Microbial ecology between Clostridioides difficile and gut microbiota.

Clostridioides difficile colonizes a polymicrobial environment in the intestine and is a causative agent for antibiotic-associated diarrhea (AAD) and pseudomembranous colitis (PMC). The most important virulence factors of C. difficile are bacterial toxins, and three toxins (toxin A, toxin B, and binary toxin) are produced by toxigenic strains. Other virulence factors include spores, flagella, capsules, biofilms, hydrolytic enzymes and adhesins. C. difficile infection (CDI) is specifically diagnosed by anaerobic culture and toxin detection by either nucleic acid amplification test (NAAT) or enzyme-linked immunosorbent assay (ELISA). For treatment of CDI, metronidazole, vancomycin and fidaxomicin are used based on the severity of CDI. Mutual interaction between C. difficile and gut microbiota is associated with pathogenesis of CDI, and decreased microbial diversity with altered gut microbiome was detected in CDI patients. Restoration of certain gut microbiota is considered to be potentially effective for the prevention and treatment of CDI, and an ideal goal for CDI patients is restoration of the gut microbiota to a healthy state. Fecal microbiota transplantation (FMT) is a highly successful method of microbiome restoration and has been reported to be effective for the prevention of recurrent CDI. In addition, approaches to restoring the gut microbiota by using probioitcs and live biotherapeutic products (LBPs) are currently being studied to examine the effect on CDI. Further microbial ecological research on C. difficile and gut microbiota could lead to a better understanding of the pathogenesis and treatment of CDI.

Establishment of a reference panel of Helicobacter pylori strains for antimicrobial susceptibility testing.

BACKGROUND: Eradication treatment for Helicobacter pylori gastritis is covered by national health insurance since 2013 in Japan. However, eradication failure due to the increase of antimicrobial resistance has become a serious problem. The present study aims to establish a reference panel of Japanese H. pylori strains for antimicrobial susceptibility testing. METHOD: A total of 28 strains were collected from 4 medical facilities in Japan. Antimicrobial susceptibility tests (ASTs) to clarithromycin (CLR), amoxicillin (AMX), and metronidazole (MNZ), were used to select standard reference strains. Complete genome sequences were also determined. RESULTS: Three H. pylori strains (JSHR3, JSHR6 and JSHR31) were selected as standard reference strains by the Japanese Society for Helicobacter Research (JSHR). The minimum inhibitory concentrations (MICs) of the antibiotics against these 3 strains by agar dilution method with Brucella-based horse-serum-containing agar medium were as follows: JSHR3 (CLR 16 µg/ml, AMX 0.032 µg/ml and MNZ 4 µg/ml), JSHR6 (CLR 0.016 µg/ml, AMX 0.032 µg/ml and MNZ 4 µg/ml), and JSHR31 (CLR 16 µg/ml, AMX 1 µg/ml and MNZ 64 µg/ml). CONCLUSIONS: A reference panel of H. pylori JSHR strains was established. The panel consisted of JSHR6, which was antibiotic-susceptible, JSHR3, which was CLR-resistant, and JSHR31, which was multi-resistant. This reference panel will be essential for standardized ASTs before the optimal drugs are selected for eradication treatment.

Prevalence estimates of Helicobacter species infection in pancreatic and biliary tract cancers.

