Figure 1.
Phylogenetic tree of the orthomyxoviridae family (PB2 gene).
A sub-tree for the Isavirus genus is observed in the lower right corner. The tree was constructed with reference sequences from GenBank database (Accession numbers; Influenza A: NC_004910, HQ117883, AJ40430; Influenza B: NC_002204, AF101982, EF626642; Influenza C: NC_006307, M28061, AF170576; ISAV: NC_006505, AY373381, DQ785183, EU118815, GU830895; Quaranjavirus: GQ499302; Thogotovirus: Y17783). The tree was inferred by using the Maximum Likelihood method based on the Tamura-Nei model with a bootstrap test (1000 replicates) using MEGA 5 software [33].
Table 1.
qRT-PCR analysis for genomic segment 8 based on Snow et al. [23].
Table 2.
Summary of sequencing results for genomic segments 6 and 5.
Figure 2.
High Resolution Melting Profiles for segment 5.
The lower right corner shows the standard reference variants, with No Insert: red line, and with Insert 4: blue line. The analyzed samples all matched with the No Insert variant.
Figure 3.
High Resolution Melting Profiles for segment 6.
The lower right corner shows the standard reference variants; HPR0: red line, and HPR7b: blue line. Two samples, GIM-1 and GIM-3 matched the HPR0 variant profile; another two, GIM-4 and GIM-5 matched the HPR7b variant profile; GIM-2 sample does not match any of the two profiles. This behavior is coherent with sequencing results, in which GIM-2 showed an overlapping sequence of the two variants (HPR0 and HPR7b).
Table 3.
Summary of HRM results for field samples.
Table 4.
Sequencing results DGGE bands.
Figure 4.
DGGE band profile for segment 5.
Lanes 1–5 corresponds to samples GIM-1 to GIM-5. The sequencing results for each band are summarized in Table 4.
Figure 5.
DGGE band profile for segment 6.
Lanes 1–5 corresponds to samples GIM-1 to GIM-5. The sequencing results for each band are summarized in Table 4.
Figure 6.
Diagram of the PCR-based experimental methodology.
Figure 7.
Target for PCR amplification on segment 5 as template.
Insert 4 and amino acid in position 266 are indicated. Arrows indicate primer locations.
Figure 8.
Target for PCR amplification on segment 6 as template.
HPR region is indicated. Arrows indicate primer locations.