RdxA Diversity and Mutations Associated with Metronidazole Resistance of Helicobacter pylori.

Metronidazole (MNZ) is administered as first-line antibiotic for Helicobacter pylori eradication therapy; however, increasing resistance to MNZ impaired the efficacy. Increasing the dose of MNZ was recommended to overcome low-level resistance, but it was difficult to determine MNZ resistance level simply based on the rdxA gene mutation. In this study, the rdxA sequences of 511 clinical H. pylori strains were analyzed to assess the genotypes associated with MNZ resistance. We observed that the prevalences of rdxA sequences with missense, nonsense, and frameshift mutations were 70.25, 11.35, and 17.03%, respectively. Regarding the amino acid substitutions, T31E, H53R, D59N, L62V, S88P, G98S/N, R131K, and V172I were present in most strains regardless of the resistance phenotype. The correlation analysis showed R16H/C, Y47C, A67V/T, and V204I substitutions were associated with MNZ resistance. The mutation resulting in RdxA truncation was observed in 36.29% of the resistant strains, and 83.45% of these strains displayed high-level MNZ resistance (MIC > 256 µg/mL). Moreover, all strains with truncated mutation positions before amino acid 70 expressed high-level MNZ resistance. Our results indicated that most amino acid mutations probably contributed to the sequence diversity of RdxA, while R16H/C, Y47C, A67V/T, and V204I were potentially helpful to identify resistant strains. Although it was difficult to determine the mutations associated with MNZ resistance, the prediction of high-level resistance based on truncated characteristics of RdxA might be an important approach, which can effectively avoid H. pylori eradication therapy with unreasonable of MNZ dose increases for patients with high-level drug resistance. IMPORTANCE The increasing resistance to metronidazole impaired the efficacy of Helicobacter pylori eradication, and increasing the dose of metronidazole was recommended to overcome low-level resistance. For patients infected with highly resistant strains, the current empirical treatments, which generally used metronidazole in double doses or more, appeared impossibly to overcome the resistance and would only increase the incidence of adverse effects. Our results indicated that high-level metronidazole resistance was predominant, and almost half of the patients with high-level drug resistance could avoid usage of metronidazole based on the truncated mutations of RdxA sequences, which can effectively avoid H. pylori eradication therapy with unreasonable increases in the metronidazole dose.

DNA starvation/stationary phase protection protein of Helicobacter pylori as a potential immunodominant antigen for infection detection.

BACKGROUND: Application of chicken egg yolk immunoglobulin Y (IgY) for Helicobacter pylori (H. pylori, HP) has gained much interest in recent years. Comparing with for treatment, IgY may be more advantageous when used for H. pylori detection. METHODS: Nine strains of H. pylori with different genetic backgrounds were inactivated and used to immunize hens, respectively, for the preparation of polyclonal anti-H. pylori immunoglobulin Y (anti-HP IgY). The proteins of H. pylori with reactivity to anti-HP IgY were detected by Western Blot. The five protein bands that can be well recognized by anti-HP IgY of each group, and were prevalent in all nine strains were excised from SDS-PAGE gel, digested and identified by Nano-HPLC-MS/MS analysis. The potential of these identified proteins as antigen detection targets was then assessed by sequence analysis. RESULTS: Anti-HP IgY derived from each group of specific strain immunized hens can recognize self-strain and non-self-strain antigens well. Five immunodominant antigens were identified as chaperonin GroEL, flagellin A, urease subunit alpha, peroxiredoxin and DNA starvation/stationary phase protection protein. Sequences analysis showed that both peroxiredoxin and DNA starvation/stationary phase protection protein were present in all 1000 strains of H. pylori queried, and the amino acid sequences were highly conserved. The highest sequence consistency between the DNA starvation/stationary phase protection protein of H. pylori and non-Helicobacter organisms was 52.59%, and the consistent sites were scattered and there was no continuous long fragment consensus sequence. CONCLUSION: DNA starvation/stationary phase protection protein was identified as an immunodominant antigen of H. pylori and sequence analysis indicated that it could serve as a potential antigen target for the diagnosis of H. pylori infection.

Distribution characteristics of the sabA, hofC, homA, homB and frpB-4 genes of Helicobacter pylori in different regions of China.

BACKGROUND: Helicobacter pylori (H. pylori) encodes numerous outer membrane proteins (OMPs), with considerable geographic heterogeneity and related to different clinical outcomes. This study aimed to investigate the distribution characteristics of five important OMP genes (sabA, hofC, homA, homB and frpB-4) in different regions of China. MATERIALS AND METHOD: A total of 266 strains were isolated from 348 stomach biopsy specimens in Shandong, Guangxi, Heilongjiang, Hunan, and Qinghai provinces. The presence of sabA, hofC, homA, homB and frpB-4 gene was detected by polymerase chain reaction (PCR) from H. pylori genomic DNA. RESULTS: Among the strains in five regions, the prevalence of frpB-4 was 100% and that of hofC was 97.7%. The prevalence of homB in the isolates from Qinghai (45.5%) was significantly lower than that in Shandong (75.3%), Guangxi (76.9%) and Hunan (69.6%) (P<0.05). The frequency of homA in Shandong (30.1%) was significantly lower than in Guangxi (57.7%) and Qinghai (63.6%) (P<0.05). The prevalence of the sabA gene in Shandong, Guangxi, Heilongjiang, Hunan and Qinghai provinces was 21.9%, 59.7%, 45.9%, 52.2%, and 18.2%, respectively (P<0.05). The sabA "on" status was significantly more frequent in isolates from Guangxi (46.8%), Heilongjiang (37.8%), and Hunan (47.8%) than Qinghai (3.0%) (P<0.05). The presence of homA and sabA genes may be negatively correlated with the development of gastritis. There was no significant association between the frpB-4, hofC, homB gene and clinical outcomes. CONCLUSION: The prevalence of homA, homB, and sabA genes and the sabA "on" or "off" status have significant geographical differences among five provinces in China. The presence of homA and sabA genes may be protective factors of gastritis.