BACKGROUND: Helicobacter pylori infection is a well-established risk factor for gastric cancer and has been linked to other gastrointestinal diseases, including pancreatic and biliary tract cancers; however, the relevance of enterohepatic non-H. pylori helicobacters to the pathophysiology of these diseases remains unclear. MATERIALS AND METHODS: We estimated the prevalence of two enterohepatic non-H. pylori helicobacters (Helicobacter hepaticus and Helicobacter bilis) in the framework of a hospital-based case-control study involving 121 patients with biliary tract cancer, pancreatic cancer, or other gastrointestinal diseases. Bile and blood samples were collected from the patients undergoing endoscopic retrograde cholangiopancreatography. The presence of H. bilis, H. hepaticus, and other Helicobacter spp. was examined using bacterial culture, PCR-based detection, and serological tests. RESULTS: Culture of Helicobacter spp. from biliary brush samples was unsuccessful. Approximately 13.0% (15/115) of the bile samples collected from patients with a variety of gastrointestinal cancers, including pancreatic and biliary tract cancers, tested positive for one of the enterohepatic non-H. pylori helicobacter species as determined by PCR. Specifically, H. bilis and H. hepaticus DNA were detected in 11 and 4 bile samples, respectively. Approximately 20%-40% of the patients tested positive for serum non-H. pylori helicobacter IgG antibodies. The seroprevalence of H. bilis and H. hepaticus in the patients without evidence of H. pylori infection appeared to be higher in the pancreatic cancer group than in the control group. CONCLUSION: Our findings suggest a role for Helicobacter spp., especially H. bilis and H. hepaticus, in the etiology of pancreatic and biliary tract cancers.

Characterization of Reproductive Microbiota of Primiparous Cows During Early Postpartum Periods in the Presence and Absence of Endometritis.

Endometritis has a major impact on fertility in postpartum dairy cows. Since previous studies showed an association between reproductive microbiota and perinatal disease, we monitored both bovine uterine and vaginal microbiota in primiparous cows to elucidate the effect of early postpartum microbiota on endometritis. Uterine and vaginal samples were collected at time points from pre-calving to 35 days postpartum (DPP), and analyzed by 16S rRNA sequencing, combined with ancillary bacterial culture. A total of seven healthy cows and seven cows diagnosed with endometritis on 35 DPP were used in the current study. The uterine and vaginal microbiota showed a maximum of 20.1% shared amplicon sequence variants (ASVs) at linked time points. 16S rRNA based analysis and traditional culture methods revealed that Trueperella showed a higher abundance in both uterus and vagina of the endometritis group compared to the healthy group on 21 DPP (U-test p < 0.05). Differential abundance analysis of the uterine microbiota showed that Enterococcus and six bacterial genera including Bifidobacterium were unique to the healthy group on the day of calving (0 DPP) and 28 DPP, respectively. In contrast, Histophilus and Mogibacteriaceae were characteristic bacteria in the vagina pre-calving in cows that later developed endometritis, suggesting that these bacteria could be valuable to predict clinical outcomes. Comparing the abundances of bacterial genera in the uterine microbiota, a negative correlation was observed between Trueperella and several bacteria including Lactobacillus. These results suggest that building an environment where there is an increase in bacteria that are generally recognized as beneficial, such as Lactobacillus, may be one possible solution to reduce the abundance of Trueperella and control endometritis.

Pathogenic Characterization of Clostridium perfringens Strains Isolated From Patients With Massive Intravascular Hemolysis.

Sepsis caused by Clostridium perfringens infection is rare but often fatal. The most serious complication leading to poor prognosis is massive intravascular hemolysis (MIH). However, the molecular mechanism underlying this fulminant form of hemolysis is unclear. In the present study, we employed 11 clinical strains isolated from patients with C. perfringens septicemia and subdivided these isolates into groups H and NH: septicemia with (n = 5) or without (n = 6) MIH, respectively. To elucidate the major pathogenic factors of MIH, biological features were compared between these groups. The isolates of two groups did not differ in growth rate, virulence-related gene expression, or phospholipase C (CPA) production. Erythrocyte hemolysis was predominantly observed in culture supernatants of the strains in group H, and the human erythrocyte hemolysis rate was significantly correlated with perfringolysin O (PFO) production. Correlations were also found among PFO production, human peripheral blood mononuclear cell (PBMC) cytotoxicity, and production of interleukin-6 (IL-6) and interleukin-8 (IL-8) by human PBMCs. Analysis of proinflammatory cytokines showed that PFO induced tumor necrosis factor-α (TNF-α), IL-5, IL-6, and IL-8 production more strongly than did CPA. PFO exerted potent cytotoxic and proinflammatory cytokine induction effects on human blood cells. PFO may be a major virulence factor of sepsis with MIH, and potent proinflammatory cytokine production induced by PFO may influence the rapid progression of this fatal disease caused by C. perfringens.

