Helicobacter pylori protein that binds to and activates platelet specifically reacts with sera of H. pylori-associated chronic immune thrombocytopenia.

Immune thrombocytopenia (ITP) is an acquired autoimmune disorder characterized by antiplatelet antibodies and/or CD8 + T cells, resulting in the destruction of platelets and decreased platelet counts. Helicobacter pylori that persistently colonizes the stomach causes various disorders, including extragastric diseases such as chronic ITP (cITP). Several studies have reported increased platelet counts in H. pylori-infected cITP patients with eradication treatment and also the pathophysiological pathways involving cross-reaction of antibodies against H. pylori with platelets, the modulation of Fcrγ receptors balance and others. We previously reported an immunocomplex pathway comprising H. pylori low-molecular-weight (LMW) antigens, their antibodies, and platelets, involved in the development of H. pylori-associated cITP; however, the LMW antigens were not identified. In the present study, we demonstrated that the H. pylori LMW antigen of the immunocomplex was identified as Lpp20 of outer membrane proteins. Lpp20 could bind to platelets and specifically react with sera of H. pylori-associated cITP patients.

Clinical utility of transthoracic echocardiography for screening abdominal aortic aneurysm: a prospective study in a Japanese population.

BACKGROUND: The aim of the present study was to evaluate the clinical utility of transthoracic echocardiography (TTE) for screening abdominal aortic aneurysm (AAA) and to identify important TTE indices associated with AAA in a Japanese population. METHODS: We prospectively studied 1912 patients who were referred for TTE. AAA was defined as ≥ 30 mm in size. RESULTS: The abdominal aorta was visualized in 95.1% (1818/1912) by TTE. AAA was identified in 2.6% (47/1818). The aortic root size was significantly larger in patients with AAA than those without (36.0 ± 4.1 vs. 31.7 ± 4.2 mm, p < 0.001). The aortic root size had a fair correlation with abdominal aortic size (r = 0.31, p < 0.001). The aortic root size of ≥ 34 mm was predictive of AAA by receiver operating characteristic curve analysis (area under the curve = 0.78, p < 0.001). Multiple logistic regression analysis revealed that aortic root size (Hazard ratio 1.23, p < 0.001) and age (Hazard ratio 1.05, p = 0.013) were the independent predictors of AAA. CONCLUSIONS: The feasibility of the abdominal aortic visualization during TTE was excellent. The aortic root size measured by TTE was the independent predictor of AAA. Screening for AAA during TTE appeared to be useful especially in the older patients with a large (≥34 mm) aortic root.

Autoimmunity and pulmonary hypertension in patients with Graves' disease.

A link between hyperthyroidism and pulmonary hypertension has been reported, but the underlying mechanisms of these two conditions have not been clearly identified. The aim of this study was to determine the clinical correlates of pulmonary hypertension in patients with Graves' disease. Among 50 consecutive patients with Graves' disease referred for echocardiography, 18 patients (36 %) had pulmonary hypertension measured by continuous-wave Doppler echocardiography (pulmonary artery systolic pressure >35 mmHg). The patients with pulmonary hypertension had significantly higher pulmonary vascular resistance (PVR), cardiac output and thyroid-stimulating hormone receptor antibody (TRAb) compared to those without (p < 0.001, p = 0.028 and p < 0.001, respectively). Pulmonary artery systolic pressure had a good correlation with TRAb (r = 0.74, p < 0.001), but was not related to free T4 (r = 0.12, p = 0.419) and free T3 (r = 0.22, p = 0.126). To determine the important variables present in patients with Graves' disease that may be related to pulmonary artery systolic pressure, 4 variables (PVR, cardiac output, TRAb and free T3) were used in the multivariate analysis. In addition to PVR (standard regression coefficient = 0.831, p < 0.001) and cardiac output (standard regression coefficient = 0.592, p < 0.001), TRAb (standard regression coefficient = 0.178, p < 0.001) emerged as a significant variable related to pulmonary artery systolic pressure. Thus, in addition to the effect of thyroid hormone on the cardiovascular system, autoimmune-mediated pulmonary vascular remodeling may play a role in Graves' disease-linked elevated pulmonary artery systolic pressure.

