We evaluated the utility of 5 commercial enzyme-linked immunosorbent assay (ELISA) kits for detecting antibodies to avian influenza viruses. The sensitivities and specificities of the ELISA kits were compared with those of the agar gel precipitation (AGP) and hemagglutination-inhibition (HI) tests. The results suggest that some ELISA kits might not be suitable for monitoring during the early stages of avian influenza virus infections. Therefore, ELISA kits should only be used in conjunction with a profound knowledge about monitoring of avian influenza.

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馬や人間にハリセファロブス感染症を引き起こすPanagrolaimida科の線虫Halicephalobus gingivalisを土壌線虫群から検出するため、DNAの抽出方法ならびにPCR法による簡易検出方法を検討した。キアゲン社のQIAamp DNA mini kitを用いて抽出したDNAとHalicephalobus属に特異的なプライマーセットを用いたPCR法によって、自活性線虫約3,000頭の中に含まれる１頭のH. gingivalisを検出することができた。

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われわれは, ヒト正常胎児肺由来の繊維芽細胞株WI-38とヒト肺扁平上皮癌由来細胞株SQ-5を用い, 非貧食細胞内へのヒト型結核菌, ウシ型結核菌およびBCGの侵入能とその内部での生存能を調べた。 ヒト型結核菌ErdmanおよびH37Rvの生菌は, ウシ型結核菌Ravenel, ヒト型結核菌H37Ra, BCG東京株およびBCGパスツール株の生菌より効率良くWI-38に侵入した。生菌のWI-38への侵入能は, Erdman≧H37Rv>BCGパスツール株≒M. bovis Ravenel≒BCG東京株>H37Raの順であった。同様の侵入能は, SQ-5細胞を用いても得られた。生菌とは対照的に, 熱処理死菌はWI-38細胞に取り込まれなかったが, SQ-5細胞には取り込まれた。これらの結果とラテックスビーズの取り込みの結果は, WI-38にはないがSQ-5細胞は貧食能を持っていることを示唆している。H37Rvは, H37RaおよびBCG東京株と比較しWI-38細胞内で最も多く増殖した。このことは, 非貧食細胞内への侵入能とその内部での生存能は, 弱毒結核菌より強毒結核菌の能動的な侵入を反映していることを示唆している。われわれが用いた系は, in vitroにおいて結核菌の毒力を解析するために有用であると思われる。

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Peroxisome proliferator-activated receptor γ (PPARγ) is a ligand-dependent transcription factor involved in various processes including the inflammation and carcinogenesis. The aim of the present study was 1) to examine the mRNA and protein expression of PPARγ in gastric cancer (GC); 2) to evaluate the effect of PPARγ ligand (ciglitazone) on the proliferation and apoptosis of GC cell line; and 3) to assess the levels of gastric tissue proinflammatory cytokines, IL-1β and IL-8, and plasma gastrin in GC patients before and after Helicobacter pylori (H. pylori) eradication. The trial material included 30 H. pylori-negative controls and 30 sex- and age-matched GC patients without or with H. pylori before and after its eradication. Expression of tissue PPARγ, tissue levels of IL-1β and IL-8, and plasma concentration of gastrin were significantly higher in H. pylori-positive GC compared to controls, but H. pylori eradication significantly reduced these parameters. Kato III cells incubated with alive H. pylori upregulated PPARγ expression and ciglitazone inhibited cell proliferation and induced apoptosis. PPARγ, proinflammatory cytokines and plasma gastrin appear to be implicated in H. pylori-related gastric carcinogenesis and PPARγ agonists may have potential in cancer therapy.