Dual interleukin-17A/F deficiency protects against acute and chronic response to cigarette smoke exposure in mice.

IL-17A and IL-17F are both involved in the pathogenesis of neutrophilic inflammation observed in COPD and severe asthma. To explore this, mice deficient in both Il17a and Il17f and wild type (WT) mice were exposed to cigarette smoke or environmental air for 5 to 28 days and changes in inflammatory cells in bronchoalveolar lavage (BAL) fluid were determined. We also measured the mRNA expression of keratinocyte derived chemokine (Kc), macrophage inflammatory protein-2 (Mip2), granulocyte-macrophage colony stimulating factor (Gmcsf) and matrix metalloproteinase-9 (Mmp9 ) in lung tissue after 8 days, and lung morphometric changes after 24 weeks of exposure to cigarette smoke compared to air-exposed control animals. Macrophage counts in BAL fluid initially peaked at day 8 and again on day 28, while neutrophil counts peaked between day 8 and 12 in WT mice. Mice dual deficient with Il17a and 1l17f showed similar kinetics with macrophages and neutrophils, but cell numbers at day 8 and mRNA expression of Kc, Gmcsf and Mmp9 were significantly reduced. Furthermore, airspaces in WT mice became larger after cigarette smoke exposure for 24 weeks, whereas this was not seen dual Il17a and 1l17f deficient mice. Combined Il17a and Il17f deficiency resulted in significant attenuation of neutrophilic inflammatory response and protection against structural lung changes after long term cigarette smoke exposure compared with WT mice. Dual IL-17A/F signalling plays an important role in pro-inflammatory responses associated with histological changes induced by cigarette smoke exposure.

Lantibiotics Produced by Oral Inhabitants as a Trigger for Dysbiosis of Human Intestinal Microbiota.

Lantibiotics are a type of bacteriocin produced by Gram-positive bacteria and have a wide spectrum of Gram-positive antimicrobial activity. In this study, we determined that Mutacin I/III and Smb (a dipeptide lantibiotic), which are mainly produced by the widespread cariogenic bacterium Streptococcus mutans, have strong antimicrobial activities against many of the Gram-positive bacteria which constitute the intestinal microbiota. These lantibiotics also demonstrate resistance to acid and temperature. Based on these features, we predicted that lantibiotics may be able to persist into the intestinal tract maintaining a strong antimicrobial activity, affecting the intestinal microbiota. Saliva and fecal samples from 69 subjects were collected to test this hypothesis and the presence of lantibiotics and the composition of the intestinal microbiota were examined. We demonstrate that subjects possessing lantibiotic-producing bacteria in their oral cavity exhibited a tendency of decreased species richness and have significantly reduced abundance of the phylum Firmicutes in their intestinal microbiota. Similar results were obtained in the fecal microbiota of mice fed with S. mutans culture supernatant containing the lantibiotic bacteriocin Mutacin I. These results showed that lantibiotic bacteriocins produced in the oral cavity perturb the intestinal microbiota and suggest that oral bacteria may be one of the causative factors of intestinal microbiota dysbiosis.

Sex-dependent differences in the gut microbiota following chronic nasal inflammation in adult mice.

A growing body of evidence suggests a relationship between olfactory dysfunction and the pathogenesis of mental disorders. Our previous studies indicated that chronic nasal inflammation caused loss of olfactory sensory neurons and gross atrophy of the olfactory bulb, which may lead to olfactory dysfunction. Simultaneously, increasing numbers of reports have elucidated the importance of gut microbiota to maintain brain function and that dysbiosis may be associated with neuropsychiatric disorders. Here we examined whether chronic nasal inflammation perturbed gut microbiota and whether there were sex differences in this pattern. Eight-week-old C57BL/6 mice repeatedly received bilateral nasal administration of lipopolysaccharide (LPS) 3 times/week to cause chronic nasal inflammation or saline as a control. At 9 weeks, cecal feces were used for 16S metagenomic analysis and whole blood and fresh tissue of spleen were used for ELISA analyses. Microbiome analysis demonstrated a remarkable change of the gut microbiota in male mice with chronic nasal inflammation which was different from that in female mice. In both mice, systemic inflammation did not occur. This has shown for the first time that chronic nasal inflammation correlates with sex-dependent changes in the gut microbiota. The detailed mechanism and potential alteration to brain functions await further studies.