Clinical Application of the DiversiLab Microbial Typing System Using Repetitive Sequence-Based PCR for Characterization of Helicobacter pylori in Japan.

We evaluated the DiversiLab (DL) system with universal primers, a semiautomated repetitive extragenic palindromic sequence-based polymerase chain reaction (PCR) (rep-PCR) system, for the characterization of Helicobacter pylori in Japan. All 135 isolates from Japanese patients with gastric cancer (GC, n = 55) or non-GC (n = 80) were used and subjected to the drug susceptibility examinations (amoxicillin, AMPC; metronidazole, MNZ; and clarithromycin, CAM) by E-test. There were 28 MNZ-resistant (20.7%), 35 CAM-resistant (25.9%), and 16 MNZ/CAM-resistant (11.9%) isolates. DL rep-PCR fingerprinting analysis at the level of 95% similarity revealed five major groups (A-E) and the other including 45 isolates. The occupation rates of GC-derived isolates in groups B (54.2%) and E (58.8%) were higher than in the other groups: A (26.7%), C (28.6%), D (30.0%), and the other (40.0%). Relative higher occupation rates of drug resistants, such as MNZ-, CAM- and double MNZ/CAM-resistant isolates, were observed in groups B (45.8%), C (42.6%), and D (40%). Five of eight GC-derived isolates with MNZ/CAM resistance were significantly assigned to group B (P = 0.0312, χ(2) -test). These results suggest that the isolates classified in group B have a potential to contribute to the development of severe gastric disorders. The DL system, rapid and high sensitive technology, would be widely available in clinical laboratory for pathological and epidemiological analyses even in H. pylori.

[Molecular-pathological analysis of Helicobacter pylori-associated disorders and clinical laboratory testing].

Helicobacter pylori (H. pylori) infection causes gastro-duodenal diseases and a wide variety of non gastrointestinal tract conditions, such as idiopathic thrombocytopenic purpura (ITP). Many reports have provided robust evidence for understanding the pathogenesis of H. pylori. However, its cell division process is little known. H. pylori exhibits marked genetic diversity to survive under various stress conditions, leading to the emergence of a variety of clones with novel characteristics. In this report, we briefly summarize our results. H. pylori urease is essential to live in a low-pH environment. Bacterial motility and urea taxisis are also important features for persistent infection. The urease is controlled in response to the pH via post translational regulation. Genetic rearrangement occurs during persistent infection of an individual's stomach, which results in the appearance of a variety of new strains with novel characteristics. The cdrA (cell division-related gene A)--dysfunctional strain had acquired such novel characteristics: increased viability, long-term survival, and tolerance to antibiotics. Furthermore, colonization by a cdrA-dysfunctional strain results in decreased IL-8 production and, hence, attenuates the host's immunity to cause persistent infection. Among the factors involved in the bacterium-host interaction and pathogenesis, we describe the H. pylori CagA, BabA, SabA, and SOD. We discovered two H. pylori phages, including a new type of spherical phage, which cannot be classified into any existing virus category. The phages probably contribute to the evolution and pathogenesis of H. pylori. Eradication therapy covered by health insurance was approved for H. pylori-associated ITP. We reported an extra mechanism, the immunocomplex model (platelets, bacterial molecules, and anti H. pylori antibodies), in the development of H. pylori associated ITP. On the other hand, increased cases of unsuccessful eradication therapy are due to the increased occurrence of drug-resistant H. pylori. Non-antibiotic substances with anti-H. pylori activity are attracting much attention.

Superoxide dismutase activity of Helicobacter pylori per se from 158 clinical isolates and the characteristics.

