Shiotani A; Matsumoto H; Fukushima S; Katsumata R; Kawano M; Saito M

Review Article

J Bacteriol Mycol. 2018; 5(8): 1086.

H. pylori and Human Gut Microbiota

Shiotani A¹*, Matsumoto H¹, Fukushima S¹, Katsumata R¹, Kawano M² and Saito M²

¹Department of Internal Medicine, Kawasaki Medical School, Japan

²Department of Microbiology, Kawasaki Medical School, Japan

*Corresponding author: Shiotani A, Department of Internal Medicine, Kawasaki Medical School, Private University in Kurashiki, Japan

Received: October 30, 2018; Accepted: November 26, 2018; Published: December 03, 2018

Abstract

The composition of the Gastrointestinal (GI) microbiome is shaped by a variety of factors including diet, additional environmental elements, and the genetic background of the host.

Recent evidence supports that H. pylori is the most relevant, but may not be the only local causative bacteria leading to gastric diseases. In several recent reports, a real, active cross talk between H. pylori and the other components of the gastric microbiota was observed. In this review, we provide a comprehensive review about gastric microbiota and discuss emerging concepts for the influence of H. pylori infection on gastric microbiome and vice versa.

H. pylori Infection and Gastric Microbiota

GI microbiota had been investigated by cultivation of luminal contents or mucosal biopsies, and the human stomach was long thought to be sterile. The new nucleotide sequencing techniques and advanced bioinformatics tools have opened the field for studying the diversity and complexity of the GI microbiome independent of traditional cultural methods. Increasing number of papers focus on non-H. pylori microbial community observed in the human stomach, so called human gastric microbiota [1]. Recent evidence supports that H. pylori is the most relevant, but may not be the only local causative bacteria leading to gastric diseases. The discrepancies of the results in the previous studies are likely due to different methods used for microbiota analysis, the limited sample sizes, difference in H. pylori infection rates, and several environmental factors such as diet, lifestyle, geography, and ethnicity. In several recent reports, a real, active cross talk between H. pylori and the other components of the gastric microbiota was observed. In this review, we provide a comprehensive review about gastric microbiota and discuss emerging concepts for the influence of H. pylori infection on gastric microbiome and vice versa.

Human stomach had been thought to be sterile, and following H. pylori discovery, it was thought to be the only bacterium able to colonize the gastric epithelium. Gastric environment is difficult to colonize mainly because of gastric acid barrier. Therefore, the microbial load is much less in the stomach than in the colon (1010- 1012 Colony-Forming Units (CFU)/mL) or been small intestine (102-104 CFU/mL) [2]. Gastric microbiota has previously identified by cultivation of gastric juice or mucosa biopsies. In the healthy stomach, the predominant bacteria belong to the species Clostridium, Lactobacillus and Veillonella based on culture analysis [3]. In the atrophic gastritis with the absence of H. pylori, urease producing members of the gastric microbiota, such as Proteus mirabilis, Klebsiella pneumonia, Staphylococcus aureus, Staphylococcus capitis, and Micrococcus species can cause false positive results in urea breath tests [4].

Recent advance of molecular biology, computer technology and bioinformatics allow genetic analysis of complex microbiota without cultivation, and the 16S rDNA sequence has opened a field for extensively revealing novel and uncultivated bacterial species in human stomach [5,6]. In the healthy stomach, the predominant bacteria belong to the phyla of Actinobacteria, Bacteroidetes, Firmicutes, and Proteobacteria (which include H. pylori), and genus of Streptococcus [7-9]. Utilizing a newer technology, tagged 454 pyrosequencing, analysis of H. pylori-negative biopsy samples identified 262 phylotypes representing 13 phyla [10]. The composition of the gastric microbiome in H. pylori-negative individuals is highly diverse. Common phylotypes present in H. pylori-uninfected subjects include Streptococcus, Prevotella, and Gemella [7]. These findings lend further support to the gastric microbiota being highly diverse, despite significant variability in the microbial composition between individuals [7,8].

