CCNE

CCNE estimates copy numbers of carbapenemase-encoding genes from whole-genome sequencing data to quantify gene dosage of blaCarb-family genes relevant to antimicrobial resistance.

Key Features:

• Target genes: Estimation focused on carbapenemase-encoding genes of the blaCarb family, including blaKPC.
• Input data: Operates on whole-genome sequencing read data mapped to reference loci.
• Reference normalization: Uses housekeeping genes as internal references to normalize read counts.
• Copy-number estimation: Compares read counts mapped to antimicrobial resistance (AMR) genes against counts mapped to reference housekeeping genes to infer copy numbers.
• Modular design: Includes CCNE-acc for comprehensive high-accuracy detection and CCNE-fast for rapid screening.
• Validation and performance: Evaluated by simulation studies and qPCR validation; CCNE-acc achieved 100% accuracy with RMSE 0.07 versus assembly-based methods (23.4% accuracy, RMSE 1.697) and OrthologsBased methods (78.9% accuracy, RMSE 0.395).
• Expression and phenotype correlation: Results were correlated with blaKPC expression by reverse transcription-qPCR (P < 0.001) and associated with MICs for imipenem and ceftazidime-avibactam (P < 0.001) in 357 KPC-producing Klebsiella pneumoniae isolates.

Scientific Applications:

• Gene copy-number quantification: Quantify carbapenemase gene copy numbers in Gram-negative bacteria such as Klebsiella pneumoniae from WGS data.
• Genotype–expression studies: Correlate estimated gene copy numbers with transcript levels measured by reverse transcription-qPCR.
• Genotype–phenotype association: Associate bla gene copy number estimates with antimicrobial susceptibility phenotypes (MICs) for agents including imipenem and ceftazidime-avibactam.
• Method benchmarking: Serve as a benchmark for comparing copy-number estimation methods using simulation and qPCR validation.

Methodology:

Maps whole-genome sequencing reads to AMR and housekeeping reference loci, normalizes AMR read counts to housekeeping gene counts to infer copy numbers, and implements two modules (CCNE-acc and CCNE-fast) for high-accuracy and rapid screening workflows.