Hand, Foot, and Mouth Disease Caused by Coxsackievirus A6: A Preliminary Report from Istanbul

Abstract

Hand, foot, and mouth disease (HFMD) is caused by various serotypes of Enterovirus genus. Coxsackievirus A16 (CV-A16) and enterovirus A71 (EV-A71) were known to be the only responsible agents for these epidemics; however, this opinion was challenged after the detection that coxsackievirus A6 (CV-A6) was the responsible species for the outbreak in Finland in 2008. HFMD is frequently seen in Turkey, and no detailed study on its clinical and microbiological epidemiology has previously been reported. The present study addresses this question. Twenty-seven patient samples collected between 2015 and 2017 were included in the study. Typing was conducted by RT-PCR and the sequencing applied directly to patient's samples and as well as to the viral cultures with pan-enterovirus and serotype-specific primers. The presence of Enterovirus in 12 of 27 HFMD samples was shown with RT-PCR. The causative agent for three of these 12 samples was CV-A16, one of the most frequent two serotypes around the world, and the remaining nine samples was CV-A6. The findings of the study are relevant since it pertains to the molecular epidemiology of HFMD in Turkey, a gateway country where different serotypes might be circulating and transmitted. The findings also support the notion that CV-A6 cases are rising in number, which has caused more severe clinical features and widespread rashes in recent outbreaks.

Hand, foot, and mouth disease (HFMD) is caused by various serotypes of Enterovirus genus. Coxsackievirus A16 (CV-A16) and enterovirus A71 (EV-A71) were known to be the only responsible agents for these epidemics; however, this opinion was challenged after the detection that coxsackievirus A6 (CV-A6) was the responsible species for the outbreak in Finland in 2008. HFMD is frequently seen in Turkey, and no detailed study on its clinical and microbiological epidemiology has previously been reported. The present study addresses this question. Twenty-seven patient samples collected between 2015 and 2017 were included in the study. Typing was conducted by RT-PCR and the sequencing applied directly to patient’s samples and as well as to the viral cultures with pan-enterovirus and serotype-specific primers. The presence of Enterovirus in 12 of 27 HFMD samples was shown with RT-PCR. The causative agent for three of these 12 samples was CV-A16, one of the most frequent two serotypes around the world, and the remaining nine samples was CV-A6. The findings of the study are relevant since it pertains to the molecular epidemiology of HFMD in Turkey, a gateway country where different serotypes might be circulating and transmitted. The findings also support the notion that CV-A6 cases are rising in number, which has caused more severe clinical features and widespread rashes in recent outbreaks.

Fig. 1.

Patients with typical lesions on the hands.

Fig. 2.

Patients with mouth ulcers.

Fig. 3.

Patient with generalized lesions on the legs.

Fig. 4.

The samples were subjected to the pan-enterovirus specific nested PCR with outer primers where 530 bp products were amplified. Left to right, line 1: 100 bp marker, lines 2 to 5: HFM-2 to HFM-5, line 6: HFM-7, line 7: HFM-9, lines 8 to 14: HFM-10 to HFM-16.

Fig. 5.

The samples were subjected to the pan-enterovirus specific nested PCR with inner primers where 389 bp products were amplified. Left to right, line 1: 100 bp marker, lines 2 to 5: HFM-2 to HFM-5, line 6: HFM-7, line 7: HFM-9, lines 8 to 14: HFM-10 to HFM-16.

Fig. 6.

CV-A6 specific primers amplified products of approximately 645 bp.

Fig. 7.

Phylogenetic analysis of clinical isolates. Phylogenetic analysis of coxsackievirus A6 nucleotide sequences that were obtained using 153F ve 541R primers, showed the relationships between the clinical CV-A6 and the CV-A16 isolate selected from the GenBank. A scale bar indicates branch distances. The phylogenetic tree was constructed using neighbour-joining method and validated with 1000 pseudo-replicates. Poliovirus (NC 002058.3) was selected as out-group.