In contrast, among H. pylori-positive subjects, the microbiota is much more uniform and H. pylori represents the most abundant phylotype. H. pylori DNA accounts for >90% of all sequence reads in H. pylori-positive subjects [7], so it greatly reduces the overall diversity of the gastric microbiota. The most abundant phyla in H. pylori-colonized stomachs are Proteobacteria, Firmicutes, and Actinobacteria [10,11]. H. pylori-infected adults are likely to have higher abundances of Spirochaetes, Acidobacteria and non- Helicobacter Proteobacteria and relatively lower abundance of Actinobacteria, Bacteroidetes, Firmicutes compared to H. pyloriuninfected adults [10]. A study performed by Hu et al. [12] identified the non-H. pylori bacterial flora in gastric biopsy specimens taken from H. pylori positive patients using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDITOF MS). The major species were Streptococcus, Neisseria, Rothia, and Staphylococcus, which differed from previous reports of healthy volunteers. Klymiuk et al. [13] recently conducted a prospective, multicenter, clinical trial of 30 gastric biopsy samples including CagA-negative and CagA-positive H. pylori. In their study, the genera Actinomyces, Granulicatella, Veillonella, Fusobacterium, Neisseria, Helicobacter, Streptococcus, and Prevotella are significantly different between the H. pylori-positive and negative sample groups. However, there is no significant correlation of H. pylori phylogeographic population or carriage of the cag PAI with microbiota composition [13].

Interestingly, Khosravi et al. [14,15] isolated Streptoccus mitis (S. mitis) and Lactobucillus fermentum (L. fermentum) from human gastric tissue biopsies, and they have shown that metabolites released by co-culturing S. mitis and H. pylori induced H. pylori to transform into viable but non-culturable coccoidal form in vitro. Streptococci seem to survive and develop in an acidic gastric environment as an indigenous microbiota of the gastric mucosa, which may inhibit the colonization by H. pylori. In contrast, L. fermentum improved S. mitis survival via secreting diffusible factors [15].

Table 1: Human gastric microbiota studies (not including gastric cancer patients).

Authors. year
Sample/Method
Samples
Results

Bik, et al. 2006
[8]
Gastric biopsies 16S rDNA clone library
23 USA adults
12 HP-positive 11
HP-negative (7 contained HP rDNA)
HP seems not affect the gastric microbiome composition.

Andersson, et al.
2008 [7]
Gastric biopsies pyrosequencing of V6 of 16SrRNA
6 healthy Swedish
3 HP-positive
3 HP-negative
Higher diversity in HP-negative patients than HP-positive patients

Andersson, et al. 2008 [7]
Gastric biopsies 16S rRNA sequencing
45 Chinese
22 healthy
23 antral gastritis
All HP-negative
Significantly higher abundance of the Firmicutes phylum and the Streptococcus genus was observed in patients with antral gastritis.

Maldonado-Contreras, et al.
2011 [10]
Gastric biopsies PhyloChip
12 adult patients
10 Amerindians 2
immigrants from South Asia and Africa
8 HP-positive
Marked differences were detected in the structure of the gastric bacterial community according to HP status.

Hu et al.
2012 [12]
Gastric biopsies MALDI-TOF MS
103 Chinese patients with dyspeptic symptoms
103 HP-positive
High prevalence of the non-HP bacterial dominated by some species.

Klymiuk, et al
2016 [13]
Gastric biopsies 16S rRNA sequencing
30 Austrian
10HP Cag A-positive
10 HP-Cag A-negative
10 HP-negative
A dramatic decrease of non HP bacterial microbiome in HP infected samples and no significant influence of CagA gene presence.

Parsons, et al. 2017 [19]
Gastric biopsies 16S rRNA sequencing
95 British patients (normal stomach, PPI treated, HP gastritis, HP induced atrophic gastritis and autoimmune atrophic gastritis)
Autoimmune and HP induced atrophic gastritis were associated with different gastric microbial profiles. PPI treated patients showed relatively few alterations in the gastric microbiota compared to healthy subjects.