The updated JSPGHAN guidelines for the management of Helicobacter pylori infection in childhood.

The Japan Pediatric Helicobacter pylori Study Group published the first guidelines on childhood H. pylori infection in 1997. They were later revised by the Japanese Society for Pediatric Gastroenterology, Hepatology and Nutrition (JSPGHAN). The H. pylori eradication rates, when employing triple therapy with amoxicillin and clarithromycin, currently recommended as the first-line therapy of H. pylori infection in Japan, have substantially decreased, creating an important clinical problem worldwide. In Japanese adults, the "test-and-treat" strategy for H. pylori infection is under consideration as an approach for gastric cancer prevention. However, the combined North American and European pediatric guidelines have rejected such a strategy for asymptomatic children. As risk for gastric cancer development is high in Japan, determining whether the "test-and-treat" strategy can be recommended in children has become an urgent matter. Accordingly, the JSPGHAN has produced a second revision of the H. pylori guidelines, which includes discussion about the issues mentioned above. They consist of 19 clinical questions and 34 statements. An H. pylori culture from gastric biopsies is recommended, not only as a diagnostic test for active infection but for antimicrobial susceptibility testing to optimize eradication therapy. Based upon antimicrobial susceptibility testing of H. pylori strains (especially involving clarithromycin), an eradication regimen including use of the antibiotics to which H. pylori is susceptible is recommended as the first-line therapy against H. pylori-associated diseases. The guidelines recommend against a "test-and-treat" strategy for H. pylori infection for asymptomatic children to protect against the development of gastric cancer because there has been no evidence supporting this strategy.

Biofilm Formation and Antibiotic Resistance Phenotype of Helicobacter pylori Clinical Isolates.

We evaluated biofilm formation of clinical Helicobacter pylori isolates from Indonesia and its relation to antibiotic resistance. We determined the minimum inhibition concentration (MIC) of amoxicillin, clarithromycin, levofloxacin, metronidazole and tetracycline by the Etest to measure the planktonic susceptibility of 101 H. pylori strains. Biofilms were quantified by the crystal violet method. The minimum biofilm eradication concentration (MBEC) was obtained by measuring the survival of bacteria in a biofilm after exposure to antibiotics. The majority of the strains formed a biofilm (93.1% (94/101)), including weak (75.5%) and strong (24.5%) biofilm-formers. Planktonic resistant and sensitive strains produced relatively equal amounts of biofilms. The resistance proportion, shown by the MBEC measurement, was higher in the strong biofilm group for all antibiotics compared to the weak biofilm group, especially for clarithromycin (p = 0.002). Several cases showed sensitivity by the MIC measurement, but resistance according to the MBEC measurements (amoxicillin, 47.6%; tetracycline, 57.1%; clarithromycin, 19.0%; levofloxacin, 38.1%; and metronidazole 38.1%). Thus, biofilm formation may increase the survival of H. pylori and its resistance to antibiotics. Biofilm-related antibiotic resistance should be evaluated with antibiotic susceptibility.

Genome-wide association meta-analysis identifies GP2 gene risk variants for pancreatic cancer.

Pancreatic cancer is the fourth leading cause of cancer-related deaths in Japan. To identify risk loci, we perform a meta-analysis of three genome-wide association studies comprising 2,039 pancreatic cancer patients and 32,592 controls in the Japanese population. Here, we identify 3 (13q12.2, 13q22.1, and 16p12.3) genome-wide significant loci (P < 5.0 × 10-8), of which 16p12.3 has not been reported in the Western population. The lead single nucleotide polymorphism (SNP) at 16p12.3 is rs78193826 (odds ratio = 1.46, 95% confidence interval = 1.29-1.66, P = 4.28 × 10-9), an Asian-specific, nonsynonymous glycoprotein 2 (GP2) gene variant. Associations between selected GP2 gene variants and pancreatic cancer are replicated in 10,822 additional cases and controls of East Asian origin. Functional analyses using cell lines provide supporting evidence of the effect of rs78193826 on KRAS activity. These findings suggest that GP2 gene variants are probably associated with pancreatic cancer susceptibility in populations of East Asian ancestry.

