Molecular events involved in cellular invasion by Ehrlichia chaffeensis and Anaplasma phagocytophilum

Ehrlichia chaffeensis and Anaplasma phagocytophilum are obligatory intracellular bacteria that preferentially replicate inside leukocytes by utilizing biological compounds and processes of these primary host defensive cells. These bacteria incorporate cholesterol from the host for their survival. Upon interaction with host monocytes and granulocytes, respectively, these bacteria usurp the lipid raft domain containing GPI-anchored protein to induce a series of signaling events that result in internalization of the bacteria. Monocytes and neutrophils usually kill invading microorganisms by fusion of the phagosomes containing the bacteria with granules containing both antimicrobial peptides and lysosomal hydrolytic enzymes and/or through sequestering vital nutrients. However, E. chaffeensis and A. phagocytophilum alter vesicular traffic to create a unique intracellular membrane-bound compartment that allows their replication in seclusion from lysosomal killing. These bacteria are quite sensitive to reactive oxygen species (ROS), so in order to survive in host cells that are primary mediators of ROS-induced killing, they inhibit activation of NADPH oxidase and assembly of this enzyme in their inclusion compartments. Moreover, host phagocyte activation and differentiation, apoptosis, and IFN-γ signaling pathways are inhibited by these bacteria. Through reductive evolution, lipopolysaccharide and peptidoglycan that activate the innate immune response, have been eliminated from these gram-negative bacteria at the genomic level. Upon interaction with new host cells, bacterial genes encoding the Type IV secretion apparatus and the two-component regulatory system are up-regulated to sense and adapt to the host environment. Thus dynamic signal transduction events concurrently proceed both in the host cells and in the invading E. chaffeensis and A. phagocytophilum bacteria for successful establishment of intracellular infection. Several bacterial surface-exposed proteins and porins are recently identified. Further functional studies on Ehrlichia and Anaplasma effector or ligand molecules and cognate host cell receptors will undoubtedly advance our understanding of the complex interplay between obligatory intracellular pathogens and their hosts. Such data can be applied towards treatment, diagnosis, and control of ehrlichiosis and anaplasmosis.     

Investigation of Staphylococcus aureus positive for Staphylococcal enterotoxin S and T genes

Staphylococcus aureus produces staphylococcal enterotoxins (SEs) and causes food poisoning. It is known that almost all SE-encoding genes are present on various types of mobile genetic elements and can mobilize among S. aureus populations. Further, plasmids comprise one of SE gene carriers. Previously, we reported novel SEs, SES and SET, harbored by the plasmid pF5 from Fukuoka5. In the present study, we analyzed the distribution of these SEs in various S. aureus isolates in Japan. We used 526 S. aureus strains and found 311 strains positive for at least one SE/SE-like toxin gene, but only two strains (Fukuoka5 and Hiroshima3) were positive for ses and set among the specimens. We analyzed two plasmids (pF5 and pH3) from these strains and found that they were different. Whereas these plasmids partially shared similar sequences involved in the ser/selj/set/ses gene cluster, other sequences were different. A comparison of these plasmids with those deposited in the NCBI database revealed that only one plasmid had the ser/selj/set/ses cluster with a stop mutation in set similar to that in pH3. In addition, the chromosomes of Fukuoka5 and Hiroshima3, positive for ses and set, were classified into different genotypes. Despite the low rate of gene positivity for these SEs, it is suggested that there is diversity in plasmids and strains carrying these two SEs. Consequently, regarding the entire feature of SE prevalence, we improved the multiplex PCR detection method for the SE superfamily to obtain further insight.     

Effects of cyazofamid against Plasmodiophora brassicae Woronin on Chinese cabbage

