Journal Article

398 Identification of Varicella Zoster by Novel Salivary and Esophageal Muscle Tissue Assay in Patients With Achalasia: A Two Center Prospective Study

Holzman, Mike, Gershon, Anne, Chen, Jason, Naik, Rishi, Higginbotham, Tina, Flores, Elizabeth, Vaezi, Michael, Gershon, Michael

2019

Overview

INTRODUCTION:Achalasia is an esophageal motor disorder characterized by impaired relaxation of the lower esophageal sphincter (LES) but the etiology of this neuronal damage is unknown. Varicella zoster virus (VZV) is an exclusively human neurotrophic alpha herpes virus that becomes latent in ganglionic neurons after initial infection. We tested the hypothesis that reactivation of latent VZV within esophageal enteric neurons initiates an inflammatory cascade that damages the innervation of the smooth muscle of the LES giving rise to achalasia. We aimed to determine whether VZV DNA, transcripts, and proteins are present in tissue and saliva of patients with achalasia and controls.METHODS:Fifteen patients with achalasia (60% Male, average age 60 years and BMI 28 kg/m2) diagnosed with high resolution manometry (7/15 with Type II Achalasia) who underwent surgical myotomy had prospective collection of saliva and LES muscle tissue with IRB approval. Salivary samples were collected by passive drool, DNA was extracted with the DNeasy Blood and Tissue Kit, and VZV DNA was amplified with PCR. All methods examined the presence of transcripts (cDNA) and DNA encoding VZV open reading frames (ORFs) 29, 40, and 67. Immunofluorescence (IMF) was used to demonstrate the neuronal markers, beta-3 tubulin or peripherin simultaneously with the VZV late protein, gE. Antigen retrieval at 100°C in citrate buffer (pH 6.0) was employed.RESULTS:VZV DNA was detected in the saliva in 12/15 (80%) of achalasia cases (Table 1). Transcripts encoding at least one ORF were found in esophageal myotomy samples 12/15 (80%), while DNA was detected in 7/15 (47%). All 15 patients had active VZV infection by salivary VZV DNA and/or VZV transcription in esophageal tissue (Figure 1). Detection of transcripts was more sensitive and specific indicator of tissue infection than DNA. IMF revealed the presence of gE within the cell bodies of enteric neurons (when present) (Figure 2). VZV DNA was not detected in the saliva of 20 controls.CONCLUSION:Direct ex-vivo examination of saliva and tissue from patients with achalasia revealed the presence of active VZV infection of the esophagus. Transcripts encoding VZV gene products, and DNA, support actively replicating virus, not just latent VZV. Immunocytochemistry suggested that enteric neurons were the site of VZV infection. These findings support VZV is a potential esophageal pathogen that reactivates from latency in enteric neurons to give rise to achalasia, which may be a therapeutic target.Table 1.Protein Detection by Saliva and Esophageal Muscle Tissue Assay

• Publisher: Wolters Kluwer Health Medical Research, Lippincott Williams & Wilkins
• Subject: Chicken pox / Esophagus / Infections