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Although the involvement of T helper (Th1) cells is central to protection against intracellular bacteria, including Mycobacterium tuberculosis, the involvement of Th2 cells, characterized by potent interleukin (IL)-4 secretion in mycobacterial infection is still unclear. In order to clarify the role of IL-4 in murine tuberculosis, IL-4-deficient mutant mice, IL-4 knockout (IL-4 KO) mice, were utilized. The mice were infected with H37Rv, Kurono or BCG Pasteur via an airborne infection route by placing them in the exposure chamber of a Middlebrook airborne infection apparatus. Their capacity to control mycobacterial growth, granuloma formation, cytokine secretion, and nitric oxide (NO) production were examined. These mice developed large granulomas, but not necrotic lesions in the lungs, liver or spleen (P<0.05). This was consistent with a significant increase in lung colony-forming units (CFU). Compared with levels in wild-type mice, upon stimulation with mycobacteria, splenic IL-10 levels were low and IL-6 levels were intermediate, but interferon (IFN)-γ and IL-12 levels were significantly higher. IL-18 levels were within the normal range. The level of NO production by alveolar macrophages of the IL-4 KO mice was similar to that of the wild-type mice. Granulomatous lesion development by IL-4 KO mice was inhibited significantly by treatment with exogenous recombinant IL-4. These findings were not specific to the IL-4 KO mice used. Our data show that IL-4 may play a protective role in defense against mycobacteria, although IFN-γ and TNF-α play major roles in it. Our data do not rule out an IFN-γ-independent function of IL-4 in controlling tuberculosis.

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Relationships between the growths of five different clonal strains of Heterocapsa circularisquama and intracellular bacteria were investigated using culture experiments. Although each H. circularisquama strain culture was established from one cell by repeated and careful washings with micropipettes, intracellular and extracellular bacteria in each strain culture were still observed under a epifluorescence microscope using the DAPI (4', 6-diamidino-2-phenylindole) staining technique. The extracellular bacteria are derived, presumably, from the inside of algal cells after the death and collapse of algal cells in culture. Using an electron microscope, bacteria were constantly observed in the cytoplasm and food vacuoles of H. circularisquama cells. The growth of five algal strains containing bacteria was compared with that of a bacteria-free strain using culture experiments under combined conditions of five different light intensities and five different strengths of culture medium. Bacteria showed no significant effect on the growth or survival of the algal cells. During the algal exponential growth phase, the intracellular bacterial cell numbers per algal cell decreased, whereas the total bacterial cell density in each algal culture increased. Final cell yield (total number) of the intracellular and extracellular bacteria varied considerably according to the algal strains. These results suggest that the intracellular bacteria of H. circularisquama grow or survive depending on the host alga, and that the alga can grow independently.

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　海水を入れたシャーレ内で実体顕微鏡を用いてヘテロボツリウムの交尾と卵形成過程を観察した。交尾は交接器を別の固体の体側面に押し当てて行われた。自家受精する固体もあった。個体差はあったが, 約2分サイクルで1個の卵が形成された。子宮内には最高1500を超える卵が計数された。卵と卵をつなぐ付属糸の長さから, 数珠状に連なった卵は最長2.8mあまりに達すると推定された。固定標本の計測から, 付属糸の長さは卵形成腔と子宮を結ぶ管の長さにほぼ匹敵することがわかった。

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　A marine raphidophyte Heterosigma akashiwo is a causative agent of harmful microalgal blooms, which often cause the massive mortality of aquacultured finfish. In the present study, the Pacific oyster Crassostrea gigas was reared with H. akashiwo, and effect of the microalga on filter-feeding behavior and microflora of the gastrointestinal tract was investigated. The intake of the raphidophyte cells inhibited the molluscan filter-feeding activities, suggesting the negative physiological effect of the microalgal cell contents. However, the bivalves ingested the H. akashiwo cells to the same extent as the diatom Chaetoceros calcitrans, a non-harmful indicator to estimate the filtration rate, showing a continuation of their non-selective ingestion of the phytoplankton. Microflora of the oyster soft tissue was dominated by bacteria affiliated with the family Rhodobacteraceae, some of which are associated with microalgae. In addition, the Bacteroidetes species, in which algicidal bacteria are included, were also found in the bivalve individuals exposed to H. akashiwo. These results suggested that the ingested phytoplankton affected the microbial flora in the gastrointestinal tracts, some constituents of which helped the mollusc assimilate the ingested red tide phytoplankton. This study will provide beneficial information to clarify mechanisms by which the oyster evades the ichthyotoxicity of harmful microalgae and the participation of the intestinal microorganisms in these processes.

