With advancements in the field of digital pathology, there has been a growing need to compare the diagnostic abilities of pathologists using digitized whole slide images (WSI) against using traditional hematoxylin and eosin (H&E)-stained glass slides for primary diagnosis. One of the most common specimens received in pathology practices is an endoscopic gastric biopsy with a request to rule out Helicobacter pylori (H. pylori) infection. The current standard of care is the identification of the organisms on H&E-stained slides. Immunohistochemical or histochemical stains are used selectively. However, due to their small size (2-4 μm in length by 0.5-1 μm in width), visualization of the organisms can present a diagnostic challenge. The goal of the study is to compare the ability of pathologists to identify H. pylori on H&E slides using a digital platform against the gold standard of H&E glass slides using routine light microscopy. Diagnostic accuracy rates using glass slide vs. digital slide were 81% vs. 72% (p=0.0142) based on H&E slides alone. When H. pylori IHC slides were provided, the diagnostic accuracy was significantly improved to comparable rates (96% glass vs. 99% digital, p=0.2199). Furthermore, differences in practice settings (academic/subspecialized vs. community/general) and the duration of sign-out experience did not significantly impact the accuracy of detecting H. pylori on digital slides. We concluded that digital WSI, although amenable in different practice settings and teaching environments, it does present some shortcomings in accuracy and precision, especially in certain circumstances and thus is not yet fully capable of completely replacing glass slide review for identification of H. pylori. We specifically recommend reviewing glass slides and/or performing ancillary stains, especially when there is a discrepancy between the degree of inflammation and the presence of microorganisms on digital image.