Enterovirus D68-associated respiratory and neurological illness in Spain, 2014–2018

Figures & data

Table 1. Panel of enterovirus (EV), parechovirus (HPeV) and rhinoviruses (RV) types used for validation of the EV-D68 RT-nested PCR assay.

Virus	EV-D68 RT-nested PCR
EV-A: CV-A6, CV-A16, CV-A10, EV-A71	negative
EV-B: E-5, E-6, E-9, E-11, E-13, E-20, E-30, CV-B3, CV-B4	negative
EV-C: CV-A24, PV1SL	negative
EV-D68 isolates:
SP_AGO10_50323_R	positive
SP_SEP10_50324_R	positive
SP_JUN11_50346_R	positive
SP_OCT10_50439_R	positive
RV-A, RV-C	negative
HPeV-1, HPeV-3	negative

Figure 1. Enterovirus genotypes detected in the respiratory specimens included in this study.

Figure 2. Seasonal distribution of EV-A, EV-B and EV-D68 infections in respiratory syndromes over the 5-year study period.

Table 2. Comparison of the clinical symptoms between EV-D68- and EV-A or EV-B.

Respiratory symptoms	EV-D68-infected patients (N = 172)	EV-A or B-infected patients (N = 174)	p value
Bronchospasm/asthma	17 (9.9%)	10 (5.7%)	0.1587
Bronchiolitis	17 (9.9%)	17 (9.8%)	0.9720
Bronchitis	38 (22.1%)	11 (6.3%)	0.0000
Pneumonia	36 (20.9%)	35 (20.1%)	0.8524
Acute respiratory distress	38 (22.1%)	27 (15.5%)	0.1205
Upper respiratory tract infections	20 (11.6%)	62 (35.6%)	0.0000
Pharyngitis/tonsillitis	6 (3.5%)	12 (6.9%)	0.1638

Figure 3. Phylogenetic analysis of 290 EV-D68 sequences in 3´-VP1 region (445 bp) from Spain, and representatives of different clades worldwide. MEGA 7.0 software was used to construct the neighbour joining and maximum composite likelihood tree. Only bootstrap values > 70% are shown. EV-D68 prototype strain Fermon was used as the outgroup. Isolates are indicated by the GenBank accession number, country and year of isolation. Spanish strains from 2014 to 2018 are represented by circles and indicated by country abbreviation, month and year of isolation and clinical manifestation. White circles represent respiratory-associated sequences and black circles represent neurological-associated sequences. Abbreviations for clinical manifestations are as follows: R-Respiratory disease, M-Meningitis, ME-Meningoencephalitis AFP-Acute flaccid paralysis, F-Fever.