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Imidacloprid is a pesticide used to control aphid infestations of cotton plants. However, poisoned aphids also serve as food for the ladybird natural predator Hippodamia variegata. We investigated whether imidacloprid-treated eggs, pupae, and adults of H. variegata and poisoned aphids altered ladybird predatory behavior. Laboratory bioassay results demonstrated that 0.72 g/L imidacloprid was lethal to ladybirds. Imidacloprid significantly reduced the hatching and emergence rates of H. variegata, and these effects were time and dose dependent. Predation was most adversely affected when the ladybirds directly consumed poisoned aphids and less so when directly exposed to the insecticide at sublethal concentrations. Imidacloprid use in cotton fields should be restricted to the initial stages of aphid infestation to avoid the period when adult ladybirds are present.

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Heterocapsa circularisquama showed much higher toxic effects on short-necked clams than Chattonella marina. Clams exposed to H. circularisquama exhibited morphological changes concomitant with an accumulation of mucus-like substances in the gills, a profound reduction in filtration activity, and lysosomal destabilization in hemocytes. Chattonella marina was less effective than H. circularisquama, and Heterocapsa triquetra was almost harmless in all these criteria. These results suggest that H. circularisquama exerted its lethal effect on short-necked clams through gill tissue damage and subsequent induction of physiological stress.

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Cross-protection between Haemophilus parasuis serovars 2 and 5 was examined in pigs using a bacterin based vaccine, and subsequently the safety and efficacy of a bivalent vaccine were evaluated. Upon intratracheal challenge of a serovar 2 or 5 strain, pigs immunized with a monovalent vaccine were protected against challenge with a homologous serovar strain, but not with a heterologous serovar strain. Immunization with a bivalent vaccine containing both serovars 2 and 5 bacterins conferred protection in pigs against lethal challenge with each of the serovar strains. A total of 86 pigs from two SPF herds were injected with the bivalent vaccine intramuscularly twice at a four-week interval. No adverse reactions following the vaccination were observed. On day 7 after the second vaccination, vaccinated and non-vaccinated control pigs from herd A were transferred to herd B, where Glasser's disease had broken out. Pigs in the control group developed clinical signs of the disease, and 6 of 8 (75%) pigs died until slaughter, in contrast with only 4 of 46 (9%) pigs in the vaccinated group. In herd C, where there was no outbreak of Glasser's disease, complement fixation antibody titer was raised only in the vaccinated group. A challenge experiment on days 20 and 79 after the second vaccination showed that only the vaccinated pigs were protected. From these findings, the safety and efficacy of the bivalent vaccine were confirmed under laboratory and field conditions.

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ラットをレ線で 800γ全身照射し, 照射後6時間内に, 予めラット肝 Homogenate で感作しておいたウサギの骨髄細胞乃至脾細胞を移植した(移植細胞数10^8).移植後ラットはアレルギー性肝病変を示すが, かゝるラットの血清中には, ラット肝 Homogenate の上清と反応する沈降抗体が, 1〜4週間に亘って証明される.また移植ラットの血清を分画し Globulin 分画を1^<131>で標識し, これを正常ラットに静注すると, このI^<131>標識 Globulin は特異的に肝に局在する.即ち移植ラット血清中には Localizing Antibody が証明される.上述の沈降抗体及び Localizing Antibodyは移植されたウサギ骨髄細胞乃至脾細胞によって産生されたものである.異種抗肝抗体を1回静注した場合は, その中に含まれる沈降抗体は比較的長く, 血清中に残存するが Localizing Antibody は短時間のうちに血清中より Clear され肝に局在する.これに反し異種抗肝感作細胞の移植の場合は, 抗肝沈降抗体もまた Localizing Antibody も血清中に証明しうることが判明した.また移植ラット肝より EluateをI^<131> で標識し, これを正常ラットに静注すると, 肝に特異的に局在した.即ち移植ラット肝は Localizing Antibody で感作されていることが判明した.

