Diagnostic moléculaire des infections respiratoires à entérovirus : apport de la PCR et du génotypage pour une meilleure approche de la circulation des souches en Basse-Normandie au cours de l’année 2008

The PCR assays are currently used in diagnosis of enterovirus (EV) meningitis. Nevertheless, the use of molecular diagnosis of EV should be investigated in respiratory tract infections (RTI).

Objectives

To perform enterovirus molecular diagnostic tools, PCR and genotyping, in nasal samples for diagnostic and epidemiologic purposes.

Methods

During 2008, 3612 nasal specimen (NS) were studied by IFD and MRC5 culture. Next, we realised successively viral isolation on HuH7 culture (for NS negative by IFD assay) and a duplex PCR enterovirus-rhinovirus for the 816 HuH7 positive supernatants. Furthermore, 327 NS collected from neonates were systematically tested by a real-time RT-PCR. This assay was used in routine for EV diagnosis setting in cerebrospinal fluid. Enterovirus genotyping was then performed for the 68 positive supernatants.

Results

Thirty-five NS (0.97%) were positive for EV by culture (MRC5). A combination of both PCR assays, PEVRV and PEV, allowed an additional identification of 41 EV, eight EV-RV and 12 RV, increasing the number of positive to 96 NS (2.6%). Among the neonates, 32 NS (11.3%) were positive for EV by PEV. Of the 98 NS tested by the two PCR assays (PEV and PEVRV), 27 were positive and we detected 10 EV, five EV-RV and 12 RV. From January to December 2008, the circulation of EV showed the usual peak in June-July when a small outbreak of aseptic meningitis occurred and an additional autumnal peak corresponding to respiratory tract infections. Five main serotypes were isolated: 19 EV68 (29.7%), 12 CB3 (18.7%), nine E3 (14,1%), six CA9 (9.4%) and six CB1 (9.4%); the 19 EV68 were isolated in October-November and 17/19 (89.5%) of positive patients were hospitalised for severe respiratory diseases.

Conclusion

The use of molecular screening techniques (PCR assays and genotyping) on nasal samples collected from patients with respiratory infections allowed a prospective, effective and precise identification of circulating strains.