| Congresso Brasileiro de Microbiologia 2023 | Resumo: 304-2 | ||||
Resumo:Antimicrobial resistance (AMR) constitutes a major threat to public health worldwide. Infections stemming from AMR bacteria are linked to increased length of hospital stay and to higher morbidity and mortality rates. Successful treatments of AMR bacterial infections are contingent upon rapid strain identification as well as AMR and virulence profiling. In addition, AMR- and virulence-encoding genes are often associated with mobile genetic elements (MGE), making their identification important for the investigation of the dissemination potential. Whole genome sequencing (WGS) has been recommended as a “one-stop” diagnostic tool due to its capability to provide multiple and simultaneous diagnostic and epidemiological data about the microorganism. In 2021, our research group developed and validated a rapid (<33h) nanopore-based WGS protocol for surveillance and outbreak investigation. To evaluate the potential of this protocol for complementary diagnostics, the current study aims to assess AMR- and virulence-related genes and MGE, comparing the results with four other slower sequencing protocols. The sequencing protocols included for each bacterial strain: three nanopore sequencing (varying the sequencing time and the assembly methodology), one sequencing on the Illumina platform, and one hybrid assembly protocol using nanopore and Illumina reads. The study enrolled 42 genomes of methicillin-resistant Staphylococcus aureus (MRSA) and six genomes of extended-spectrum β-lactamases (ESBL)-producing Klebsiella pneumoniae. MGE, AMR- and virulence-encoding genes were detected using online and free-of-charge bioinformatics tools. There was no significant difference in the ability to detect virulence- and AMR-related genes among the protocols for MRSA (p values > 0,05). When considering ESBL-K. pneumoniae, the protocol based on Illumina sequencing detected considerably fewer AMR-related genes than the other protocols (p = 0,003), although there was no difference in the detection of virulence genes (p > 0,05). Additionally, the detection of plasmids and MGE were significantly different among the protocols for MRSA (p values = 0,0005 and 0,04, respectively) and ESBL-K. pneumoniae (p values = 0,0003 and 0,002, respectively). Accuracy data in inferring resistance from analyzes of genomes obtained by the rapid protocol developed previously reports a sensitivity of 100% (Confidence Interval [CI] of 94.04% to 100%) and specificity of 99,58% (CI of 98.78% to 99.96%) for MRSA and a sensitivity of 100% (CI of 93.15% to 100%) and specificity of 80% (CI of 56.34% to 94.27%) for ESBL-K. pneumoniae. The rapid protocol developed previously showed results as good and even better than the other slower protocols tested, strengthening its potential for application in the clinical routine. Overall, genomic analysis based on nanopore sequencing is a promising alternative in the effort to provide rapid and accurate diagnostics and epidemiological data. However, more studies are needed for its implementation, considering that the analyses depend on the cost of the inputs, the availability of bioinformatics tools, and the quality of their reference databases. Palavras-chave: Genomic analysis, Antimicrobial resistance, diagnosis, virulence, whole genome sequencing Agência de fomento:UNIEDU/FUMDES PÓS-GRADUAÇÃO | |||||