Pereira, et al.
2018 [30]
Gastric biopsies MALDI-TOF MS
74 HP-positive
21with chronic dyspepsia
53 without
Staphylococcus and Lactobacillus spp. were more commonly identified in patients with chronic dyspepsia.

Schulz, et al. 2018 [33]
�Saliva, gastric and duodenal aspirates and biopsies
16S rRNA sequencing
24 German with chronic gastritis
8 HP-positive
16 HP-negative
Helicobacter spp. dominate the mucosa-associated community in the stomach, and to signi?cantly in?uence duodenal and oral communities.

GC: Gastric Cancer; HP: Helicobacter Pylori; MALDI-TOF MS: Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry.



Table 1: Human gastric microbiota studies (not including gastric cancer patients).

Table 2: Human gastric microbiota studies (including gastric cancer patients).




  
    Authors. year 
    Sample/Method
    Samples
    Results
  
  
    Dicksved,    et al. 2009 [23]
    Gastric    biopsies T-RFLP 16S rRNA cloning and sequencing
    Swedish    10 GC (5 intestinal type, 5 diffuse type; 8 HP-positive) 5 HP-negative    dyspeptic controls
    Microbiota    in cancer patients was dominated by different species of Streptococcus,    Lactobacillus, Veillonella, and Prevotella.
  
  
    Eun,    et al. 2014 [24]
    Gastric    biopsies 454 pyrosequencing of V5 of 16S rRNA
    31    Korean patients (11 noncardia GC 10 intestinal metaplasis 10 chronic    gastritis) 18 HP-positive
    The    microbial compositions of gastric mucosa from gastric cancer patients are    significantly different to the other groups.
  
  
    Khosrav,    et al. 2014 [14]
    Gastric    biopsies

      MALDI-TOF MS 16S rRNA sequencing
    215    Malaysian patients (185 FD, 22 PUD 8 GC) 
      131    HP-positive

        84 HP-negative
    The    presence of HP did not significantly modify the diversity of the gastric    microbiota. There may be geographical variations in the diversity of the    gastric microbiome.
  
  
    Yang,    et al. 2016 [25]
    Gastric    biopsies

      16S rDNA deep sequencing
    40    Colombian
      20    High GA risk town
      20    low GA risk town

        39 HP-positive
    The    gastric microbiota compositions differed between the two population.
  
  
    Yu,    et al. 2017 [27]
    Surgical    tissue 16S rRNA sequencing PICRUSt
    77    Chinese with operated GC

      Differentiated 29; Poorly; 48 Metastasis 54
    Non-malignant    tissue microbiota features were associated with family history of UGI cancer,    tumor grade and metastasis.
  
  
    Li,    et al 2017 [29]
    Gastric    biopsies 16S rDNA V3-V4 sequencing Gastric biopsies 16S rDNA V3-V4 sequencing
    33    Korean (Normal 8 Gastritis 9 IM 9; GA 7)
      11    HP-positive (before and after eradication)
    Eradication    resulted in restoration of gastric microenvironment to that of HP-negative    subject and an increase in the bacterial diversity index.
  
  
    Hsieh,    et al. 2018 [26]
    Gastric    biopsies 16S rRNA sequencing
    27    Taiwanise 
      9    gastritis (5 HP-positive)
      7    IM (4 HP-positive)

        11 GC (3 HP-positive)
    Clostridium and Fusobacterium frequently colonize in patients with GC.
  
  
    GC:    Gastric Cancer; HP: Helicobacter Pylori;    MALDI-TOF MS: Matrix-Assisted Laser Desorption Ionization Time-Of-Flight Mass    Spectrometry; FD: Functional Dyspepsia; PUD: Peptic Ulcer Disease; T-RFLP:    Terminal Restriction Fragment; PICRUSt: Phylogenetic Investigation of    Communities by Reconstruction of Unobserved States.



Table 2: Human gastric microbiota studies (including gastric cancer patients).

