TABLE 3.
Detection of single nucleotide polymorphisms in the NA of viruses recovered from lungs of mice (day 6 p.i. of passage 2) infected with the A/WSN/33 (H1N1) and A/Victoria/3/75 (H3N2) viruses by automated (Sanger) sequencing, colony screening (cloning), and deep sequencinga
| Virus/regimenb | NA positionc | Referenced | SNP detected by: |
Depthe | ||
|---|---|---|---|---|---|---|
| Sanger sequencing (%) | Cloning (%) | Deep sequencing (%) | ||||
| A/WSN/33 (H1N1) | ||||||
| P2-ose/P1-ose 0.05 mg/kg | 389 | M | M | M/T (95/5) | M/T (99/1) | 124,898 |
| 434 | D | D | D/E (95/5) | D/Ef (98/2) | 77,999 | |
| A/Victoria/3/75 (H3N2) | ||||||
| P2-unt/P1-unt | 46 | A | A | A/P (94/6) | A/P (94/6) | 57,911 |
| 73 | I | I | I/T (94/6) | I/T (96/4) | 72,738 | |
| 131 | Q | Q | Q/H (88/12) | Q/H (96/4) | 78,897 | |
| 398 | V | V | V/I (94/6) | V/I (93/7) | 66,338 | |
| P2-ose/P1-ose 0.05 mg/kg | 200 | N | N | N/D (89/11) | N/D (88/12) | 81,069 |
| 222 | I | I | I/T (84/16) | I/T (85/15) | 78,541 | |
| 398 | V | V | V/I (95/5) | V/I (87/13) | 56,232 | |
| P2-ose/P1-ose 1 mg/kg | 131 | Q | Q | Q/H (94/6) | Q/H (98/2) | 128,656 |
| 222 | I | I/T (80/20) | I/T (78/22) | I/T (72/28) | 167,234 | |
| 398 | V | V | V/I (94/6) | V/I (95/5) | 104,690 | |
Data correspond to viruses recovered from pooled lung homogenates on day 6 p.i. of the second passage (P2). Only treatment groups in which single nucleotide polymorphisms (SNPs) were detected are shown (deletions are not shown). The residues in bold type confer reduced susceptibility to oseltamivir.
P1, passage 1; P2, passage 2; unt, untreated; ose, oseltamivir.
N2 numbering.
Reference; the predominant residues found in these positions in the reference sequences.
Mean number of reads that cover that position in the deep-sequencing analysis.
A D/V (93/7) substitution was also found at this position by deep sequencing but not by cloning in the P2-ose/P1-ose 1 mg/kg group.