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Image quality assessment method is a methodology that measures the difference of quality between the reference image and its distorted one. In this paper, we propose a novel reduced-reference (RR) quality assessment method for JPEG-2000 compressed images, which exploits the statistical characteristics of context information extracted through partial entropy decoding or decoding. These statistical features obtained in the process of JPEG-2000 encoding are transmitted to the receiver as side information and used to estimate the quality of images transmitted over various noisy channels at the decompression side. In the framework of JPEG-2000, the context of a current coefficient is determined depending on the pattern of the significance and/or the sign of its neighbors in three bit-plane coding passes and four coding modes. As the context information represents the local property of images, it can efficiently describe textured pattern and edge orientation. The quality of transmitted images is measured by the difference of entropy of context information between received and original images. Moreover, the proposed quality assessment method can directly process the images in the JPEG-2000 compressed domain without full decompression. Therefore, our proposed can accelerate the work of assessing image quality. Through simulations, we demonstrate that our method achieves fairly good performance in terms of the quality measurement accuracy as well as the computational complexity.

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Strain-specific immune responses may play a critical role in the acute exacerbation of chronic obstructive pulmonary disease (COPD) caused by Haemophilus influenzae (NTHi), and the outer membrane protein P2 is one of surface antigens of NTHi, which may contribute to the strain-specific protective immunity. We examined whether repeated airway immunizations with killed-NTHi strains bearing different P2 molecules were capable of inducing protective immunity against homologous or heterologous strains in the lungs of a mouse model. Three different strains of NTHi were used in this study. Three serial intratracheal (IT) immuizations of a single strain or three different strains of NTHi led to the production of cross-reactive immunoglobulins G and A in bronchoalveolar lavage fluids. Three serial IT immunizations with a single strain enhanced the bacterial clearance of the homologous strain in the lungs, but no enhancement of bacterial clearance was found with three serial IT immunizations of heterologous strains. The enhancement in bacterial clearance, therefore, appears to be primarily strain-specific. Enhanced bacterial clearance of a hetrologous strain was also found after three serial IT immunizations of a single strain among two of the three strains employed for bacterial challenge. These findings suggest that P2 molecules and surface antigens other than P2 are involved in the development of pulmonary defense against NTHi in mice. Our data may explain, in part, why patients with COPD experience recurrent NTHi infections.

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Macrophages separated from the granulomatous lungs of tuberculous mice had a high amount of cholesterol esters.
Resident peritoneal macrophages of normal mice were very low in the ester content. However, when the cells were incubated with mycobacteria in Hanks' solution, the ester content of the mixture increased greatly.
Peritoneal macrophages harvested by induction with casein had a much larger amount of cholesterol esters than unstimulated resident cells. When such stimulated macrophages were incubated alone in Hanks' solution, the ester content went down probably due to hydrolysis into free form. This reduction was markedly inhibited by incubation with mycobacteria.
These observations at a macrophage level presented a cytological explanation for our previous finding that cholesterol ester content increased in the mouse lungs with the development of granulomatous lesions.

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The red tide dinoflagellate Heterocapsa circularisquama Horiguchi has intracellular bacteria. The isolation and cultivation of these bacteria were attempted for three-clonal strains of H. circularisquama (HY9423, HA92-1, and HU9433) using various culture media containing cell components of the host alga prepared by homogenization and heat killing, in addition to different organic materials (peptone and extracts). Only one intracellular bacterium of the bacterial population UBb in H. circularisquama strain HU9433 was successfully cultivated on many kinds of culture media. The intracellular bacterial populations of Yb and Ab in the algal clones HY9423 and HA92-1, respectively, could not grow in the culture media used. The presence of live algal cells is thought to be essential for the growth and survival of the bacterial populations Yb and Ab. These results indicate that the degree of dependence of the intracellular bacteria on the host algal cells differs greatly among the algae-bacteria sets of strains.

