Transposable Element ISHp608 of Helicobacter pylori: Nonrandom Geographic Distribution, Functional Organization, and Insertion Specificity

A new member of the IS605 transposable element family, designated ISHp608, was found by subtractive hybridization in Helicobacter pylori. Like the three other insertion sequences (ISs) known in this gastric pathogen, it contains two open reading frames (orfA and orfB), each related to putative transposase genes of simpler (one-gene) elements in other prokaryotes; orfB is also related to the Salmonella virulence gene gipA. PCR and hybridization tests showed that ISHp608 is nonrandomly distributed geographically: it was found in 21% of 194 European and African strains, 14% of 175 Bengali strains, 43% of 131 strains from native Peruvians and Alaska natives, but just 1% of 223 East Asian strains. ISHp608 also seemed more abundant in Peruvian gastric cancer strains than gastritis strains (9 of 14 versus 15 of 45, respectively; P = 0.04). Two ISHp608 types differing by approximately 11% in DNA sequence were identified: one was widely distributed geographically, and the other was found only in Peruvian and Alaskan strains. Isolates of a given type differed by < or = 2% in DNA sequence, but several recombinant elements were also found. ISHp608 marked with a resistance gene was found to (i) transpose in Escherichia coli; (ii) generate simple insertions during transposition, not cointegrates; (iii) insert downstream of the motif 5"-TTAC without duplicating target sequences; and (iv) require orfA but not orfB for its transposition. ISHp608 represents a widespread family of novel chimeric mobile DNA elements whose further analysis should provide new insights into transposition mechanisms and into microbial population genetic structure and genome evolution.     

Conservation and Diversity of the Helicobacter pylori Copper-Transporting ATPase Gene (copA) Sequence Among Helicobacter Species and Campylobacter Species Detected by PCR and RFLP

Background Helicobacter pylori is a causative pathogen of such human stomach diseases as chronic type B gastritis, ulcer, and possibly gastric carcinoma. As a co-factor in various redox enzymes and an essential trace metal required for the synthesis of metalloproteins, copper might play a role in the pathogenesis of H. pylori. A gene, copA , associated with copper transport, has been isolated from H. pylori UA802. In this study, conservation and diversity of this gene were analyzed among some Helicobacter and Campylobacter species.
Materials and Methods. Twenty-one clinical isolates and strains of helicobacters and campylobacters were used in this study. Methods including polymerase chain reaction (PCR) amplification, restriction fragment-length polymorphisms (RFLPs), and hybridization were employed to carry out this work.
Results. The copA gene was highly conserved in all the H. pylori isolates tested ( Helicobacter nemestrinae and Helicobacter felis but not in Helicobacter mustelae and the Campylobacter species), whereas the sequence downstream of the copA appears to diverge among H. pylori isolates. In addition, two restriction patterns of the PCR-amplified copA fragments from seven H. pylori isolates and H. nemestrinae were identified, and the RFLP of H. nemestrinae was identical to that of one of the H. pylori isolate group.
Conclusions. The adenosine triposphatase-derived copper-transporting mechanism is employed by various H. pylori strains, H. nemestrinae, H. felis , and perhaps by other Helicobacter species. The nucleotide mutations have risen in the copA gene. It appears that there is a genetic relatedness of the copA gene to H. pylori and H. nemestrinae.     

Distinctiveness of genotypes of Helicobacter pylori in Calcutta, India

The genotypes of 78 strains of Helicobacter pylori from Calcutta, India (55 from ulcer patients and 23 from more-benign infections), were studied, with a focus on putative virulence genes and neutral DNA markers that were likely to be phylogenetically informative. PCR tests indicated that 80 to 90% of Calcutta strains carried the cag pathogenicity island (PAI) and potentially toxigenic vacAs1 alleles of the vacuolating cytotoxin gene (vacA), independent of disease status. This was higher than in the West (where cag PAI(+) vacAs1 genotypes are disease associated) but lower than in east Asia. The iceA2 gene was weakly disease associated in Calcutta, whereas in the West the alternative but unrelated iceA1 gene at the same locus is weakly disease associated. DNA sequence motifs of vacAm1 (middle region) alleles formed a cluster that was distinct from those of east Asia and the West, whereas the cagA sequences of Calcutta and Western strains were closely related. An internal deletion found in 20% of Calcutta iceA1 genes was not seen in any of approximately 200 strains studied from other geographic regions and thus seemed to be unique to this H. pylori population. Two mobile DNAs that were rare in east Asian strains were also common in Calcutta. About 90% of Calcutta strains were metronidazole resistant. These findings support the idea that H. pylori gene pools differ regionally and emphasize the potential importance of studies of Indian and other non-Western H. pylori populations in developing a global understanding of this gastric pathogen and associated disease.     

