whole-genome sequencing (WGS) was used to analyse the genetic context of the strains. Five VREfm isolates were recovered from sterile sites and surveillance cultures of two paediatric patients with acute lymphoblastic leukaemia. Species identification was performed using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS), and the minimum inhibitory concentration (MIC) was assessed according to the European Committee for Antimicrobial Susceptibility Testing (EUCAST). WGS was performed to analyse the genetic context of virulence and resistance genes, and in silico multilocus sequence typing was performed to identify the sequence typing of the strains. High-level vancomycin resistance was observed in all isolates (≥256 mg/L). WGS revealed the presence of mobile genetic elements, such as plasmids (rep₂, rep_11a, rep_US15, rep₁₇, and rep_18a), insertion sequences, and phages. Multiple resistance genes (aac(6')-aph(2"), dfrG, ermB, and vanA) and virulence genes (acm and efaAfm) were identified. All the isolates were assigned to ST117 (ST1133 - via a novel MLST), an important epidemic lineage associated with nosocomial infections and outbreaks. Our results show that the ST117 (ST1133) VREfm isolates are circulating in paediatric patients, which raises a great concern. The development of new drugs as well as the implementation of an antimicrobial stewardship program are necessary for their correct management, limiting the spread of resistance in oncohematological patients.

Methicillin-resistant Staphylococcus aureus (MRSA) is a leading cause of hospital- and community-acquired infections (HA-MRSA and CA-MRSA, respectively). Although this microorganism is associated with childhood bacteremia, its epidemiology and predictive factors are not well established. In this study, the susceptibility profiles and molecular characteristics of 46MRSA isolates obtained from blood samples and other sterile fluids of pediatric patients between January 2017 and May 2022 were characterized. The isolates were identified using matrix-assisted laser desorption ionization-time of flight mass spectrometry. After that, their antimicrobial susceptibility profiles were assessed trough broth microdilution, and their molecular characteristics was performed by whole genome sequencing. Antibiotic susceptibility tests showed that the isolates were 100% sensitive to vancomycin, linezolid, and sulfamethoxazole-trimethoprim. In terms of SCCmec, types IV and II were the most prevalent, accounting for 47.82% (22/44) and 47.72% (21/44) of the isolates, respectively, whereas type I was identified in just one isolate (2.27%). Thus, 50% of infections were caused by CA-MRSA and HA-MRSA. Sequencing analysis revealed polyclonal spread, with several sequence types (STs) identified. Five STs belonging to the CC5 clonal complex (ST5, ST105, ST1649, ST1176, and ST1307) accounted for most of the isolates (n=38). The detected virulence genes were sak, tst-1, ACME, hlgA, lukF-PV/lukS-PV, and lukD/lukE. The Panton–Valentine Leukocidin (PVL) toxin gene was found in nine isolates, all of which were CA-MRSA and carried SCCmec type IV. Several antimicrobial resistance genes encoding resistance to different antibiotic classes were identified. In conclusion, we conclude that CA-MRSA strains are currently ubiquitous in hospital environments, causing serious infections, with CC5 being predominant over the past five years.

Background/Objectives: Acute lymphoblastic leukemia (ALL) is the most common childhood malignancy. Understanding its epidemiological characteristics is essential for guiding public health strategies. In this study, we characterized the epidemiological profiles that may contribute to the risk of ALL in children in southern Brazil. Methods: Clinical and epidemiological data from 71 children (1–15 years old) admitted and newly diagnosed with ALL at four hospitals in Curitiba, Paraná, Brazil, were retrieved and analyzed. Results: Among the 71 children with ALL, the majority were male (n = 43, 60.6%), with an age range of 1–3 years (n = 26, 36.6%), self-identified as White (n = 47, 66.2%), and were born in Paraná state (n = 61, 85.9%). Nearly half had a family history of cancer (n = 33, 46.5%), primarily among grandparents (n = 36, 61%). Parental environmental exposures included smoking (n = 30, 42.3%) and occupational exposure to chemicals or radiation (n = 17, 23.9%). At diagnosis, most patients (n = 43, 60.5%) had a bone marrow blast count > 70%, and 27 patients (38%) had a peripheral blood blast count > 70%. B-cell ALL was the predominant subtype (n = 61, 85.9%). In B-cell ALL cases, the most frequent molecular subtype was high hyperdiploidy (n = 17, 23.9%). White blood cell counts differed significantly between the B-cell ALL and T-cell ALL groups (p = 0.029). Conclusions: Our findings provide insights into ALL epidemiology in southern Brazil and highlight regional differences across the country.

Methicillin-resistant Staphylococcus aureus (MRSA) is a leading cause of hospital- and community-acquired infections (HA-MRSA and CA-MRSA, respectively). Although this microorganism is associated with childhood bacteremia, its epidemiology and predictive factors are not well established. In this study, the susceptibility profiles and molecular characteristics of 46MRSA isolates obtained from blood samples and other sterile fluids of pediatric patients between January 2017 and May 2022 were characterized. The isolates were identified using matrix-assisted laser desorption ionization-time of flight mass spectrometry. After that, their antimicrobial susceptibility profiles were assessed trough broth microdilution, and their molecular characteristics was performed by whole genome sequencing. Antibiotic susceptibility tests showed that the isolates were 100% sensitive to vancomycin, linezolid, and sulfamethoxazole-trimethoprim. In terms of SCCmec, types IV and II were the most prevalent, accounting for 47.82% (22/44) and 47.72% (21/44) of the isolates, respectively, whereas type I was identified in just one isolate (2.27%). Thus, 50% of infections were caused by CA-MRSA and HA-MRSA. Sequencing analysis revealed polyclonal spread, with several sequence types (STs) identified. Five STs belonging to the CC5 clonal complex (ST5, ST105, ST1649, ST1176, and ST1307) accounted for most of the isolates (n=38). The detected virulence genes were sak, tst-1, ACME, hlgA, lukF-PV/lukS-PV, and lukD/lukE. The Panton–Valentine Leukocidin (PVL) toxin gene was found in nine isolates, all of which were CA-MRSA and carried SCCmec type IV. Several antimicrobial resistance genes encoding resistance to different antibiotic classes were identified. In conclusion, we conclude that CA-MRSA strains are currently ubiquitous in hospital environments, causing serious infections, with CC5 being predominant over the past five years.