We investigated the correlation between the SOD activity of Helicobacter pylori (H. pylori) and gastroduodenal diseases and the characteristics of strains exposed to oxidative stress. Two sequenced strains, 26695 and J99, and clinical isolates from 156 Japanese patients with gastroduodenal diseases such as gastric cancer (n= 59) and non-cancer (n= 97) were used. SOD activities of all 158 isolates were measured and were divided into three groups: high-SOD activity (>0.22, n= 2), moderate-SOD activity (0.15≦≦0.22, n= 16) and low-SOD activity (<0.15, n= 140). Expressions of H. pylori Fe-SOD were examined by western blotting with anti-H. pylori Fe-SOD antibody prepared inhouse, and the profiles of Fe-SOD activity were investigated by zymogram with activity staining in native-PAGE. The characteristics of strains from high-SOD and low-SOD groups were examined under oxidative stress by paraquat. The average of H. pylori SOD activity was significantly higher in the cancer group than in the non-cancer group (P < 0.05). However, irrespective of SOD activity level, the amount of Fe-SOD expressed was variable among individual strains. Zymogram revealed a single band in moderate-SOD and low-SOD strains, but multiple bands in high-SOD strains were observed. These bands were confirmed as H. pylori Fe-SOD. Under oxidative stress with paraquat, low-SOD strains were drastically eliminated without inducible SOD activity; however, high-SOD strains were still viable with increased SOD activity. This study is the first to exhibit the characteristics of high-SOD activity strains representing multiple bands in zymograms and the correlation between H. pylori SOD activity and gastric cancer.

[Molecular identification of human Diphyllobothrium nihonkaiense using mitochondrial cytochrome c oxidase subunit 1 (cox1) gene sequence].

Identification of Diphyllobothrium species has been carried out based on their morphology, especially sexual organs. In addition to these criteria, PCR-based identification methods have been developed recently. A 20 year-old Japanese living in Kochi Prefecture passed tapeworm. He was successfully treated with single dose of gastrografin. We examined the morphologic features of the proglottids and eggs using histology and scanning electron microscope. We also analyzed mitochondrial cytochrome c oxidase subunit 1 (cox1) gene of the proglottids. The causative tapeworm species was identified as D. nihonkaiense based on the results of morphologic features and genetic analysis. We discussed the advantage of PCR-based identification methods of Diphyllobothrium species using cox1 sequence in the clinical laboratory.

End-of-season outbreaks of nosocomial influenza caused by waning vaccine immunity.

The waning of vaccine protection may be responsible for outbreaks toward the end of the influenza season. Three of five outbreaks occurred at the beginning of April following an interval of >100 days from the date of vaccination; the reported index case was a nurse or office worker, and >50% of those affected were healthcare workers. The results are consistent with intra-seasonal waning of vaccine immunity that resulted in outbreaks at the end of season.

Prevalence and characteristics of methicillin-resistant Staphylococcus aureus colonization among healthcare professionals in a university hospital in Japan.

BACKGROUND: Asymptomatic carriers of methicillin-resistant Staphylococcus aureus (MRSA) are important sources of nosocomial transmission. MRSA may be transmitted from hospitalized patients to healthcare professionals and vice versa. METHODS: The prevalence of MRSA colonization among forty-five healthcare professionals in a Japanese hospital was determined by performing surveillance cultures to identify unrecognized carriers of MRSA. All MRSA isolates were evaluated using multilocus sequence typing (MLST) to identify the transmission routes. RESULTS: The proportion of MRSA colonization was significantly higher in healthcare professionals (11.1%) than in community residents (0.72%; P < 0.0001) or admission case (2.5%; P = 0.018). MLST analysis revealed that both the ST8 and ST764 strains were identified in residents, patients, and healthcare professionals. MRSA colonization was more frequently observed among physicians (4/13; 31%) than nurses (1/32; 3%) (P = 0.020). CONCLUSION: Multilocus sequence typing results suggest that ST8 and ST764 are involved in the occurrence of nosocomial MRSA infections. These findings emphasize the necessity for the effective education of physicians to prevent MRSA transmissions.