Hypochlorhydria and Gastric Microbiota

Gastric cancer usually develops via gastric atrophy, and the resulting hypochlorhydria potentially leads to alterations in the composition of the gastric microbiota by providing a more favourable environment for colonization. It is generally thought that the hypochlorhydria observed following proton pump inhibitor (PPI) long use does not increase the risk of gastric cancer. Moreover, autoimmune atrophic gastritis is more frequently associated with the development of gastric neuroendocrine Tumor (NET), although it also increases the risk of gastric cancer.

Two recent studies reported by Imhann et al and Jackson et al. have independently identified that long term use of PPI alter the gut microbiota to predispose to enteric infection including Clostridium difficile [16,17]. More recent study indicated influence of potassium competitive acid blocker (vonoprazan) on the gut microbiome H. pylori-negative healthy individuals. Its stronger effect was reported to be increase in the genus Streptococcus as compared with PPI (>20- fold by vonoprazan vs approximately seven fold by lansoprazole). In addition, vonoprazan, but not PPI, induced a significant increase in the core members of the oral microbiome, such as the genera Actinomyces and Rothia, in the gut microbiome [18].

In a recent British study including 95 patients consisting of five groups (20 normal, 19 PPI treated, 22 H. pylori gastritis, 23 H. pylori induced atrophic gastritis and 11 autoimmune atrophic gastritis), the microbial profiles in the stomachs of patients with H. pyloriinduced atrophic gastritis and autoimmune atrophic gastritis were quite different. Interestingly, PPI-treated patients showed more similarities in microbial diversity and abundance to the patients who had autoimmune atrophic gastritis, than to the patients who had H. pylori-induced atrophic gastritis [19]. The results agree with other previous reports that PPIs do not significantly influence the gastric microbiota [20,21], however, samples from PPI-treated patients contained significantly more Streptococcus at the Operational Taxonomic Unit (OUT) level [22]. Early interactions between H. pylori and human tissues may alter the structure of the gastric microbiome, by promoting gastric atrophy.

Gastric Cancer and Microbiota

Less-acidic conditions lead to new microbiome populations that might promote carcinogenesis, especially development gastric cancer in patients with severe atrophy. Studies examining imbalance of the gastric microbiota in H. pylori-associated gastric cancer are more limited. Dicksved, et al. [23] performed 16S rRNA gene sequencing analysis of gastric mucosa of patients with gastric cancer and found that the diversity was equal to the dyspeptic controls. The microbiota of cancer patients was predominantly composed of the genera Lactobacillus, Streptococcus, Prevotella, and Veillonella. S. mitis and S. parasanguinis were the most common species among S. genera.

More recently, Eun, et al. [24] investigated the gastric microbiota by pyrosequencing and proposed that there were marked differences in the composition and diversity among patients with gastric cancer, intestinal metaplasia and chronic gastritis, especially in Helicobacterdominant group. The relative abundance of Helicobacteraceae family was significantly lower in gastric cancer than chronic gastritis and intestinal metaplasia, while the relative abundance of Streptococcaceae family significantly increased. Furthermore, in an Unweighted Pair Group Method with Arithmetic mean (UPGMA) clustering of Helicobacter-dominant group, the chronic gastritis group and gastric cancer group were clearly separated while the intestinal metaplasia group was distributed between the two groups [24]. Their findings also suggest that the carcinogenic role of H. pylori may partly due to its impact on the gastric commensal flora, which become a favorable environment for gastric cancer development.

In a recent study comparing the gastric microbiota between the two Colombian populations; one at high-risk and one at low-risk of developing gastric cancer, significant correlations were found with the town of origin [25]. Leptotrichia wadei, which is associated with necrotizing enterocolitis, bacteremia, and a Veillonella sp., were significantly more abundant in the high risk population. Interestingly, there was no significant correlation of H. pylori phylogeographic population or carriage of the cagPAI with microbiota composition [25].

