INTRODUCTION: Noting that the activated eosinophils and mast cells (MCs) that accumulate in the esophagus of eosinophilic esophagitis (EoE) patients secrete myoactive, pro-inflammatory, and cytotoxic products capable of causing the motor abnormalities and neuronal degeneration of achalasia, we proposed that an allergy-mediated, esophageal muscle-predominant form of EoE might cause achalasia. In earlier studies exploring that proposal, we found that LES muscle of achalasia patients exhibits striking MC degranulation. Although the hypercontraction and poor relaxation that characterize achalasic LES muscle have been attributed solely to its loss of inhibitory neurons, we hypothesized that abnormalities intrinsic to LES muscle cells themselves might contribute to motor disturbances in achalasia associated with MC degranulation. This study aimed to explore genetic factors that might underlie such intrinsic abnormalities. METHODS: We performed qPCR for a panel of genes known to mediate smooth muscle contraction and Ca2+ handling on LES muscle specimens obtained from 7 achalasia patients (2 type I, 4 type II, 1 type III) and 3 EGJ outflow obstruction (EGJOO) patients during Heller myotomy; control LES muscle was taken from 2 organ donors. RESULTS: Hierarchical clustering analysis revealed two discrete clusters that we call “mototype” gene patterns; one control subject grouped within each cluster (Figure 1). Mototype Cluster 1, comprised of the 2 type I and 1 type III achalasia patients as well as 2 EGJOO patients, exhibited upregulation of genes involved in smooth muscle contraction and Ca2+ handling (gene expression relative to corresponding control, Figure 2). In contrast, Mototype Cluster 2, comprised of all 4 achalasia type II patients and 1 EGJOO patient, exhibited downregulation of those same genes. Principal component analysis recapitulated this clustering of mototypes that separated achalasia types I and III from type II (Figure 3), and Rand Index calculations demonstrated that this clustering was not due to chance. CONCLUSION: We show that the genetic mototype of LES muscle can distinguish the manometric phenotypes of achalasia that are associated with MC degranulation. These findings support our proposal that there is an allergy-mediated form of achalasia, and suggest that when patients acquire this form of achalasia, their underlying genetic mototype might contribute to the resulting manometric phenotype.Figure 1.: Heat-map and hierarchical clustering with dendrogram generated from qPCR gene expression data of LES muscle for 2 control subjects, 7 achalasia patients (2 type I, 4 type II, 1 type III) and 3 EGJ outflow obstruction (EGJOO) patients reveals clustering of achalasia types I and III in mototype 1, and achalasia type II into mototype 2. Red color corresponds to high relative expression and blue color corresponds to low relative expression.Figure 2.: Heat-map and hierarchical clustering with dendrogram generated from qPCR gene expression data of LES muscle from 7 achalasia patients and 3 EGJOO patients relative to their corresponding control highlights differences between the two mototype clusters. Note the relative upregulation of the calcium handling genes PLN, RYR3, IP3R1 and the smooth muscle contraction and contractility genes RHOA, COL1A2, MYLK, and ACTA2 in mototype 2 patients and the relative downregulation of these same genes in mototype 1 patients. Red/blue colors indicate up/down-regulation with respect to corresponding control in each mototype.Figure 3.: Principal component (PC) analysis of gene expression data confirms mototype clustering of achalasia types 1 and III distinct from achalasia type II.