Effect of Probiotics on the Fecal Resistome During Helicobacter Pylori Eradication Therapy
Condition: Gut Resistome
Study type: Interventional
Phase: Phase 4

Summary: 
      Eradication therapy against Helicobacter pylori (Hp) carries adverse effects, such as
      altering the intestinal microbiota's structure and function and selecting commensals and
      pathogens resistant to antibiotics. This last undesirable effect turns the microbiota into a
      reservoir of resistance genes. Saccharomyces boulardii CNCM I-745 (Sb) can improve dysbiosis
      and reduce the abundance of multi-resistant bacteria. The objective of the current project is
      to characterize the resistome of individuals treated with anti-H. pylori therapy in the
      presence or absence of Sb in fecal samples. Applying metagenomics and using next-generation
      sequencing tools, the investigators seek to demonstrate the beneficial effect of Sb on the
      gut microbiota by reducing the abundance of resistance genes and characterizing bacteria
      modulated by this probiotic-yeast. The investigators expect to find an increase in the
      diversity and relative abundance of antibiotic resistance genes (ARGs) in the intestine of
      participants who did not receive Sb during Hp eradication therapy and one month after
      completing treatment. The increase in ARGs is probably correlated with the presence of
      Clostridia and Bacteroides.
    

Detailed description: 
      Antibiotic use is one of the most common therapeutic strategies of modern medicine. Together
      with the significant beneficial effects of antibiotic use, side effects accompany the use of
      these drugs, including the increasing appearance of antibiotic-resistant microbes, a global
      public health problem. Other significant side effects include gastrointestinal-related
      symptoms such as diarrhea, pain, intestinal discomfort, and flatulence, which can be the
      consequence of dysbiosis. A common strategy to limit antibiotic-associated side effects is
      the addition of probiotics, as S. boulardii CNCM I-745 (Sb). The use of bacterial probiotics
      to reduce antibiotic-associated side effects has several potential limitations, including the
      destruction of probiotics, development of probiotics strains resistant to antibiotics, and
      passage of antibiotic-resistant genes to pathogenic bacteria through horizontal gene
      transfer. Sb is a probiotic yeast whose benefits on intestinal dysbiosis have been associated
      with establishing a favorable growth environment for the normal intestinal microbiota. Due to
      the yeast nature of this probiotic, the limitations of bacterial-probiotics cannot be
      accounted for during Sb use. Consequently, it is crucial to study the molecular mechanism
      elicited by Sb on intestinal microbiota, including changes in resistome.

      In our previous study regarding the supplementation with Sb on the treatment against H.
      pylori (Hp) infection, patients that received the probiotic had a significantly lower
      frequency of gastrointestinal symptoms. Additionally, those patients have a higher number of
      bacterial diversity evenness (P=0.0156), higher abundance of Enterobactereacea, and lower
      abundance of Bacteroides and Clostridia upon treatment completion and one month later.
      Bacteroides and Clostridia have been previously implicated as antibiotic multi-resistant
      pro-inflammatory strains. The objective of the current project is to characterize the
      resistome of individuals treated with anti-H. pylori therapy in the presence or absence of Sb
      in fecal samples.

      Specific aims To determine the resistome changes before and after Hp eradication treatment in
      patients that have received or not Sb.

      To compare the resistome patterns between patients treated for Hp infection that have
      received or not SB, at the time of completion of treatment and one month after finishing
      them.

      Based on the shotgun metagenomics, the researchers seek to characterize the microbiome and
      resistome in fecal samples and understand the microbiome's functional characteristics,
      including virulence genes, metabolic pathways, and mobile genetic elements.

      Methodology DNA obtained from the previous microbiome characterization will be used in this
      project. In case DNA is not enough, the researchers will extract DNA from the frozen stored
      samples using the Power Soil kit.

      DNA will be shipped to UNC Chapel Hill or the University of Minnesota, where genomic
      libraries will be constructed. Hi-Seq Illumina runs will be used to produce a sequencing
      deepness of 10 million per sample.

      FASTQC will be used for reads quality control and multiQC as reports generator from the raw
      reads. Trimmomatic will be used to trim Illumina adapters and to cut reads on the average
      quality. As human DNA is present on the fecal samples, the investigators have to remove the
      host DNA using the mapping software BWA and SAMtools. Using the Resfinder database, the
      investigators must align the filtered reads and generate a count table using
      ResistomeAnalyzer. Finally, the researchers can create rarefaction curves based on the table
      made using AMRPlusPlus.

      Expected results The researchers expect to find a higher diversity and abundance of
      antibiotic resistance genes (genome intrinsic and carried by mobile genetic elements) on the
      group that did not receive Sb at the end of treatment and one month after its completion.
      This would be correlated with the lower abundance of Clostridia and Bacteroides encountered
      in the 16S rRNA gene microbiome characterization.

      The investigators will also be able to sub-characterize the resistome diversity and abundance
      of antibiotic resistance genes encountered typically on Enterobacteriaceae. A previous
      analysis found a higher abundance of Enterobacteriaceae in the group that received Sb
      (previously reported in a mouse model by Sovran et al. 2018). Additionally, since the
      researchers will obtain the raw reads on the shotgun sequencing, they could further
      characterize the bacteria modulated by the use of Sb at strain level (Bacteroides,
      Clostridia, and Enterobacteriaceae). Together, the researchers expect to obtain valuable data
      on the use of Sb regarding antibiotic resistance and further microbiome characteristics,
      including genes of bacterial pathogenicity. These results will contribute to clarify the
      beneficial effects of Sb addition in patients treated for Hp infection.
    

Intervention name(s): Saccharomyces Boulardii-Containing Product in Oral Dose Form
Intervention type(s): Drug
Intervention description(s): S. boulardii CNCM I-745 (approximately 22.5 x109 CFU)
Allocation: Randomized
Intervention model: Parallel Assignment
Intervention model description: A total of 72 H. pylori-positive patients were enrolled, and they were randomly assigned to group conventional (A) and group conventional plus S. boulardi (B). Patients in group A received 14-day triple therapy consisting of amoxicillin 1 g three times a day, tinidazole 1 g once a day, and omeprazole 40 mg twice a day. Patients in group B received triple therapy supplemented with S. boulardii CNCM I-745 (approximately 22.5 x 10^9 CFU) for 14 days.
Masking: Single (Participant)
Primary purpose: Treatment
Keywords: Saccharomyces boulardii; eradication therapy; Helicobacter Pylori [H. Pylori]