Transmission of methicillin-resistant Staphylococcus aureus in an acute care hospital in Japan.

BACKGROUND: Asymptomatic carriers of methicillin-resistant Staphylococcus aureus (MRSA) are important sources of nosocomial transmission. However, the route of transmission of MRSA is not completely understood. The purpose of this study was to calculate MRSA transmission rates in a hospital with a high MRSA infection/colonization density and inadequate hand hygiene compliance. METHODS: The prevalence of MRSA colonization among 157 patients at the time of admission to and discharge from a medical school hospital in Japan was determined by performing surveillance cultures. All MRSA isolates were evaluated using multilocus sequence typing (MLST) to identify the transmission routes. RESULTS: Methicillin-resistant S. aureus was prevalent in 1.9% of our study population. MRSA was acquired during hospitalization at a rate of 4.0/1000 patient-days. At discharge, 5.1% of the patients exhibited MRSA colonization; this was significantly higher than the prevalence noted upon admission (P < 0.001). MLST documented three possible nosocomial transmission events. MRSA colonization was detected using surveillance cultures prior to being identified by conventional, clinically oriented examinations. CONCLUSIONS: Multilocus sequence typing results suggested that patients who were colonized with MRSA acquired it during hospitalization. These results reinforce the importance of infection control for preventing nosocomial MRSA transmission in hospitalized patients.

Prevalence and characteristics of methicillin-resistant Staphylococcus aureus in community residents of Japan.

BACKGROUND: To implement effective precautions to avoid methicillin-resistant Staphylococcus aureus (MRSA) nosocomial infections, it is important to clarify when, how, and from whom MRSA was transmitted to the patients. However, MRSA strains obtained from outpatient population were not analyzed, and the transmission routes of MRSA in the community are not completely understood. The purpose of this study was to clarify whether MRSA is spreading in community settings or whether MRSA transmission still occurs only in healthcare institutions. METHODS: Surveillance cultures of 1274 residents living in a community were performed in two different areas, Kochi and Osaka prefectures of Japan. All isolated MRSA strains were evaluated using multilocus sequence typing (MLST) to clarify the transmission routes of MRSA. The results were compared with those of inpatients. Moreover, written questionnaires and medical records were analyzed. RESULTS: Analysis of surveillance cultures from residents living in the community in Japan revealed an MRSA colonization rate of 0.94%. The proportion of MRSA to S. aureus colonization was 2.6% in the 310 residents, which was significantly lower than in the 393 hospitalized patients (63.1%; P < .0001). MRSA strains in residents are different from the endemic strains in the hospitalized patients. Previous hospital admission is a risk factor for MRSA infection of the endemic strain in hospital. CONCLUSIONS: Methicillin-resistant Staphylococcus aureus colonization in community setting is rare in Japan. MLST results suggest that some MRSA strains are moving to the community through previous hospital admissions; however, MRSA is not spreading in community settings.

Helicobacter pylori CagA status associated with gastric cancer incidence rate variability in Costa Rican regions.

BACKGROUND: We evaluated several risk factors for gastric cancer in Costa Rican regions having contrasting gastric cancer incidence rates, despite the small dimensions of the country. METHODS: A total of 180 dyspeptic patients were classified into two groups according to the gastric cancer incidence (GCI) rate in their Costa Rican region: group A, with a high GCI rate (n = 91) and group B, with a low GCI rate (n = 89). Helicobacter pylori infection was detected by rapid urease test, Gram staining, and histological observation. Antral and corpus specimens were obtained to assess the grade of inflammation, topography of gastritis, gastric atrophy, and intestinal metaplasia by histological examination. Serum CagA antibody was measured by an antigen-specific enzyme-linked immunosorbent assay. RESULTS: There was no significant difference in H. pylori prevalence between groups A (73%) and B (63%); however, serum CagA antibody was more frequently detected in group A (79%) than in group B (54%) [P = 0.02; odds ratio (OR), 2.68]. Among patients under 60 years of age, serum CagA antibody was even more frequently detected in group A (81%) than in group B (49%) (P < 0.01; OR, 4.50). The prevalence of corpus-predominant gastritis, atrophic gastritis, and moderate/severe grades of neutrophilic infiltration was higher in serum CagA antibody-positive patients than in CagA antibody-negative patients (P = 0.003, 0.04, and 0.002, respectively). CONCLUSIONS: Infection with H. pylori possessing the cagA gene is associated with the development of severe gastric damage such as gastric atrophy, leading to gastric cancer, and probably influences the differences in GCI between Costa Rican regions.