Acanthamoeba castellanii supports extracellular survival of Helicobacter pylori in co-culture.

This study aimed to demonstrate whether Helicobacter pylori is able to survive in co-culture with a protozoan, Acanthamoeba castellanii, in order to further investigate a possible aqueous environmental mode of transmission. Numbers of H. pylori in co-culture with A castellanii were assessed by colony forming unit (CFU) assay and cell morphology was observed by electron microscopy. Viable and intact H. pylori in co-culture were detected and the number of H. pylori in co-culture with A. castellanii was significantly higher than in bacterial single culture. It was also shown that co-culture of H. pylori with A. castellanii physically separated by a filter membrane negated this survival effect, suggesting that adherence of H. pylori to A. castellanii affects its survival. Scanning electron microscopy revealed helical forms of H. pylori in co-culture with A. castellanii, but not in single culture. These results imply that mutual interaction between H. pylori and A. castellanii in the environment is critical for survival of H. pylori. In addition, the H. pylori gene expression profile was found to differ between single and co-cultured cells using RNA-sequence analysis.

Serum Pepsinogen Values in Japanese Junior High School Students With Reference to Helicobacter Pylori Infection.

BACKGROUND: Distributions of serum pepsinogen (PG) values were assessed in Helicobacter pylori-infected and non-infected junior high school students (aged 12-15 years) in Japan. METHODS: All junior high school students (1,225 in total) in Sasayama city, who were basically healthy, were asked to provide urine and serum samples, which were used to measure urine and serum H. pylori antibodies using ELISA kits and PG values. The subjects, whose urine and serum antibodies were both positive, were considered H. pylori infected. RESULTS: Of the 187 subjects who provided urine and blood samples, 8 were infected, 4 had discrepant results, 4 had negative serum antibody titers no less than 3.0 U/ml, and 171 were non-infected. In the H. pylori non-infected subjects, the median PG I and PG II values and PG I to PG II ratio (PG I/II) were 40.8 ng/mL, 9.5 ng/mL, and 4.4, respectively, whereas in the infected subjects, these values were 55.4 ng/mL, 17.0 ng/mL, and 3.3, respectively (each P < 0.01). In the non-infected subjects, PG I and PG II were significantly higher in males than in females (P < 0.01). CONCLUSIONS: The PG I and PG II values were higher, and the PG I/II was lower in H. pylori infected students than in non-infected students. In H. pylori non-infected students, males showed higher PG I and PG II values than females. The distributions of PG values in junior high school students differed from those in adults.

Skin and Soft Tissue Infections Caused by Different Genotypes of PVL-Positive Community-Acquired Methicillin-Resistant Staphylococcus aureus Strains.

Panton-Valentine leukocidin (PVL) is a causative agent of lethal necrotizing pneumonia and is associated with epidemic strains of community-acquired (CA) methicillin-resistant Staphylococcus aureus (MRSA). PVL-producing strains have rarely been isolated in Japan. However, PVL-positive CA-MRSA has been isolated much more frequently in recent years. To investigate the relevance of pvl genes (lukS/F-PV) and clinical traits in epidemic S. aureus strains, we genotyped four PVL-positive CA-MRSA strains isolated from patients with skin and soft tissue infections and measured their susceptibility to antibiotics. Three of the isolates matched the genotype of the USA300 clone, which has predominantly been isolated in the USA. The remaining strain matched the ST217 genotype, and its spa type was identical to that of PVL-positive strains previously reported in India and China. Abscess drainage was necessary in all cases, and deep cutaneous ulcers were formed in three out of four cases regardless of the genotype. The ST217 genotype strain was resistant to clindamycin, in addition to quinolones, macrolides, and aminoglycosides. Thus, diagnostic determination of lukS/F-PV should be used as a guide for selecting the treatment regimen.

