mNGS Signal Calculator

1 Sample Information ▾

Sample Type

WBC (×10⁹/L) 6.0

Normal Infection Sepsis

Sample Volume (mL) 1.0

2 Pathogen ▾

E. coli S. aureus K. pneumoniae MTB HBV C. albicans CMV

Pathogen Name

Concentration 1000

copies/mL

Note: Model assumes 1 CFU ≈ 1 Genome Copy for simplicity.

Genome Size (Mb) 4.6

Use Unique Reads
(recommended for viruses)

3 Sequencing ▾

Total Reads (M) 10

2M 10M 20M 50M

4 DNA Extraction (Advanced) ▾

Recovery (%) 50

Overall DNA extraction yield; commercial kits typically 30-70%

Bias 1.0

Cell wall structure affects extraction efficiency: G- = 1.0, G+ ≈ 0.6, fungi ≈ 0.3, viruses ≈ 1.2

5 Host Depletion (Advanced) ▾

Enable Host Depletion

Depletion Rate (%) 99

95% 99% 99.9%

Host DNA removal rate; saponin ~95-99%, CRISPR up to 99.9%

Microbe Loss (%) 50

50% 90% 99%

Co-loss of pathogen DNA during host depletion; depends on method selectivity

6 Library & Analysis (Advanced) ▾

Library Efficiency (%) 80

Extracted DNA converted to sequenceable library; PCR-free 60-80%, PCR-based 30-60%

Analysis Recall (%) 85

Bioinformatics pipeline correctly identifies and retains pathogen reads; Recall = TP/(TP+FN)

Duplicate Rate (%) 5

Low (5%) Med (30%)

Physical PCR duplicates; high depth or low input leads to higher duplication

Specificity Ratio (%) 100

Percentage of reads that are uniquely mapped to this specific species

Database Coverage (%) 95

Reference genome database completeness for the target pathogen

Background Noise (RPM) 0.5

Baseline signal from reagent contamination, environmental microbes, and cross-contamination

Gross Observation 0.00

- Background Noise 0.00

Estimated Clinical Load

0.0000

reads per million

S/N Ratio: 0.0x

Low Marginal Reliable

Total Mapped

De-duplicated

Unique Reads

Key Insight

Calculating...

Calculation Steps

Expand

Human Cells in Sample

human_cells = WBC × 10⁹ × volume(mL) / 1000

Pathogen DNA Total

pathogen_DNA = CFU × volume × genome_size

Human DNA Total

human_DNA = cells × 6.5 pg × 978 Mb/pg ≈ 6357 Mb

After Host Depletion

human_DNA' = human_DNA × (1 - depletion_rate)

Pathogen DNA After Extraction

pathogen_DNA'' = pathogen_DNA × recovery × bias

Human DNA After Extraction

human_DNA'' = human_DNA' × recovery

Library Prep Ratio

ratio = pathogen_DNA'' / (pathogen_DNA'' + human_DNA'') × library_eff

Analysis & RPM

RPM = ratio × Recall × db_coverage × 10⁶ - bg_noise

Virus Mode: Unique Reads

Unique Reads = Reads_mapped × (1 - Duplicate_rate) × Specificity_ratio

Parameter Impact Analysis

Seq Depth

Recovery

Depletion

Recall

Library

Reads Distribution

LOD Analysis

Laboratory Evidence Strength

Reliable
≥10 RPM

Marginal
1-10 RPM

Undetectable
<1 RPM

Core Calculation Formulas ▾

# 1. Human cells from WBC

human_cells = WBC × 10⁹ × volume_mL / 1000

# 2. Pathogen DNA total

pathogen_DNA = CFU × volume × genome_size (Mb)

# 3. Human DNA total

human_DNA = human_cells × 6.5 pg × 978 Mb/pg (≈ 6357 Mb)

# 4. After host depletion

human_DNA' = human_DNA × (1 - D)

# D = depletion rate

# 5. Pathogen DNA after extraction

pathogen_DNA'' = pathogen_DNA × R × B

# R = recovery, B = bias factor

# 6. Human DNA after extraction

human_DNA'' = human_DNA' × R

# 7. Library prep ratio

ratio = pathogen_DNA'' / (pathogen_DNA'' + human_DNA'') × L

# L = library efficiency

# 8. Analysis & RPM

RPM = ratio × Recall × C_db × 10⁶ - N_bg

# Recall = analysis recall, C_db = DB coverage, N_bg = background noise

# 9. Virus mode: Unique Reads

Unique Reads = (RPM × total_reads / 10⁶) × U_ratio

# U_ratio = unique reads ratio (background noise already deducted)

Extraction Bias Reference ▾

Pathogen Type	Bias	Note
G- (E. coli)	1.0	Standard
G+ (S. aureus)	0.5-0.8	Thick cell wall
Fungi (Candida)	0.2-0.5	Chitin cell wall
Virus (non-enveloped)	1.2-1.5	Easy lysis
Mycobacteria	0.1-0.3	Waxy cell wall