Helicobacter pylori is a gram-negative, microaerophilic, and spiral-shaped bacterium and causes gastrointestinal diseases in human. IL-1β is a representative cytokine produced in innate immune cells and is considered to be a key factor in the development of gastrointestinal malignancies. However, the mechanism of IL-1β production by neutrophils during H. pylori infection is still unknown. We designed this study to identify host and bacterial factors involved in regulation of H. pylori-induced IL-1β production in neutrophils. We found that H. pylori-induced IL-1β production is abolished in NLRP3-, ASC-, and caspase-1/11-deficient neutrophils, suggesting essential role for NLRP3 inflammasome in IL-1β response against H. pylori. Host TLR2, but not TLR4 and Nod2, was also required for transcription of NLRP3 and IL-1β as well as secretion of IL-1β. H. pylori lacking cagL, a key component of the type IV secretion system (T4SS), induced less IL-1β production in neutrophils than did its isogenic WT strain, whereas vacA and ureA were dispensable. Moreover, T4SS was involved in caspase-1 activation and IL-1β maturation in H. pylori-infected neutrophils. We also found that FlaA is essential for H. pylori-mediated IL-1β production in neutrophils, but not dendritic cells. TLR5 and NLRC4 were not required for H. pylori-induced IL-1β production in neutrophils. Instead, bacterial motility is essential for the production of IL-1β in response to H. pylori. In conclusion, our study shows that host TLR2 and NLRP3 inflammasome and bacterial T4SS and motility are essential factors for IL-1β production by neutrophils in response to H. pylori.
Two virulence factors of Helicobacter pylori , cagA and vacA , have been known to play a role in the development of severe gastric symptoms. However, they are not always associated with peptic ulcer or gastric cancer. To predict the disease outcome more accurately, it is necessary to understand the risk of severe symptoms linked to other virulence factors. Several other virulence factors of H. pylori have also been reported to be associated with disease outcomes, although there are many controversial descriptions. H. pylori isolates from Koreans may be useful in evaluating the relevance of other virulence factors to clinical symptoms of gastric diseases because the majority of Koreans are infected by toxigenic strains of H. pylori bearing cagA and vacA . In this study, a total of 116 H. pylori strains from Korean patients with chronic gastritis, peptic ulcers, and gastric cancers were genotyped. The presence of virulence factors vacAs1c , alpA , babA2 , hopZ , and the extremely strong vacuolating toxin was found to contribute significantly to the development of severe gastric symptoms. The genotype combination vacAs1c / alpA / babA2 was the most predictable determinant for the development of severe symptoms, and the presence of babA2 was found to be the most critical factor. This study provides important information on the virulence factors that contribute to the development of severe gastric symptoms and will assist in predicting clinical disease outcomes due to H. pylori infection.
Mycobacterium intracellulare is a leading cause of nontuberculous mycobacterial pulmonary disease, with a rapidly increasing prevalence worldwide. This bacterium, commonly distributed in soil and water, is known to be transmitted through the environment rather than between people. Therefore, it is imperative to establish distinguishable genotyping methods to understand the clinical outcome, disease relapses, and epidemiology. Therefore, in this study, representative band-based genotyping methods were performed using M. intracellualre clinical isolates, and their Hunter–Gaston discriminatory index (HGDI) was 0.947, 0.994, and 1 for variable number tandem repetition (VNTR), VNTR-mycobacterial interspersed repetitive units, pulsed field gel electrophoresis, and repetitive sequence based-PCR, respectively. Although VNTR showed relatively low HGDI, co-infection with other M. intracellualre strains could be determined by loci showing allele diversity from 0 to 0.69. Additionally, genetic distance of clinical isolates from Gyeongnam/Korea, and other regions/countries were visualized by minimum spanning tree (MST) using the globally available VNTR profiles. The results of MST revealed that M. intracellulare isolated from patients in Gyeongnam/Korea had specific VNTR genotypes, which may be evidence of the geographic distribution of M. intracellulare specific genotypes. The comparative results of genotyping techniques and geographical characteristics in this study may provide fundamental information for the epidemiology of M. intracellulare.
Background. Nontuberculous mycobacteria (NTM) are widely present in environments, such as soil and water, and have recently been recognized as important pathogenic bacteria. The incidence of NTM-related infections is steadily increasing. As the diagnosis and treatment of NTM infection should be distinguished from tuberculosis, and the treatment should be specific to the species of NTM acquired, accurate species identification is required. Methods. In this study, two-step multiplex PCR (mPCR) and multigene sequence-based analysis were used to accurately identify NTM species in 320 clinical isolates from Gyeongsang National University Hospital (GNUH). In particular, major mycobacterial strains with a high isolation frequency as well as coinfections with multiple species were diagnosed through two-step mPCR. Multigene sequencing was performed to accurately identify other NTM species not detected by mPCR. Variable regions of the genes 16S rRNA, rpoB, hsp65, and 16S-23S rRNA internal transcribed spacer were included in the analysis. Results. Two-step mPCR identified 234 (73.1%) cases of M. intracellulare, 26 (8.1%) cases of M. avium subsp. avium, and 13 (4.1%) cases of M. avium subsp. hominissuis infection. Additionally, 9 (2.8%) M. fortuitum, 9 (2.8%) M. massiliense, 2 (0.6%) M. abscessus, and 4 (1.2%) M. kansasii isolates were identified. Coinfection was identified in 7 (2.2%) samples. The sixteen samples not classified by two-step mPCR included 6 (1.9%) cases of M. chimaera, 4 (1.3%) M. gordonae, 1 (0.3%) M. colombiense, 1 (0.3%) M. mageritense, and 1 (0.3%) M. persicum identified by sequence analysis. Conclusions. The results of this study suggest a strategy for rapid detection and accurate identification of species using two-step mPCR and multigene sequence-based analysis. To the best of our knowledge, this study is the first to report the identification of NTM species isolated from patients in Gyeongnam/Korea.
Johne's disease (JD) is a chronic granulomatous enteritis of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP), which induces persistent diarrhea and cachexia. JD causes huge economic losses to the dairy industry due to reduced milk production and premature culling. Infected animals excrete MAP via feces during the prolonged subclinical stage without exhibiting any clinical signs. Therefore, accurate detection of subclinical stage animals is crucial for successful eradication of JD in the herd. In the current study, we analyzed serum samples of MAP-infected and non-infected cattle to identify potential biomarker candidates. First, we identified 12 differentially expressed serum proteins in subclinical and clinical shedder groups compared to the healthy control group. Second, we conducted ELISA for three selected biomarkers (alpha-2-macroglobulin (A2M), alpha-1-beta glycoprotein, and transthyretin) and compared their diagnostic performance with that of two commercial ELISA diagnostic kits. Serum A2M levels were significantly higher in the MAP-exposed, subclinical shedder, subclinical non-shedder, and clinical shedder groups than in the healthy control group, suggesting its possible use as a diagnostic biomarker for MAP infection. Furthermore, A2M demonstrated a sensitivity of 90.4%, and a specificity of 100% while the two commercial ELISA kits demonstrated a sensitivity of 67.83 and 73.04% and a specificity of 100%, respectively. In conclusion, our results suggest that measuring A2M by ELISA can be used as a diagnostic tool to detect MAP infection, considerably improving the detection rate of subclinical shedders and MAP-exposed animals that are undetectable using current diagnostic tools.
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