Long-term investigation of environmental radioactivity levels and public health around the Qinshan Nuclear Power Plant, China.

To evaluate the impact of the Qinshan Nuclear Power Plant (Qinshan NPP) in normal operation on the surrounding environment and population, the radioactivity levels of drinking water and the ambient environment, as well as the residents' cancer incidence, were continuously monitored for a period of 9 years (2012-2020). All of the gross α and ß radioactivity concentrations in drinking water were less than the WHO recommended values (0.5 Bq/L for gross α and 1 Bq/L for gross ß). The results of ambient environment accumulated dose monitored by thermoluminescent dosimeters (TLDs) indicated that the ambient environment radioactive level around the Qinshan NPP is consistently at natural background radiation levels. The age-dependent annual effective doses due to the ingestion of tap water or exposure to the outdoor ambient environment are lower than the reference dose of 0.1 mSv/year. The corresponding excess risks are at relatively low levels. Thus, the consumption of drinking water and outdoor activities are not expected to give rise to any detectable adverse effects on the health of the public around the Qinshan NPP. For all cancers combined, the age-standardized incidence rate by the Chinese 2000 standard population of the inhabitants living around Qinshan NPP is consistent with that of Zhejiang Province as a whole. Based on current radiation risk estimates, radiation exposure is not a plausible explanation for any excess cancers observed in the vicinity of the Qinshan NPP.

Diversity of 3' variable region of cagA gene in Helicobacter pylori strains isolated from Chinese population.

BACKGROUND: The cytotoxin-associated gene A (cagA) is one of the most important virulence factors of Helicobacter pylori (H. pylori). There is a highly polymorphic Glu-Pro-Ile-Tyr-Ala (EPIYA) repeat region in the C-terminal of CagA protein. This repeat region is thought to play an important role in the pathogenesis of gastrointestinal diseases. The aim of this study was to investigate the diversity of cagA 3' variable region and the amino acid polymorphisms in the EPIYA segments of the CagA C-terminal region of H. pylori, and their association with gastroduodenal diseases. METHODS: A total of 515 H. pylori strains from patients in 14 different geographical regions of China were collected. The genomic DNA from each strain was extracted and the cagA 3' variable region was amplified by polymerase chain reaction (PCR). The PCR products were sequenced and analyzed using MEGA 7.0 software. RESULTS: A total of 503 (97.7%) H. pylori strains were cagA-positive and 1,587 EPIYA motifs were identified, including 12 types of EPIYA or EPIYA-like sequences. In addition to the four reported major segments, several rare segments (e.g., B', B″ and D') were defined and 20 different sequence types (e.g., ABD, ABC) were found in our study. A total of 481 (95.6%) strains carried the East Asian type CagA, and the ABD subtypes were most prevalent (82.1%). Only 22 strains carried the Western type CagA, which included AC, ABC, ABCC and ABCCCC subtypes. The CagA-ABD subtype had statistical difference in different geographical regions (P = 0.006). There were seven amino acid polymorphisms in the sequences surrounding the EPIYA motifs, among which amino acids 893 and 894 had a statistical difference with gastric cancer (P = 0.004). CONCLUSIONS: In this study, 503 CagA sequences were studied and analyzed in depth. In Chinese population, most H. pylori strains were of the CagA-ABD subtype and its presence was associated with gastroduodenal diseases. Amino acid polymorphisms at residues 893 and 894 flanking the EPIYA motifs had a statistically significant association with gastric cancer.

Global Deletion of TSPO Does Not Affect the Viability and Gene Expression Profile.

Translocator Protein (18kDa, TSPO) is a mitochondrial outer membrane transmembrane protein. Its expression is elevated during inflammation and injury. However, the function of TSPO in vivo is still controversial. Here, we constructed a TSPO global knockout (KO) mouse with a Cre-LoxP system that abolished TSPO protein expression in all tissues and showed normal phenotypes in the physiological condition. The birth rates of TSPO heterozygote (Het) x Het or KO x KO breeding were consistent with Mendel's Law, suggesting a normal viability of TSPO KO mice at birth. RNA-seq analysis showed no significant difference in the gene expression profile of lung tissues from TSPO KO mice compared with wild type mice, including the genes associated with bronchial alveoli immune homeostasis. The alveolar macrophage population was not affected by TSPO deletion in the physiological condition. Our findings contradict the results of Papadopoulos, but confirmed Selvaraj's findings. This study confirms TSPO deficiency does not affect viability and bronchial alveolar immune homeostasis.