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Helicobacter pylori (H. pylori) infection and nonsteroidal anti-inflammatory drugs (NSAIDs) are two major causes of gastric ulceration, but relation between H. pylori infection and use of these drugs in gastric mucosal injury is controversial. Neutrophils have been implicated in the pathogenesis of gastric mucosal damage induced by H. pylori or NSAIDs. H. pylori itself and H. pylori extract induce neutrophil activation, such as superoxide production, expression of adhesion molecule (CD11b/CD18) and transendothelial migration, capillary plugging. Several clinical and experimental studies demonstrated that the degree of H. pylori-induced gastric mucosal injury was closely correlated with the extents of H. pylori infection and of neutrophil infiltration, suggesting implication in extravascular neutrophils for H. pylori-induced gastric mucosal injury. On the other hand, aspirin promotes neutrophil-endothelial adhesive interactions via increasing CD11b/CD18, but not neutrophil migration to extravascular space. In addition, our recent in vivo study suggests that neutrophils adhering to the blood vessels, but not neutrophils migrating to the interstitium, are implicated in aspirin-induced gastric mucosal injury. Recently, we found that administration of aspirin to gerbils three weeks after H. pylori inoculation produced severe gastric mucosal injury via marked infiltration of neutrophils. In this animal model, pretreatment with anti-neutrophil serum, elastase inhibitor or scavengers of reactive oxygen species remarkably inhibited gastric mucosal injury. These results suggest that H. pylori infection potentiates aspirin-induced gastric mucosal injury by mechanisms that include accumulation of activated neutrophils.

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Although some studies have indicated a correlation between Helicobacter pylori infection and the risk of colorectal neoplasms, these findings have not been consistent and are controversial. This case-control study aimed to investigate the association between endoscopic gastric mucosal atrophy and colorectal polyp occurrence. Records of 7,394 participants who underwent colonoscopy examinations from August 2008 to July 2018 were reviewed retrospectively. A total of 2,404 subjects were registered; 1,565 (65.1%) were in the gastric mucosal atrophy-positive group and 1,138 (47.3%) had colorectal polyps. The multivariate analysis adjusted by age, sex, smoking habits, alcohol habits, hemoglobin A1c, and systolic blood pressure indicated that patients in the gastric mucosal atrophy-positive group more frequently had colorectal polyps compared with patients in the gastric mucosal atrophy-negative group (odds ratio, 3.27; 95% confidence interval, 2.68–4.01; p<0.001). An analysis of the association between gastric mucosal atrophy degree and colorectal polyp status indicated that, compared with mild gastric mucosal atrophy, severe gastric mucosal atrophy was associated with a higher risk of proximal colon polyps (odds ratio, 1.47; 95% confidence interval, 1.05–2.07; p = 0.024) and two or more colorectal polyps (odds ratio, 1.80; 95% confidence interval, 1.30–2.49; p<0.001). In conclusion, gastric mucosal atrophy found during esophagogastroduodenoscopy may be an indication for complete colon screening.

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Helicobacter pylori (H. pylori) urease is a key protein for persistent infection of the bacteria in the stomach. Although H. pylori generally induce anti-H. pylori-specific antibodies (Abs), these Abs do not usually work for eradication or prevention of the H. pylori infection. In our previous study, we identified a linear epitope composed of 19-mer peptides termed UB-33, CHHLDKSIKEDVQFADSRI, within the large subunit of H. pylori urease. Anti-UB-33-specific Abs neutralized the enzymatic activity of H. pylori urease in vitro. In the present study, we evaluated the effect of immunization of BALB/c mice with H. pylori UB-33 peptide. After confirming the production of anti-UB-33-specific Abs, mice were challenged orally with H. pylori Sydney Strain-1 (SS-1). Mice producing anti-UB-33-specific Abs were not infected with SS-1, and the amount of SS-1 isolate in their stomach was significantly reduced. Also, the urease-negative mutant of H. pylori, HPP1801, did not colonize in the stomach, indicating that H. pylori urease was a critical element for infection of H. pylori in the gastric mucosa. Moreover, mice producing UB-33-specific Abs apparently suppressed H. pylori infection in the stomach where anti-UB-33 Abs were secreted in the gastric juice, indicating that H. pylori colonization was inhibited in the presence of anti-UB-33 Abs. In addition, the neutralization activity of sera from mice immunized with purified urease was less potent than that in the sera from mice immunized with UB-33. Furthermore, the recognition of epitope UB-33 was mediated through Toll-like receptor 2 (TLR2) on the B-1 cells using TLR2-knockout BALB/c mice in vivo. These results indicate that liner peptide UB-33 should be used for immunization to induce neutralizing Abs instead of purified H. pylori urease to prevent H. pylori infection and their colonization in the stomach.

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