Helicobacter pylori genotyping from American indigenous groups shows novel Amerindian vacA and cagA alleles and Asian, African and European admixture

It is valuable to extend genotyping studies of Helicobacter pylori to strains from indigenous communities across the world to better define adaption, evolution, and associated diseases. We aimed to genetically characterize both human individuals and their infecting H. pylori from indigenous communities of Mexico, and to compare them with those from other human groups. We studied individuals from three indigenous groups, Tarahumaras from the North, Huichols from the West and Nahuas from the center of Mexico. Volunteers were sampled at their community site, DNA was isolated from white blood cells and mtDNA, Y-chromosome, and STR alleles were studied. H. pylori was cultured from gastric juice, and DNA extracted for genotyping of virulence and housekeeping genes. We found Amerindian mtDNA haplogroups (A, B, C, and D), Y-chromosome DYS19T, and Amerindian STRs alleles frequent in the three groups, confirming Amerindian ancestry in these Mexican groups. Concerning H.pylori cagA phylogenetic analyses, although most isolates were of the Western type, a new Amerindian cluster neither Western nor Asian, was formed by some indigenous Mexican, Colombian, Peruvian and Venezuelan isolates. Similarly, vacA phylogenetic analyses showed the existence of a novel Amerindian type in isolates from Alaska, Mexico and Colombia. With hspA strains from Mexico and other American groups clustered within the three major groups, Asian, African or European. Genotyping of housekeeping genes confirmed that Mexican strains formed a novel Asian-related Amerindian group together with strains from remote Amazon Aborigines. This study shows that Mexican indigenous people with Amerindian markers are colonized with H. pylori showing admixture of Asian, European and African strains in genes known to interact with the gastric mucosa. We present evidence of novel Amerindian cagA and vacA alleles in indigenous groups of North and South America.     

Expression of histo-blood group antigens by lipopolysaccharides of Helicobacter pylori strains from asian hosts: the propensity to express type 1 blood-group antigens

Past studies have shown that the cell surface lipopolysaccharides (LPSs) of the ubiquitous human gastric pathogen Helicobacter pylori (a type 1 carcinogen) isolated from people residing in Europe and North America express predominantly type 2 Lewis x (Le(x)) and Le(y) epitopes and, infrequently, type 1 Le(a), Le(b), and Le(d) antigens. This production of Lewis blood-group structures by H. pylori LPSs, similar to those found in the surfaces of human gastric cells, allows the bacterium to mimic its human niche. In this study, LPSs of H.pylori strains extracted from patients living in China, Japan, and Singapore were chemically and serologically analyzed. When compared with Western H.pylori LPSs, these Asian strains showed a stronger tendency to produce type 1 blood groups. Of particular interest, and novel observations in H.pylori, the O-chain regions of strains F-58C and R-58A carried type 1 Le(a) without the presence of type 2 Le(x), strains R-7A and H607 were shown to have the capability of producing the type 1 blood group A antigen, and strains CA2, H507, and H428 expressed simultaneously the difucosyl isomeric antigens, type 1 Le(b) and type 2 Le(y). The apparent proclivity for the production of type 1 histo-blood group antigens in Asian H.pylori LPSs, as compared with Western strains, may be an adaptive evolutionary effect in that differences in the gastric cell surfaces of the respective hosts might be significantly dissimilar to select for the formation of different LPS structures on the resident H.pylori strain.     

Clinical and pathological importance of cagA-positive Helicobacter pylori strains in children with abdominal complaints

Background. The aim of this study was to assess the correlation between the prevalence of Helicobacter pylori strains possessing cytotoxin-associated gene A ( cag A) in children and the intensity of clinical complaints and morphological changes of the gastric mucosa.
Materials and Methods. A group of 80 children with gastrointestinal complaints was included in this study. Pathologists examined mucosal biopsy specimens from these patients. The urease test and multiplex polymerase chain reaction (MPCR) were used to identify H. pylori strains.
Results. In the group of children infected with cag A-positive H. pylori strains, fourth-degree gastritis was more frequent than in the group with cag A-negative H. pylori colonization. In histopathological assessment, infection with cag A-positive H. pylori was associated also with higher grades of inflammatory intensity and activity.
Conclusions. Marked inflammation of the antral mucosa was significantly more frequent in children infected with cag A-positive H. pylori than in those infected with cag A-negative H. pylori , as assessed endoscopically and histopathologically. No specific symptoms for cag A-positive and cag A-negative H. pylori infection were observed.     