Genotyping Giardia intestinalis by Using DNA Extracted from Long-Term Preserved Human Specimens Stained with Chlorazol Black E.

Giardia intestinalis is a parasitic protozoan that causes diarrhea and abdominal pain in humans. Studies of the Giardia genotypes are thought to be important for understanding their infection routes and prevalence. However, few have reported pathogen genotyping in human giardiasis cases in Japan. In this study, we genotyped G. intestinalis by using DNA extracted from chlorazol black E-stained fecal smears from patients. The triosephosphate isomerase gene was amplified from 21 (91.3%) of 23 human fecal samples. Twelve (52.2%) of pathogens detected were of the genotype A, and 9 (39.1%) of the genotype B. A restriction fragment length polymorphism analysis showed that all genotype A found in the present study were of the genotype AI, which were presumed to be zoonotic. The source of Giardia infections was unclear in the present study. However, patients' histories of international travel appeared not to be associated with the Giardia genotypes. Thus, most cases were thought to be acquired sporadically and domestically.

Intrinsic characteristics of Min proteins on the cell division of Helicobacter pylori.

Helicobacter pylori divides in the human stomach resulting in persistent infections and causing various disorders. Bacterial cell division is precisely coordinated by many molecules, including FtsZ and Min proteins. However, the role of Min proteins in H. pylori division is poorly understood. We investigated the functional characteristics of Min proteins in wild-type HPK5 and five HPK5-derivative mutants using morphological and genetic approaches. All mutants showed a filamentous shape. However, the bacterial cell growth and viability of three single-gene mutants (minC, minD, minE) were similar to that of the wild-type. The coccoid form number was lowest in the minE-disruptant, indicating that MinE contributes to the coccoid form conversion during the stationary phase. Immunofluorescence microscopic observations showed that FtsZ was dispersedly distributed throughout the bacterial cell irrespective of nucleoid position in only minD-disruptants, indicating that MinD is involved in the nucleoid occlusion system. A chase assay demonstrated that MinC loss suppressed FtsZ-degradation, indicating that FtsZ degrades in a MinC-dependent manner. Molecular interactions between FtsZ and Min proteins were confirmed by immunoprecipitation (IP)-western blotting (WB), suggesting the functional cooperation of these molecules during bacterial cell division. This study describes the intrinsic characteristics of Min proteins and provides new insights into H. pylori cell division.

Hypoxia-inducible factor-1alpha is involved in the attenuation of experimentally induced rat glomerulonephritis.

BACKGROUND/AIM: Among various kidney disease models, there are few rat glomerulonephritis (GN) models that develop in a short time, and with mainly glomerular lesions. Hypoxia-inducible factor (HIF)-1alpha is a transcriptional factor that induces genes supporting cell survival, but the involvement of HIF-1alpha in attenuating the progression of GN remains to be elucidated. We developed a new model of rat GN by coadministration of angiotensin II (AII) with Habu snake venom (HV) and investigated whether HIF-1alpha is involved in renal protection. METHODS: Male Wistar rats were unilaterally nephrectomized on day 1, and divided into 4 groups on day 0; N group (no treatment), HV group, A group (AII), and H+A group (HV and AII). To preinduce HIF-1alpha, cobalt chloride (CoCl2) was injected twice before injections of HV and AII in 11 rats. RESULTS: GN was detected only in the H+A group; observed first on day 2 and aggravated thereafter. HIF-1alpha was expressed in the glomeruli and renal tubules in the A and H+A groups. In the H+A group, GN was remarkably reduced by CoCl2 pretreatment (44.9 to 12.2%, p < 0.01). CONCLUSION: Both HV and AII were critical for the development of GN, and HIF-1alpha remarkably attenuated the progression of GN.