Cyazofamid (4-chloro-2-cyano-N,N-dimethyl-5-p-tolylimidazole-1-sulfonamide) is a novel fungicide with high levels of activity against Oomycetes fungi and Plasmodiophora brassicae Woronin. The effects of cyazofamid were investigated against P. brassicae, the causal agent of clubroot disease in Chinese cabbage. Cyazofamid at 0.3 mg litre(-1) inhibited resting spore germination of this pathogen by about 80%. Cyazofamid at 3-10 mg litre(-1) exhibited fungicidal activity to resting spores of P. brassicae 1-10 days after treatment. When cyazofamid was applied to infested soil, both root-hair infections and club formation caused by P. brassicae were strongly inhibited at 1-3 mg kg(-1) dry soil. These results suggest that cyazofamid directly inhibits resting spore germination, thereby leading to the inhibition of root-hair infection and club formation. Cyazofamid at 3 mg kg(-1) dry soil also exhibited complete control of clubroot disease. The effect of broadcast soil application using a dust formulation at 2 kg AI ha(-1) (equivalent to 1.3 mg AI kg(-1) dry soil), and plug seedling tray application by a suspension concentrate formulation at 200 and 400 mg AI tray(-1) (30 x 60 x 4 cm3) against P. brassicae was also evaluated. Cyazofamid exhibited good efficacy against the pathogen. The sequential treatment including plug seedling tray application with cyazofamid and pre-plant broadcast soil application with the fungicide fluazinam also exhibited excellent levels of control. These results indicate that cyazofamid has a high potential to be an effective fungicide for the control of clubroot disease.     

Characterization of ISPsy2 and ISPsy3, Newly Identified Insertion Sequences in Pseudomonas syringae pv. eriobotryae

Two new active insertion sequences, ISPsy2 and ISPsy3, were isolated from Pseudomonas syringae pv. eriobotryae, the causal agent of stem cankers of loquat trees. ISPsy2 is 1194-bp long, has 16-bp imperfect terminal inverted repeats, and generates a 4-bp target site duplication upon insertion into the selective cartridge of the entrap vector pSHI1063. The nucleotide sequence of ISPsy2 is completely identical with that of the previously identified IS-like element located adjacent to the virulence gene psvA of Pseudomonas syringae pv. eriobotryae NAE6. The single open reading frame of ISPsy2 encodes a 323-amino-acid protein that has similarity to the transposase of the IS5 subgroup of the IS5 family. The ISPsy3 belonging to the IS91 family is 1507 bp in length, does not duplicate its target sequence, GAAC, and presents an 81% sequence homology with IS801 in P. s. pv. phaseolicola. The transposase of ISPsy3 possesses the conserved amino acid motifs found in the rolling-circle replication protein. Southern blot analysis indicated that multiple copies of ISPsy2 and ISPsy3 are present in the genomes of P. s. pv. eriobotryae and some of the other P. s. pathovars tested. Received 16 August 2001/ Accepted in revised form 19 October 2001     

Effects of slaughter age on the levels of free amino acids and dipeptides in fattening cattle

The effects of slaughter age, breed type and postmortem period were studied on the concentrations of free amino acids (FAA) and dipeptides (carnosine and anserine) in Longissimus dorsi muscle of beef. Slaughter age affected the levels of most FAA and dipeptides, and the concentrations of these compounds were significantly lower at 35 months of age than either 15 or 25 months of age. Slight increases were observed at 25 months compared with 15 months, but the differences were not significant except for a few FAA. These tendencies still remained when the concentrations were recalculated on the basis of protein. A significant breed effect was observed for taurine, carnosine and anserine. Almost all FAA were significantly increased during postmortem conditioning, but the level of each FAA fluctuated, and the fluctuations could not be explained statistically by slaughter age or breed differences.     

Mycotic aneurysm caused by Graphium species in a dog

A 10-year-old castrated male mixed-breed dog exhibited vomiting, lethargy and anorexia. An abdominal ultrasound examination revealed a focal dilation of the abdominal aorta. The dog died 2 days after examination, and necropsy revealed a ruptured aneurysm at the abdominal aorta between the kidneys. Histological examination revealed severe granulomatous and necrotizing pan-arteritis with fungal organisms. Graphium species was identified through DNA sequence analysis of the PCR product from formalin-fixed paraffin-embedded samples. To our knowledge, this is the first report of aortic aneurysm caused by Graphium species in a dog.     

Swine abortion caused by Candida parapsilosis

In July 2020, a sow in a breeding herd in the Chiba Prefecture, Japan, suffered abortion. A necropsy revealed pale pulmonary foci scattered in the two fetuses. Histologically, multifocal pulmonary necrosis was detected with numerous yeasts. The yeast was positively stained using the periodic acid-Schiff reaction and Grocott’s silver stain. Molecular identification indicated that the yeast was Candida parapsilosis. In conclusion, our results suggested that C. parapsilosis caused multifocal necrotizing pneumonia in the two fetuses. This study is the first report of a swine abortion with C. parapsilosis infection.     