Identification of the repeated number of C and D regions of tyrosine phosphorylation motifs in Helicobacter pylori cagA using multiplex PCR

Various tyrosine phosphorylation motif regions of H. pylori cagA exist. The number of these regions was found to have some influence on cell signaling, which was found to be more pronounced when in D (ESS) region than in C (WSS) region. A molecular biological method with multiplex PCR was developed to distinguish C and D regions, and to identify the repetition number of tyrosine phosphorylation of the cagA gene. Multiplex PCR using novel primer sets was performed on 73 strains of H. pylori isolated from Korean patients with upper gastrointestinal diseases. The Western cagA was identified in only 3 strains (4.1%) whereas East Asia cagA was identified in 69 strains (94.5%). These results were reconfirmed through a sequencing analysis. The method developed in this study would be useful for monitoring the repeated number of C and D regions of tyrosine phosphorylation motifs in H. pylori cagA.     

cagA gene variants in Malaysian Helicobacter pylori strains isolated from patients of different ethnic groups

Helicobacter pylori infection of a distinct subtype of cagA may lead to different pathological manifestation. The aim of this study is to determine the presence of cagA gene and its variants in H. pylori infection among different ethnic groups and its effect on gastroduodenal diseases. Overall detection of cagA among the 205 clinical isolates of H. pylori was 94%. Variations in size of the 3' region of cagA gene were examined among 192 Malaysian H. pylori cagA-positive strains. Results showed that three cagA variants differing in fragment length of PCR products were detected and designated as type A (621-651bp), type B (732-735bp) and type C (525 bp). Although there was no association between any of the cagA subtypes with peptic ulcer disease (p>0.05), an association between cagA subtypes with a specific ethnic group was observed. Specific-cagA subtype A strains were predominantly isolated from Chinese compared to Malays and Indians (p<0.0005), and cagA subtype B strains were predominantly isolated from Malays and Indians compared to Chinese (p<0.05). The cagA type A strains of H. pylori is commonly found in the Chinese patients who have a higher risk of peptic ulcer disease, thus indicating that it could be used as an important clinical biomarker for a more severe infection.     

AP—PCR结合进化树分析在幽门螺杆菌地域起源特征研究中的价值

目的探讨随机引物PCR（Arbitrary primerPCR,AP—PCR）结合克隆测序及生物信息学分析等方法在研究幽门螺杆菌（Helicobacter pylori,Hpylori）菌株地域起源特征中的价值和意义。方法针对临床分离培养的Hpylori菌株的基因组DNA,采用一组10nt的寡核苷酸引物进行随机PCR扩增,选取相对保守的片段进行回收、克隆及测序,测序的基因序列提交GenBank数据库进行序列相似性的BLAST比对,收集BLAST比对得到的同源性较高不同地域来源螺杆菌的对应序列,用ClustalX软件进行排序,采用Mega4．0软件中的邻位相连法（Neighbor-joining）和最大简约法（Maximum—parsimony）进行进化树分析。结果随机引物扩增及筛选克隆测序得到的基因产物为NADH脱氢酶G和H亚单位的部分编码序列,与27株不同地域来源H．pylori及1株猫科动物来源螺杆菌菌株的同源性均高达90％以上,表明Hpylori中NADH脱氢酶基因序列为保守结构,进化树分析显示：采用AP．PCR方法得到的Hpylori临床菌株的基因序列,显示出东亚菌株来源的遗传特征,与具有东亚菌株特征的美洲秘鲁Sat464和Shi470菌株、韩国的52、51菌株、日本的1757菌株遗传距离较近,与南亚、欧洲菌株距离较远,与非洲的SouthAffica7菌株和猫科动物来源的Sheeba菌株的遗传距离最远。结论不仅某些特殊基因可以反映地域差异,随机定位相对保守的基因片段同样可以反映Hpylori的地域起源特征。AP—PCR、测序等技术方法与进化树分析相结合是探讨Hpylori地域起源特征的一种更为便捷有效的新方法。     

Metronidazole activation is mutagenic and causes DNA fragmentation in Helicobacter pylori and in Escherichia coli containing a cloned H. pylori RdxA(+) (Nitroreductase) gene

