Here, we analyzed bacteria colonizing the mucosal tissue in Japanese people with a normal esophagus, reflux esophagitis, and Barrett’s esophagus. Our study showed no significant difference in the total amount of bacterial DNA between the three groups (p > 0.1), but unexpected diversity in bacterial populations on the esophageal epithelia in these subjects.
The evolutionary idea that natural selection generally favors the good of the species is currently accepted . This evolutionary history suggests that the bacterial population of the esophagus should remain relatively invariable. Pei Z et al. have reported a complex but conserved bacterial population in the normal distal esophagus . Generally, the relationship between bacteria and the normal esophagus must be balanced, and disruption of this homeostasis may result in esophageal diseases, or esophageal diseases may cause the change in bacterial biota. This prompted us to investigate the relationship between the bacterial biota and esophageal diseases. Our findings suggest the presence of highly complex bacterial populations in the distal esophagus in Japanese subjects, no matter whether they have a normal esophagus, reflux esophagitis, or Barrett’s esophagus. We also found that esophageal bacterial composition differs between these groups. Traditionally, the human esophagus has not been viewed as a hospitable environment for microorganisms because of anatomical structure, and culture-based approaches that have been used to analyze microbiota showed that the esophagus was either sterile or contained only a few transient bacteria [20, 21]. However, conventional culturing techniques cannot detect all the bacteria in the gut because of the requirements of an anaerobic and complex environment . The recent use of molecular methods and, in particular, genetic sequencing have revealed a much more diversified flora, of which some are cultivable by traditional techniques whereas many others are not . Pei Z et al. studied bacterial biota in the human distal esophagus using a phylogenetic approach based on 16S rDNA sequences . In the present study, we analyzed bacterial 16S rDNA sequences using a standard nucleotide BLAST search of GenBank for homology with known bacterial 16S rDNA sequences.
Regarding the amount of bacteria in the distal esophagus, we used a real-time PCR method to amplify a 466-bp fragment of the bacterial 16S rDNA gene. Results demonstrated the presence of numerous bacteria in the esophagus, in contrast to previous reports using culture-based approaches [20, 21, 24]. Interestingly, our study showed no significant difference in the amount of bacteria between groups with a normal esophagus, reflux esophagitis, or Barrett’s esophagus (p > 0.1).
Phylum-level analysis of our present samples revealed that the bacterial communities differed among groups. First, each group had a different number of phyla: populations could be classified into four phyla in patients with normal esophagus, six in those with reflux esophagitis, and five in those with Barrett’s esophagus. Second, phyla composition differed among groups. For example, Fusobacteria was found in patients with reflux esophagitis or Barrett’s esophagus but not in those with a normal esophagus. We found differences in composition at the genus level among the normal esophagus, reflux esophagitis, and Barrett’s esophagus groups. We compared not only the distribution of bacterium 16S rDNA gene clone libraries but also the bacteria-positive patient numbers. The most prevalent genus was Streptococcus in patients with a normal esophagus or reflux esophagitis, versus Veillonella in patients with Barrett’s esophagus. Among other genera, we found that Fusobacterium was not detected in normal esophagus but was detected at 9% in both reflux esophagitis and Barrett’s esophagus. A number of other disease-associated differences in genera were noted (Figure 2). These findings therefore show that bacterial populations differ among normal subjects and patients with reflux esophagitis or Barrett’s esophagus at both the phylum and genus levels.
According to an estimate by Ashktrab et al. in African Americans , the H. pylori positivity was much smaller in patients with esophagitis (4%) than in normal controls (34%). The prevalence of Helicobacter pylori (H. pylori) infection varies widely by geographic area, age, race and socioeconomic status . Generally, H. pylori is a common stomach bacteria among Asian populations . It has been suggested that Asians could be protected against GERD by their high prevalence of H. pylori infection . The declining prevalence of H. pylori infection due to improved hygiene conditions or use of H. pylori eradication therapy might have contributed to the recent increased frequency of reflux esophagitis in Asia . Here, we found Helicobacter in four of six subjects with a normal esophagus, two of six with reflux esophagitis, and three of six with Barrett’s esophagus. Allowing for the small number of subjects in each group, we consider that these findings may support the idea that a high prevalence of H. pylori infection protects against reflux esophagitis.
Several limitations of our study warrant mention. First, we evaluated bacterial microbiota in the distal esophagus in patients with gastro-esophageal reflux disease (GERD), Barrett’s esophagus, and normal esophageal mucosa using a small data set. Second, the normal subjects were younger than the reflux esophagitis and Barrett’s esophagus patients, which may have affected bacterial variability. Third, we defined species as homologous when their 16S rDNA sequences had >97% identity with known bacterial species. However, comparative studies have clearly revealed the limitations of sequence analysis of this conserved gene and gene product in the determination of relationships at the strain level, for which DNA-DNA reassociation experiments still constitute the superior method . Accordingly, additional study with more refined data collection and an increased number of normal older subjects is required.