Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2021 Jun;1(1-2):10.1016/j.jcvp.2021.100018.
doi: 10.1016/j.jcvp.2021.100018.

Detection of coxsackievirus A6 in formalin-fixed, paraffin-embedded skin biopsy specimens using immunohistochemistry and real-time reverse-transcriptase PCR

Amy M Denison  1 , Julu Bhatnagar  1 , Richard R Jahan-Tigh  2 , Pamela Fair  1 , Gillian L Hale  1
Affiliations

Detection of coxsackievirus A6 in formalin-fixed, paraffin-embedded skin biopsy specimens using immunohistochemistry and real-time reverse-transcriptase PCR

Amy M Denison et al. J Clin Virol Plus. 2021 Jun.

Abstract

Background: Hand, foot, and mouth disease (HFMD), classically a childhood viral infection, has an atypical and severe clinical presentation in adults. Coxsackievirus A6 is a leading cause of atypical HFMD, but current diagnostic methods utilizing formalin-fixed, paraffin-embedded skin biopsy specimens often lack sensitivity and specificity.

Methods: Formalin-fixed, paraffin-embedded skin biopsies from seven case patients with clinical and histopathological suspicion of atypical HFMD were evaluated by coxsackievirus A6 (CVA6) immunohistochemistry, enterovirus-specific conventional reverse transcriptase-PCR with subsequent Sanger sequencing targeting the 5'UTR, and CVA6-specific real-time PCR targeting the VP1 gene.

Results: The CVA6-specific antibody demonstrated appropriate antigen distribution and staining intensity in keratinocytes in all cases. Conventional RT-PCR and sequencing also detected the presence of enterovirus, and CVA6-specific real-time RT-PCR analysis identified CVA6.

Conclusion: Applying these immunohistochemistry and molecular techniques to formalin-fixed, paraffin-embedded tissues, CVA6 was determined to be the causative infectious agent in seven cases of atypical hand, foot, and mouth disease.

Keywords: Atypical HFMD; Coxsackievirus A6 (CVA6) real-time; Enterovirus; Formalin-fixed; Hand, foot, and mouth disease (HFMD); Immunohistochemistry (IHC); Paraffin-embedded (FFPE) tissue; reaction (rRT-PCR); reverse-transcriptase polymerase chain.

PubMed Disclaimer

Conflict of interest statement

Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Figures

Fig. 1.
Fig. 1.
Hematoxylin and eosin (H&E) staining (A&B) and immunohistochemical (IHC) staining (C&D) as Exemplified by Case Patient Number Seven. A. H&E 10X. The shave biopsy of the right dorsal index finger joint shows intraepidermal vesiculation with associated neutrophilic inflammation, papillary edema and a lymphocytic dermatitis. B. H&E 20X. Higher power demonstrates epidermal necrosis in the area of vesiculation (arrows), intralesional neutrophils and edema. C. IHC 10X. CVA6 immunostain using a red chromogen highlights the extensive intracytoplasmic viral antigens throughout the lesion. D. IHC 20X. A higher power image of the CVA6 immunostain shows extensive cytoplasmic staining of the infected cells in the area of vesiculation (arrows).
Fig. 2.
Fig. 2.
5’ to 3’ nucleotide sequence of the portion of the EV VP1 gene in which the forward and reverse primers and probe anneal (primer/probe sequences and orientation in gray arrows) from cases 1 and 3 as well as the Gdula strain. Nucleotide mismatches are also shaded in gray, demonstrating the larger number of divergent nucleotides present in the Gdula strain.
See this image and copyright information in PMC

References

    1. Long L, Xu L, Xiao Z, Hu S, Luo R, et al. Neurological complications and risk factors of cardiopulmonary failure of EV-A71-related hand, foot and mouth disease, Sci. Rep 6 (2016) 23444. - PMC - PubMed
    1. Feder HM Jr., Bennett N, Modlin JF, Atypical hand, foot, and mouth disease: a vesiculobullous eruption caused by Coxsackie virus A6, Lancet Infect. Dis 14 (2014) 83–86. - PubMed
    1. Kimmis BD, Downing C, Tyring S, Hand-foot-and-mouth disease caused by coxsackievirus A6 on the rise, Cutis 102 (2018) 353–356. - PubMed
    1. Esposito S, Principi N, Hand, foot and mouth disease: current knowledge on clinical manifestations, epidemiology, aetiology and prevention, Eur. J. Clin. Microbiol. Infect. Dis 37 (2018) 391–398. - PubMed
    1. Laga AC, Shroba SM, Hanna J, Atypical hand, foot and mouth disease in adults associated with coxsackievirus A6: a clinico-pathologic study, J. Cutan. Pathol 43 (2016) 940–945. - PubMed

LinkOut - more resources