Another study from Taiwan indicated that dominant bacterial species in the H. pylori-negative patients were Burkholderia, Enterobacter, and Leclercia. The abundance of those bacteria was similar between the cancer and non-cancer groups, whereas the frequency and abundance of H. pylori were significantly lower in the cancer group. Clostridium colicanis and Fusobacterium nucleatum were significantly more abundant in patients with gastric cancer in H. pylori-negative patients [26]. Another recent study investigating nonmalignant tissue microbiota using resected stomach due to cancer indicated that Bacteroidetes was associated with lower tumor grade and Lactobacillales was negatively associated with metastasis [27]. Bacterioidetes might be involved in protection against gastric cancer progression, and the most abundant genus was Prevotella, which is commonly detected in stomach and oral samples.

Overall, there is emerging evidence of the presence of non- Helicobacter bacteria in the human stomach besides H. pylori. However, the specific role of individual microorganisms in human gastric carcinogenesis remains unclear, and further studies are needed to investigate the potential role of non- H. pylori bacteria in tumor progression.

The results of two previous studies on the bacterial diversity during the gastric cancer progression were conflicting. One study indicated the progressive decline and the other indicated the increase during progression from gastritis to cancer [28,24]. Li, et al [29] first examined the effect of H. pylori eradication on gastric microbiota and found that eradication resulted in restoration of gastric microenvironment to that of H. pylori -negative subject and an increase in the bacterial diversity index. There was an inverse association between H. pylori abundance and bacterial diversity in non-cancer gastric samples including normal, gastritis and intestinal metaplasia, this inverse association was weak in cancer samples. The bacterial diversity seems to be reduced in cancer samples with low H. pylori abundance. Eradication may possibly prevent development of gastric cancer by improvement of gastric dysbiosis and restoration of the disturbed gastric homeostasis.

Chronic Dyspepsia and Microbiota

In a study performed by Hu, et al., the prevalence of non-H. pylori bacterial was observed higher in non-ulcer dyspepsia group than in gastric ulcer group indicating the potential roles of non-H. pylori bacterial played in the pathogenesis of stomach disorders [12]. In another recent study investigating gastric microbiota using the same method, Staphylococcus spp. and Lactobacillus spp. were significantly more commonly identified in patients with chronic dyspepsia; Streptococcus spp., Pseudomonas mosselii, Escherichia coli and Klebsiella pneumoniae were more common in non-dyspeptic patients [30]. Some or all of these organisms possibly play a role in the causation of symptoms in H. pylori positive patients.

H. pylori Infection and Intestinal Microbiota

Changes of the microbiota in the duodenum and the proximal small bowel due to H. pylori infection have yet to be studied despite the causal relationship between a gastric infection with H. pylori and duodenal ulcer disease [31,32]. A recent study by Schulz, et al. [33] investigating the influence of H. pylori on duodenal and oral communities indicated the only significant influence of H. pylori infection on duodenal communities at the phylum level, which was the increased abundance of Proteobacteria probably due to transfer of Helicobacter from the stomach to the duodenal lumen. The influence of Helicobacter on the community was more evident in the duodenal samples compared to oral samples, although phylotypes in saliva were different between the H. pylori-positive and negative subjects.

In agreement of animal experiments, a study investigating faecal samples obtained from 39 H. pylori-infected patients and 19 H. pylorinegative volunteers showed that the composition of the microbiota between H. pylori-positive versus H. pylori-negative control individuals differed with regard to Clostridia and the total number of anaerobes [34]. Another study by Buhling, et al. [35] investigating the intestinal microbiota using faecal samples from 51 H. pyloripositive patients found that the microbiota of H. pylori-infected patients, characterized by an increase in growth of Lactobacilli, was different from that in H. pylori negative controls. However, fecal compartment is predominantly resident in the lumen and influenced by diet, while mucosa-adherent compartment known as mucosaassociated microbiota (MAM) consists of dense cohesive microbial communities that adhere to mucosal surface of GI tract [36]. Thus evaluating MAM in precise and accurate manners is recognized as a clinically important trial to characterize and understand the H. pylori and gut microbiome interactions and their roles in the development of health promotion and diseases.

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Citation: Shiotani A, Matsumoto H, Fukushima S, Katsumata R, Kawano M and Saito M. H. pylori and Human Gut Microbiota. J Bacteriol Mycol. 2018; 5(8): 1086.

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