Much of the normal high sensitivity of wild-type Helicobacter pylori to metronidazole (Mtz) depends on rdxA (HP0954), a gene encoding a novel nitroreductase that catalyzes the conversion of Mtz from a harmless prodrug to a bactericidal agent. Here we report that levels of Mtz that partially inhibit growth stimulate forward mutation to rifampin resistance in rdxA(+) (Mtz(s)) and also in rdxA (Mtz(r)) H. pylori strains, and that expression of rdxA in Escherichia coli results in equivalent Mtz-induced mutation. A reversion test using defined lac tester strains of E. coli carrying rdxA(+) indicated that CG-to-GC transversions and AT-to-GC transitions are induced more frequently than other base substitutions. Alkaline gel electrophoretic tests showed that Mtz concentrations near or higher than the MIC for growth also caused DNA breakage in H. pylori and in E. coli carrying rdxA(+), suggesting that this damage may account for most of the bactericidal action of Mtz. Coculture of Mtz(s) H. pylori with E. coli (highly resistant to Mtz) in the presence of Mtz did not stimulate forward mutation in E. coli, indicating that the mutagenic and bactericidal products of Mtz metabolism do not diffuse significantly to neighboring (bystander) cells. Our results suggest that the widespread use of Mtz against other pathogens in people chronically infected with H. pylori may stimulate mutation and recombination in H. pylori, thereby speeding host-specific adaptation, the evolution of virulence, and the emergence of resistance against Mtz and other clinically useful antimicrobials.     

Identification of a novel mutation affecting domain V of the 23S rRNA gene in Helicobacter pylori

BACKGROUND: This study analyzes clarithromycin resistance status and 23S rRNA gene mutations in Helicobacter pylori strains from Central Italian patients. MATERIALS AND METHODS: H. pylori strains from 235 dyspeptic patients (205 with no history of clarithromycin exposure and 30 referred for failure of eradication therapy) were tested for clarithromycin resistance by screening agar method and E-test. Resistant strains were analyzed for mutations of the 23S rRNA gene by PCR-RFLP and sequencing. RESULTS: Primary resistance was observed in strains from 43/205 (21%) patients with no history of clarithromycin exposure and secondary resistance in 30/30 (100%) strains from previously treated patients. A single mutant strain was detected in 54/73 (74%) cases, a mixture of one or more mutant(s) plus the wild type in the remaining 19/73 (26%) cases. One 23S rRNA gene mutation (A-->T transversion at nucleotide 2144) in the peptidyltransferase region of domain V was novel. CONCLUSIONS: This study shows: (a) a high prevalence of H. pylori strains with primary or secondary clarithromycin resistance in an urban area of Central Italy; (b) colonization by both mutant and wild-type H. pylori in the same patient; (c) a novel variant of the H. pylori 23S rRNA gene.     

Native supercolonies of unrelated individuals in the invasive Argentine ant

Kinship among group members has long been recognized as a main factor promoting the evolution of sociality and reproductive altruism, yet some ants have an extraordinary social organization, called unicoloniality, whereby individuals mix freely among physically separated nests. This type of social organization is not only a key attribute responsible for the ecological dominance of these ants, but also an evolutionary paradox because relatedness between nestmates is effectively zero. Recently, it has been proposed that, in the Argentine ant, unicoloniality is a derived trait that evolved after its introduction into new habitats. Here we test this basic assumption by conducting a detailed genetic analysis of four native and six introduced populations with five to 15 microsatellite loci and one mitochondrial gene. In contrast to the assumption that native populations consist of family-based colonies with related individuals who are aggressive toward members of other colonies, we found that native populations also form supercolonies, and are effectively unicolonial. Moreover, just as in introduced populations, the relatedness between nestmates is not distinguishable from zero in these native range supercolonies. Genetic differentiation between native supercolonies was very high for both nuclear and mitochondrial markers, indicating extremely limited gene flow between supercolonies. The only important difference between the native and introduced populations was that supercolonies were several orders of magnitude smaller in the native range (25-500 m). This size difference has important consequences for our understanding of the evolution and stability of unicolonial structures because the relatively small size of supercolonies in the native range implies that competition can occur between supercolonies, which can act as a break on the spread of selfish mutants by eliminating supercolonies harboring them.     

