Comparative analysis of bacterial community and antibiotic-resistant strains in different developmental stages of the housefly (Musca domestica)

The housefly (Musca domestica) is an important host for a variety of bacteria, including some pathogenic and antibiotic-resistant strains. To further investigate the relationship between the housefly and the bacteria it harbors, it is necessary to understand the fate of microorganisms during the larval metamorphosis. The major bacterial communities in three developmental stages of the housefly (maggot, pupa, and adult fly) were investigated by a culture-independent method, polymerase chain reaction–denaturing gradient gel electrophoresis (PCR?DGGE) analysis of 16S rRNA genes. The bacteria that were identified using DGGE analysis spanned phyla Proteobacteria, Firmicutes, and Bacteroidetes. Changes in the predominant genera were observed during the housefly development. Bacteroides, Koukoulia, and Schineria were detected in maggots, Neisseria in pupae, and Macrococcus, Lactococcus, and Kurthia in adult flies. Antibiotic-resistant bacteria were screened using a selective medium and tested for antibiotic susceptibility. Most resistant isolates from maggots and pupae were classified as Proteus spp., while those from adult flies were much more diverse and spanned 12 genera. Among 20 tested strains across the three stages, 18 were resistant to at least two antibiotics. Overall, we demonstrated that there are changes in the major bacterial communities and antibiotic-resistant strains as the housefly develops.     

Properties of Nucleoside Phosphorylase from Enterobacter aerogenes

The properties of uridine Phosphorylase (UPase) and purine nucleoside Phosphorylase (PNPase) at high temperature were investigated. Both enzymes were found to be distributed in a wide range of bacteria and were partially purified from Enterobacter aerogenes AJ 11125 by heat treatment, ammonium sulfate fractionation and column chromatographies onDEAE-cellulose and Sephadex G-150. The UPase was purified 109-fold, and it showed an optimum pH of 8.5 and optimum temperature of 65°C, and activity toward uridine, 2′-deoxyuridine, thymidine and uracil arabinoside but not cytidine. The Km values of UPase for uridine were 0.7 mm at 40°C and 1.8 mm at 60°C. The PNPase was purified 83-fold, and it showed an optimum pH of 6.8 and optimum temperature of 60°C, and significant activity toward purine arabinosides as well as purine ribosides. The Km values of PNPase for inosine were 0.8 mm at 40°C and 2.2 mm at 60°C.     

Application of N-(9-Acridinyl)maleimide as a Fluorescent Detection Reagent of Cysteine and Related Thiols and Disulfide Compounds on Thin-layer Chromatogram

Fluctuations of pungent principles of hot pepper fruits (capsaicinoid), chlorophylls, carotenoid, and fresh fruit weight in Capsicum annuum var. annuum cv. Karayatsubusa at different growth stages after flowering were examined. Capsaicinoid was first detected 20 days after flowering, and reached maximal level around 40 days after flowering, then later decreased gradually. The capsaicinoid composition did not show any appreciable change throughout the stages after flowering. CAP and DC were the major components in all of the stages examined. By using radioisotopic technique, it was found that the main formation and accumulation sites of capsaicinoid are in the placenta of the fruits.     

Phosphorylated TandeMBP: A unique protein substrate for protein phosphatase assay

To analyze a variety of protein phosphatases, we developed phosphorylated TandeMBP (P-TandeMBP), in which two different mouse myelin basic protein isoforms were fused in tandem, as a protein phosphatase substrate. P-TandeMBP was prepared efficiently in four steps: (1) phosphorylation of TandeMBP by a protein kinase mixture (Ca²⁺/calmodulin-dependent protein kinase Iδ, casein kinase 1δ, and extracellular signal-regulated kinase 2); (2) precipitation of both P-TandeMBP and protein kinases to remove ATP, Pi, and ADP; (3) acid extraction of P-TandeMBP with HCl to remove protein kinases; and (4) neutralization of the solution that contains P-TandeMBP with Tris. In combination with the malachite green assay, P-TandeMBP can be used to detect protein phosphatase activity without using radioactive materials. Moreover, P-TandeMBP served as an efficient substrate for PPM family phosphatases (PPM1A, PPM1B, PPM1D, PPM1F, PPM1G, PPM1H, PPM1K, and PPM1M) and PPP family phosphatase PP5. Various phosphatase activities were also detected with high sensitivity in gel filtration fractions from mouse brain using P-TandeMBP. These results indicate that P-TandeMBP might be a powerful tool for the detection of protein phosphatase activities.     

EcoBalance 2018—Nexus of ideas: innovation by linking through life cycle thinking (9–12 October 2018, Tokyo,Japan)

The International Journal of Life Cycle Assessment -     

Neural expression of G protein-coupled receptors GPR3, GPR6, and GPR12 up-regulates cyclic AMP levels and promotes neurite outgrowth

Cyclic AMP regulates multiple neuronal functions, including neurite outgrowth and axonal regeneration. GPR3, GPR6, and GPR12 make up a family of constitutively active G protein-coupled receptors (GPCRs) that share greater than 50% identity and 65% similarity at the amino acid level. They are highly expressed in the central nervous system, and their expression in various cell lines results in constitutive stimulation of cAMP production. When the constitutively active GPCRs were overexpressed in rat cerebellar granule neurons in culture, the transfected neurons exhibited significantly enhanced neurite outgrowth and overcame growth inhibition caused by myelin-associated glycoprotein. GPR12-mediated neurite outgrowth was the most prominent and was shown to depend on G(s) and cAMP-dependent protein kinase. Moreover, the GPR12-mediated rescue from myelin-associated glycoprotein inhibition was attributable to cAMP-dependent protein kinase-mediated inhibition of the small GTPase, RhoA. Among the three receptors, GPR3 was revealed to be enriched in the developing rat cerebellar granule neurons. When the endogenous GPR3 was knocked down, significant reduction of neurite growth was observed, which was reversed by expression of either GPR3 or GPR12. Taken together, our results indicate that expression of the constitutively active GPCRs up-regulates cAMP production in neurons, stimulates neurite outgrowth, and counteracts myelin inhibition. Further characterization of the GPCRs in developing and injured mammalian neurons should provide insights into how basal cAMP levels are regulated in neurons and could establish a firm scientific foundation for applying receptor biology to treatment of various neurological disorders.