IL-5-Induced Eosinophils Suppress the Growth of Leishmania amazonensis In Vivo and Kill Promastigotes In Vitro in Response to Either IL-4 or IFN-gamma.

In IL-5 transgenic mice (C3H/HeN-TgN(IL-5)-Imeg), in which 50% of peripheral blood leukocytes are eosinophils, the development of infection by Leishmania amazonensis was clearly suppressed. To determine mechanistically how this protozoan parasite is killed, we performed in vitro killing experiments. Either IL-4 or IFN-gamma effectively stimulated eosinophils to kill Leishmania amazonensis promastigotes, and most of the killing was inhibited by catalase but not by the NO inhibitor L-N5-(1-iminoethyl)-ornithine, suggesting that hydrogen peroxide is responsible for the killing of L. amazonensis by eosinophils. There was no significant degranulation of eosinophils in the culture, because eosinophil peroxidase was not detected in culture supernatants when L. amazonensis promastigotes were killed by activated eosinophils. Such resistance was also observed in BALB/c mice, which are highly susceptible to L. amazonensis. Expression plasmids for IL-4, IL-5, and IFN-gamma were transferred into muscle by electroporation in vivo starting 1 week before infection. Expression plasmid for IL-5 was most effective in slowing the development of infection among three expression plasmids. Expression plasmid for IL-4 was slightly effective and that for IFN-gamma had no effect on the progress of disease. These results suggest that IL-5 gene transfer into muscle by electroporation is useful as a supplementary protection method against L. amazonensis infection.

[Incidence and clinical features of Giardia lamblia].

To determine the incidence and clinical features of Giardia lamblia infection, we studied 1790 patients at Kochi Medical School Hospital from April 1998 to July 2001. Fecal samples were examined microscopically by the direct smear method, direct immunofluorescent assay and by Kohn's one-step staining for G. lamblia cysts. Cysts of G. lamblia were found in 17 of 1,790(0.95%) stool samples, indicating that G. lamblia infection is not rare in Kochi. The most characteristic feature was that G. lamblia-positive cases were more frequent in the advanced age group(41-79 years old) and most of the subjects (except 2 cases) with G. lamblia had no history of traveling overseas. Four subjects had symptoms related to G. lamblia infection. Thus, more attention should be given to parasitic infections in laboratory stool examinations in order to detect cyst carriers as potential sources of infection.

Natural products and food components with anti-Helicobacter pylori activities.

The bacterial pathogen Helicobacter pylori (H. pylori) colonizes in over half of the world's population. H. pylori that establishes life-long infection in the stomach is definitely associated with gastro-duodenal diseases and a wide variety of non-gastrointestinal tract conditions such as immune thrombocytopenia. Triple therapy which consists of a proton pump inhibitor and combinations of two antibiotics (amoxicillin, clarithromycin or amoxicillin, metronidazol) is commonly used for H. pylori eradication. Recently, the occurrence of drug-resistant H. pylori and the adverse effect of antibiotics have severely weakened eradication therapy. Generally antibiotics induce the disturbance of human gastrointestinal microflora. Furthermore, there are inappropriate cases of triple therapy such as allergy to antibiotics, severe complications (liver and/or kidney dysfunction), the aged and people who reject the triple therapy. These prompt us to seek alterative agents instead of antibiotics and to develop more effective and safe therapy with these agents. The combination of these agents actually may result in lower a dose of antibiotics. There are many reports world-wide that non-antibiotic substances from natural products potentially have an anti-H. pylori agent. We briefly review the constituents derived from nature that fight against H. pylori in the literature with our studies.