Helicobacter acinonychis: genetic and rodent infection studies of a Helicobacter pylori-like gastric pathogen of cheetahs and other big cats

Insights into bacterium-host interactions and genome evolution can emerge from comparisons among related species. Here we studied Helicobacter acinonychis (formerly H. acinonyx), a species closely related to the human gastric pathogen Helicobacter pylori. Two groups of strains were identified by randomly amplified polymorphic DNA fingerprinting and gene sequencing: one group from six cheetahs in a U.S. zoo and two lions in a European circus, and the other group from a tiger and a lion-tiger hybrid in the same circus. PCR and DNA sequencing showed that each strain lacked the cag pathogenicity island and contained a degenerate vacuolating cytotoxin (vacA) gene. Analyses of nine other genes (glmM, recA, hp519, glr, cysS, ppa, flaB, flaA, and atpA) revealed a approximately 2% base substitution difference, on average, between the two H. acinonychis groups and a approximately 8% difference between these genes and their homologs in H. pylori reference strains such as 26695. H. acinonychis derivatives that could chronically infect mice were selected and were found to be capable of persistent mixed infection with certain H. pylori strains. Several variants, due variously to recombination or new mutation, were found after 2 months of mixed infection. H. acinonychis ' modest genetic distance from H. pylori, its ability to infect mice, and its ability to coexist and recombine with certain H. pylori strains in vivo should be useful in studies of Helicobacter infection and virulence mechanisms and studies of genome evolution.     

球形与螺旋形幽门螺杆菌基因表达差异的研究

目的：从基因转录水平了解球形幽门螺杆菌(Helicobacter pylori,Hp)基因表达的变化。方法：幽门螺杆菌标准株NCTC ll637和2株临床分离株,经抗生素—灭滴灵诱导球变。提取等菌量螺旋形和球形Hp的RNA,RT—PCR扩增Hp25种基因,这些基因包括毒力基因(kat、aphA、rdxA、frxA、cysS、ureB glmM、cagA、vacA、fldA、iceAl、hpaA、fliD、napA、oipA、alpAB、babB、hopZ);看家基因(scoB、atpD、g1nA、recA)和功能不明的外膜蛋白基因(hopW、hopX)。扩增产物经分子定量成像系统进行扫描定量。结果和结论：等菌量的球形HpRNA量比螺旋形Hp减少。NCTC ll637、F44和F49菌株的螺旋菌分别有25、20、19个基因RT—PCR阳性,对应的球形菌相应的基因RT—PCR也为阳性。定量分析结果表明：被检测的基因在球形菌中的表达均比螺旋菌降低。提示球形菌致病性的降低与基因的低表达有关。3株球形菌不同基因表达的变化并不完全一致,表达量变化较小且各菌间较一致的是g1mM、oipA、g1nA;表达量变化较大且菌问较一致的基因是：napA、sodB、recA、fliD。     

The rdxA gene plays a more major role than frxA gene mutation in high-level metronidazole resistance of Helicobacter pylori in Taiwan

BACKGROUND: Metronidazole-resistant H. pylori associating with mutations of rdxA or frxA is still a debated topic. This study investigates whether rdxA and frxA mutations of H. pylori accounted for the high MIC value (>/= 64 micro g/ml) of metronidazole (Mtz). MATERIAL AND METHODS: From 126 clinical H. pylori isolates, we examined 14 Mtz-sensitive, 18 Mtz-resistant H. pylori, and eight pairs of Mtz-sensitive and Mtz-resistant colonies simultaneously present within a single gastric biopsy. The paired strains from one single biopsy were proven identical by PCR-RFLP. MICs of Mtz were checked by the E-test and agar dilution method. The mutations of rdxA and frxA sequencing were matched with the Mtz-susceptible ATCC 26695 and J99. RESULTS: There were 89% (16/18) of Mtz-resistant isolates with mutation of RdxA. Half of the 14 Mtz-sensitive strains, all without mutation of RdxA, still contained truncation of FrxA. Within the paired isolates from a single biopsy, rdxA mutation (86%) was more common than frxA mutation (43%) in those isolates with high-level Mtz-resistant H. pylori. RdxA truncation was more prevalent in Mtz-resistant strains with high MICs than in those with low to moderate MICs (75% vs. 20%, p =.01, OR: 12, 95% CI: 1.8-81.7). CONCLUSION: Mutations in the rdxA gene rather than the frxA gene generally determine a high MIC level of Mtz-resistant H. pylori in Taiwan.     