Analysis of the colonic mucosa associated microbiota (MAM) using brushing samples during colonic endoscopic procedures.

The mucosa-associated microbiota is an important component in human microbiota. The aim was to investigate mucosa-associated microbiota using brush samples during endoscopic procedures and compare with fecal microbiota. Seven patients who were planning to undergo a routine colonoscopy were recruited. Mucosal brushing samples were taken from 3 sites (terminal ileum, ascending and sigmoid colon), and a fecal sample was taken on the morning of colonoscopy. The samples were immediately placed in microcentrifuge tubes containing DNA stabilization reagent and analyzed using the next generation sequencer. The individual differences in microbiota were more evident than the differences of the sampling sites. Actinobacteria was more abundant and Bacteroidetes was less in the brush samples than those in the fecal samples. Taxonomic composition at the genus level and the proportion of genes responsible for some functions in the brushing samples tended to be different from those in the fecal samples. Bulleidia and Oribacteriumi were significantly more abundant and the proportions of genes responsible for transcription factors and phosphotransferase system were higher in ileal mucous than those in fecal samples. Brushing during colonoscopic procedure instead of using feces samples might be useful to analyze mucosa-associated microbiota.

Role of Probiotics in Eradication Therapy for Helicobacter pylori Infection.

Probiotics are defined as, "Live microorganisms that, when administered in adequate amounts, confer a health benefit on the host", and have various effects including inhibitory capabilities on pathogens, stimulation of organ functions and activation of immune responses in the human. Probiotics were reported to inhibit Helicobacter pylori not only in vitro, but also in vivo studies. The mechanisms by which probiotics inhibit H. pylori infection include competition for nutrients, production of bactericidal substances, competitive inhibition of adherence and stimulation of host functions and immunity. In addition, probiotics are clinically used for eradication therapy of H. pylori infection, and the effects of probiotics as single treatment and combination use with other drugs including proton pump inhibitors and antibiotics against H. pylori are reported. It has been testified that probiotics increase the eradication rate and prevent adverse events including diarrhea, nausea, vomiting and taste disorder. In the Maastrich V/Florence Consensus Report 2017, it was stated that some probiotics may have a beneficial effect on H. pylori eradication and are effective in reducing side effects of eradication therapy, but more research is needed to answer several questions concerning the mechanisms of probiotics action. In addition, strain specificity, dosages and duration times of probiotics used for H. pylori eradication therapy need to be clarified in future studies.

Effect of Helicobacter pylori biofilm formation on susceptibility to amoxicillin, metronidazole and clarithromycin.

The human gastric pathogen Helicobacter pylori forms biofilms in vitro and in vivo. We previously demonstrated that H. pylori biofilm formation in vitro decreased its susceptibility to clarithromycin (CAM). The aim of this study was to evaluate the effects of biofilm formation on amoxicillin (AMPC) and metronidazole (MNZ) susceptibility. In addition, we assessed the influence of biofilms of CAM resistant H. pylori on CAM susceptibility. It was shown that high levels of efflux pump gene transcripts were detected in biofilm cells of all H. pylori strains used in this study. H. pylori biofilm biomass was significantly decreased compared to initial biomass after treatment with the minimum inhibitory concentration (MIC) of AMPC. Similarly, the biofilm biomass of H. pylori decreased after treatment with MIC of MNZ, although the difference was not statistically significant. However, minimum bactericidal concentrations (MBCs) of AMPC or MNZ to biofilm cells were higher than those of planktonic cells. The biofilm biomasses of all of the CAM resistant strains were significantly decreased compared to initial biomass after treatment with 2x MIC of CAM. However, the viability of the CAM treated biofilm cells with 2x MIC of CAM was not significantly reduced compared to initial cell numbers with the exception of one strain. The viability of biofilm cells of all strains was higher than that of planktonic cells after treatment with various concentrations of CAM. These results indicate that biofilm cells were more resistant to these antibiotics than planktonic cells and that the assessment of the ability to form biofilms in H. pylori is important for eradication of this microorganism.