A review of tower flux observation sites in Asia

Aggregating and sharing the metadata of flux observation sites results in a strong collaboration among various fields of study. Such data sharing will also be a part of the future design of a tower flux observation network in Asia. The aim of this review is to comprehend the state of tower flux observation sites in Asia. There are 109 tower flux observation sites in Asia including 51 forest sites. There are more new sites under construction in Asia than in America and Europe. These sites range from the taiga in Siberia to the rainforest in Southeast Asia, and from the equatorial to polar Koeppen climate zones. There are many highly humid areas in Asia, not only at low latitudes but also at middle latitudes. This climate condition has developed unique vegetation such as lucidophyllous (evergreen broadleaf) forest, which is distributed in warm areas with high precipitation in the growing season. However, there are only a few observations taking place in lucidophyllous forest. Rice paddy fields are also unique land cover in Asia. It is important to accumulate long-term data for rice fields with their management records, because plant activity depends highly on both climate conditions and land-use management. Flux data, especially net ecosystem exchange and related elements, are used for widespread studies not only within the flux-research community but also in other fields of study, for example remote sensing. At present, however, both the quantity and quality of the data are not sufficient for these studies. Regarding the quantity, there are many recently established sites that have not published data yet; regarding quality, flux data include uncertainties caused by methodological problems. Flux researchers are required not only to obtain flux data but also to improve their quality. Meanwhile, data users must understand there are still uncertainties in flux data. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Solubility and standard Gibbs free energy of formation of natural imogolite at 25°c and 1 atm

We carried out a dissolution experiment to determine the solubility and the Gibbs free energy of formation of natural imogolite at 25°C and 1 atm. Purified gelatinous film of natural imogolite, synthetic gibbsite, and imogolite plus gibbsite were equilibrated in 1 mmol L^-1 HCI solutions and Si, Al concentrations and pH were monitored during 700 d. Imogolite plus gibbsite and gibbsite systems reached an equilibrium after 120 d. The logarithmic values of the equilibrium constants of dissolution reactions were 12.10±0.01 for imogolite and 8.01±0.01 for gibbsite. The calculated Gibbs free energy of formation was -2929.19±3.28 kJ mol^-1 for imogolite and -1155.11 ± 1.42 kJ mol^-1 for gibbsite. These values predict that the silicic acid concentration at the imogolite-gibbsite equilibrium would be 0.120 mmol L^-1.     

Growth characteristics,phytate contents,and coagulation properties of soymilk from a low-phytate Japanese soybean (Glycine max (L.) Merr.) line

The phytic acid (myo-inositol hexakisphosphate or InsP₆) content of seed crops is important to their nutritional quality. Since it represents 75?±?10% of the total seed phosphorus (P), phytic acid is also important regarding the management of P in agricultural production. A low-phytate F5 line, No. T-2-250-4-20, was selected from the progeny of a cross between the low-phytate soybean line CX1834 and the Japanese commercial cultivar Tanbakuro. This line and its parents were grown in a field nursery, and the growth characteristics, phytate accumulation, and processing suitability for tofu were evaluated. At full maturity, the weight of seeds per plant of line T-2-250-4-20 was 5.2- and 1.3-fold higher than that of CX1834 and Tanbakuro, respectively. The amount of phytate-phosphorus as a percentage of the total P content in seeds was 23% in line T-2-250-4-20-34, 30% in CX1834, and 69% in Tanbakuro. No significant difference was observed among the three cultivars/lines in their seed magnesium (Mg), potassium (K), crude protein, and sugar. However, the calcium (Ca), crude fat and ash contents in seeds of line T-2-250-4-20-34 and Tanbakuro was lowered compared to that of CX1834. The breaking stress of tofu was estimated employing a rheometer with a decreasing concentration of the coagulant magnesium chloride (MgCl₂), starting at 15.7?mmol?L^?1. In tofu made from Tanbakuro, the concentration of MgCl₂ required to achieve the maximum breaking stress was 12.6?mmol?L^?1; however, it was 9.5?mmol?L^?1 for tofu made from T-2-250-4-20-34 and CX1834. The tofu made from Tanbakuro was soft and broke at 6.3?mmol?L^?1 MgCl₂, but, in line T-2-250-4-20-34, harder tofu was made with lower MgCl₂ concentrations. No difference was observed among the cultivars/lines in the SDS-PAGE patterns of protein in soymilk. These results indicate that we have developed a low-phytate soybean with adequate productivity, and confirmed that tofu made from the low-phytate T-2-250-4-20-34 soybean becomes coagulated and harder at a lower MgCl₂ concentration than that from high-phytate soybean cultivars.