Comparative genomics of Helicobacter pylori strains of China associated with different clinical outcome

In this study, a whole-genome CombiMatrix Custom oligonucleotide tiling microarray with 90,000 probes covering six sequenced Helicobacter pylori (H. pylori) genomes was designed. This microarray was used to compare the genomic profiles of eight unsequenced strains isolated from patients with different gastroduodenal diseases in Heilongjiang province of China. Since significant genomic variation was found among these strains, an additional 76 H. pylori strains associated with different clinical outcomes were isolated from various provinces of China. These strains were tested by polymerase chain reaction to demonstrate this distinction. We identified several highly variable regions in strains associated with gastritis, gastric ulceration, and gastric cancer. These regions are associated with genes involved in the bacterial type I, type II, and type III R-M systems. They were also associated with the virB gene, which lies on the well-studied cag pathogenic island. While previous studies have reported on the diverse genetic characterization of this pathogenic island, in this study, we find that it is conserved in all strains tested by microarray. Moreover, a number of genes involved in the type IV secretion system, which is related to horizontal DNA transfer between H. pylori strains, were identified in the comparative analysis of the strain-specific genes. These findings may provide insight into new biomarkers for the prediction of gastric diseases.     

Roles of FrxA and RdxA nitroreductases of Helicobacter pylori in susceptibility and resistance to metronidazole

The relative importance of the frxA and rdxA nitroreductase genes of Helicobacter pylori in metronidazole (MTZ) susceptibility and resistance has been controversial. Jeong et al. (J. Bacteriol. 182:5082--5090, 2000) had interpreted that Mtz(s) H. pylori were of two types: type I, requiring only inactivation of rdxA to became resistant, and type II, requiring inactivation of both rdxA and frxA to become resistant; frxA inactivation by itself was not sufficient to confer resistance. In contrast, Kwon et al. (Antimicrob. Agents Chemother. 44:2133--2142, 2000) had interpreted that resistance resulted from inactivation either of frxA or rdxA. These two interpretations were tested here. Resistance was defined as efficient colony formation by single cells from diluted cultures rather than as growth responses of more dense inocula on MTZ-containing medium. Tests of three of Kwon's Mtz(s) strains showed that each was type II, requiring inactivation of both rdxA and frxA to become resistant. In additional tests, derivatives of frxA mutant strains recovered from MTZ-containing medium were found to contain new mutations in rdxA, and frxA inactivation slowed MTZ-induced killing of Mtz(s) strains. Northern blot analyses indicated that frxA mRNA, and perhaps also rdxA mRNA, were more abundant in type II than in type I strains. We conclude that development of MTZ resistance in H. pylori requires inactivation of rdxA alone or of both rdxA and frxA, depending on bacterial genotype, but rarely, if ever, inactivation of frxA alone, and that H. pylori strains differ in regulation of nitroreductase gene expression. We suggest that such regulatory differences may be significant functionally during human infection.     

Isogenic variation of Helicobacter pylori strain resulting in heteroresistant antibacterial phenotypes in a single host in vivo

BACKGROUND: Antibiotic-susceptible and -resistant Helicobacter pylori can be present simultaneously in the same host. The aim of this study was to evaluate the genomic diversity of H. pylori strains resulting in heteroresistant antibacterial phenotypes. MATERIALS AND METHODS: Twenty-one pairs of H. pylori strains isolated from the antrum and body displaying heteroresistant antibacterial phenotypes were included. We compared the genotypes of paired-isolates by random arbitrarily primed polymerase chain reaction (PCR), flagella gene PCR-based restriction fragment length polymorphism, and flaA gene sequencing. In metronidazole-heteroresistant isolates, the sequence variation of rdxA and frxA genes was analyzed using phylogenetic analysis. RESULTS: The DNA fingerprinting patterns of the paired isolates revealed that 12 pairs (57.1%) were identical, whereas one pair (3.8%) was different. The remaining eight pairs (38.1%) of isolates showed minor heterogenecity in fingerprinting patterns. In flaA gene sequencing, these identical and similar isolates showed close sequence similarity between the antrum and body, whereas different isolate showed 31 points of different nucleotide sequences. Phylogenetic analysis of the metronidazole-heteroresistant pairs showed consistent genetic relatedness of each paired isolates despite the sequence variation of the rdxA or frxA genes in five pairs (71.4%). CONCLUSIONS: These results suggest that continuing genomic diversities in the same strain may play an important role in modulating the antibiotic-heteroresistant H. pylori in vivo.