Comparative evaluation of VITEK® 2 and three commercial gradient strip assays for daptomycin susceptibility testing of Staphylococcus aureus

Background Physicochemical and microbiological methods are found in the literature for the analysis of daptomycin, an antimicrobial. Objective This paper brings a miniaturized turbidimetric microbiological method for analysis of daptomycin in lyophilized powder. Methods The method was performed using 96-well microplates, 4-hours incubation, 2, 4 and 8 μg/mL, S. aureus ATCC 6538 IAL 2082 at 7% and BHI broth. Results Linearity was proven by obtaining analytical curves with correlation coefficient greater than 0.99 and statistical evaluation by Analysis of Variance (ANOVA). The method was also selective, since the standard and sample analytical curves were parallel, proving that the excipient does not interfere with daptomycin analysis. Intraday, interday and inter-analyst precision presented RSD of 2%, 2.27% and 1.08%, respectively. Accuracy was assessed by the recovery test, where known quantities of standard solution are added to the sample and an average recovery value of 100.73% (RSD = 0.71%) was obtained. The present method was robust when minor changes were made in the parameters of used antimicrobial volume, inoculum volume and incubation time. Conclusions This work has an innovative and ecological proposal and advantages such as (i) less waste generation, (ii) miniaturized quantities of sample, culture media and inoculum, (iii) no need to use formaldehyde as in the traditional turbidimetric method, (iv) lower volume of glassware used and (v) shorter incubation time compared to other methods as agar diffusion requiring approximately 24 hours. Highlights This work is extremely important and contemplates a current, innovative and sustainable theme for pharmaceutical analysis around the world.

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An Ecological and Miniaturized Biological Method for the Analysis of Daptomycin Potency

Richardi

Kogawa

Belavenuto

et al. 2020

Journal of AOAC INTERNATIONAL

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Genome-Wide Association Studies for the Detection of Genetic Variants Associated With Daptomycin and Ceftaroline Resistance in Staphylococcus aureus

et al. 2021

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BackgroundAs next generation sequencing (NGS) technologies have experienced a rapid development over the last decade, the investigation of the bacterial genetic architecture reveals a high potential to dissect causal loci of antibiotic resistance phenotypes. Although genome-wide association studies (GWAS) have been successfully applied for investigating the basis of resistance traits, complex resistance phenotypes have been omitted so far. For S. aureus this especially refers to antibiotics of last resort like daptomycin and ceftaroline. Therefore, we aimed to perform GWAS for the identification of genetic variants associated with DAP and CPT resistance in clinical S. aureus isolates.Materials/methodsTo conduct microbial GWAS, we selected cases and controls according to their clonal background, date of isolation, and geographical origin. Association testing was performed with PLINK and SEER analysis. By using in silico analysis, we also searched for rare genetic variants in candidate loci that have previously been described to be involved in the development of corresponding resistance phenotypes.ResultsGWAS revealed MprF P314L and L826F to be significantly associated with DAP resistance. These mutations were found to be homogenously distributed among clonal lineages suggesting convergent evolution. Additionally, rare and yet undescribed single nucleotide polymorphisms could be identified within mprF and putative candidate genes. Finally, we could show that each DAP resistant isolate exhibited at least one amino acid substitution within the open reading frame of mprF. Due to the presence of strong population stratification, no genetic variants could be associated with CPT resistance. However, the investigation of the staphylococcal cassette chromosome mec (SCCmec) revealed various mecA SNPs to be putatively linked with CPT resistance. Additionally, some CPT resistant isolates revealed no mecA mutations, supporting the hypothesis that further and still unknown resistance determinants are crucial for the development of CPT resistance in S. aureus.ConclusionWe hereby confirmed the potential of GWAS to identify genetic variants that are associated with antibiotic resistance traits in S. aureus. However, precautions need to be taken to prevent the detection of spurious associations. In addition, the implementation of different approaches is still essential to detect multiple forms of variations and mutations that occur with a low frequency.

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Comparative evaluation of VITEK® 2 and three commercial gradient strip assays for daptomycin susceptibility testing of Staphylococcus aureus

Cited by 2 publications

References 14 publications

An Ecological and Miniaturized Biological Method for the Analysis of Daptomycin Potency

An Ecological and Miniaturized Biological Method for the Analysis of Daptomycin Potency

Genome-Wide Association Studies for the Detection of Genetic Variants Associated With Daptomycin and Ceftaroline Resistance in Staphylococcus aureus

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