Community-acquired fulminant colitis caused by binary toxin-producing Clostridium difficile in Japan.

We report a case of community-acquired fulminant colitis caused by Clostridium difficile in Japan. A 46-year-old woman was diagnosed with severe infectious enterocolitis and was admitted at another hospital. The stool culture was positive for toxigenic C. difficile. Since the patient presented with fulminant C. difficile infection (CDI) with toxic megacolon, respiratory insufficiency, and circulatory failure, she was transferred to Kyorin University Hospital for intensive care. Intubation and antibiotic therapy were performed. The general condition improved with conservative treatment, and she was discharged without sequelae. While the recovered isolate was toxin A and B-positive and binary toxin-positive, it was identified as polymerase chain reaction (PCR) ribotype ts0592 and slpA sequence type ts0592. The isolate was different from PCR ribotype 027 epidemic in Europe and North America. In Japan, binary toxin-producing strains are rare and have not caused an epidemic to date. Furthermore, there are few data on community-acquired CDI in Japan. In this case, a non-elderly woman with no major risk factors such as antibiotic use, administration of proton pump inhibitor and history of gastrointestinal surgery developed community-acquired fulminant CDI caused by the binary toxin-positive strain, and ICU treatment was required. Further studies focusing on the role of binary toxin-positive C. difficile in the severity of community-acquired CDI are necessary.

Enhanced infectivity of strains of Helicobacter pylori isolated from children compared with parental strains.

PURPOSE: Intra-familial infection, mother-to-child infection, is considered to be one of the main routes of transmission for Helicobacter pylori, in developed countries such as Japan. A major role for intra-familial spread in the pathogenicity of H. pylori is now beyond controversy, although the major route of transmission remains poorly understood. We performed this study to clarify the factors determining intra-familial transmission. METHODOLOGY: We used several H. pylori strains isolated from family members to compare infectivity. H. pylori K21 and K22 strains were isolated from the father and mother, and the K25 strain was isolated from the third child of the family. Mongolian gerbils were inoculated with H. pylori strains and the infectivity of three strains was compared in each experiment. In addition, the whole genome sequence, adhesion to gastric epithelial cells and the growth of static condition or continuous flow culture among three strains of H. pylori were analysed.Results/Key findings. Most of the colonies were determined as the same molecular type K25 in all of the four grouped animals and H. pylori K25 was observed as the dominant strain. The stronger adhesion capacity of the K25 strain was observed in comparison with the other two strains through in vitro analysis. By assessing the genomic profiles of H. pylori isolates from three strains, identified TnPZ regions were detected only in the K25 strain. CONCLUSION: The infectivity of H. pylori isolates intra-familial infection and animal infection were prescribed by the adhesion capacity and molecular type of each strain.

Effects of Probiotic Lactobacillus acidophilus Strain INMIA 9602 Er 317/402 and Putative Probiotic Lactobacilli on DNA Damages in the Small Intestine of Wistar Rats In Vivo.

Double-strand breaks in the DNA of the small intestine in male Wistar rats were studied using a neutral comet assay after 7 days of feeding with a single strain probiotic formulation Narine (Vitamax-E, Armenia), containing Lactobacillus acidophilus strain Er-2317/402 Narine, and putative probiotics L. rhamnosus Vahe and L. delbrueckii IAHAHI. Type 0 (undamaged DNA), type 1 (head diameter 13.18-17.08 µm), and type 2 (14.15-µm head diameter) damaged DNA comets were studied in control and lactobacilli-fed rats using the neutral comet assay. Lactobacilli-fed rats were shown to carry only type 0 (undamaged) DNA.Thus, the effects of probiotic Lactobacillus acidophilus strain INMIA 9602 Er 317/402 and putative probiotic lactobacilli on DNA damage in the small intestine of Wistar rats in vivo was shown, and the neutral comet assay is suggested as a potential tool for the in vivo selection of putative probiotics with DNA-protective activity.