Changes in the microbial community structure in soils treated with a mixture of glucose and peptone with reference to the respiratory quinone profile

Based on the respiratory quinone profile, changes in the structure of microbial communities in the soil samples from Nagoya University Farm were monitored after the treatment with 1% of a mixture of glucose and peptone. Samples of two soils differing in the fertilization history were examined: CF-soil with the application of only chemical fertilizers and FYM-soil with the application of only farmyard manure at a high rate. In the CF-soil, the amount of water-soluble organic carbon (WOC), indicator of the mixture of glucose and peptone, decreased to the original level after 14 d. After 7 d, the soil pH reached the maximum level, then decreased gradually. Changes in the inorganic nitrogen levels in the water extract also reflected the 14-d period of mineralization. The amount of respiratory quinones reached maximum levels after 7 d and gradually decreased, reflecting the changes in the microbial biomass. The quinone composition significantly changed during the 14-d period and returned to a profile similar to the original one after 28 d. Diversity of quinones significantly decreased during the 14-d period due to the predominance of ubiquinone with 9 isoprenoid units. In the FYM-soil, the amount of WOC decreased to the original level after 1 d, and the pH and inorganic nitrogen levels in the water extract reflected the one-day mineralization period, and nitrification started after 3 d. Although the amount of quinones indicated an increase in the microbial biomass for 14 d, the quinone composition did not change. These findings suggested that long-term application of farmyard manure resulted in stable microbial communities in response to the incorporation of organic matter in soil.     

Matrix-metalloproteinases-2 and -9 production in bovine endometrial cell culture

In vitro cell culture is a convenient tool for studying cellular mechanisms. In the present study, production of matrix-metalloproteinases (MMPs) in bovine endometrial (containing both epithelial and stromal cells) monolayer cells was examined. Blastocysts attached to the endometrial cells in a monolayer culture were examined for their effects on MMP-2 production. Initial attachment of blastocysts to the monolayer inhibited MMP-2 production by endometrial cells. But once trophoblast cells began to migrate into the endometrial cell layer, MMP-2 production increased, and at the same time MMP-9 production also became evident in the medium. In order to understand how blastocysts affected MMP-2 production, we examined the effect of progesterone, estradiol, insulin-like growth factors (IGFs), tumor necrosis factors (TNFs), and interferon-tau (IFN-tau) supplementation. It was IFN-tau that inhibited the production of MMP-2. In addition, progesterone at a lower dose appeared to inhibit MMP-2 production. Both TNF-alpha and TNF-beta strongly stimulated the production of MMP-2 and MMP-9, whereas IGFs had no effect. Based on these findings, it appears that conceptus has the capacity to inhibit MMP activity.     

In vitro characterization of the inhibitory effects of ketoconazole on metabolic activities of cytochrome P-450 in canine hepatic microsomes

OBJECTIVE: To evaluate the inhibitory potency of ketoconazole (KTZ) on the metabolic activities of isozymes of cytochrome P-450 (CYP) in dogs. ANIMALS: 4 healthy 1-year-old male Beagles. PROCEDURE: Hepatic microsomes were harvested from 4 dogs after euthanasia. To investigate the effects of KTZ on CYP metabolic activities, 7-ethoxyresorufin, tolbutamide, bufuralol, and midazolam hydrochloride were used as specific substrates for CYP1A1/2, CYP2C21, CYP2D15, and CYP3A12, respectively. The concentrations of metabolites formed by CYP were measured by high-performance liquid chromatography, except for the resorufin concentrations that were measured by a fluorometric method. The reaction velocity-substrate concentration data were analyzed to obtain kinetic variables, including maximum reaction velocity, Michaelis-Menten constant, and inhibitory constant (Ki). RESULTS: KTZ competitively inhibited 7-ethoxyresorufin O-deethylation and midazolam 4-hydroxylation; it noncompetitively inhibited tolbutamide methylhydroxylation. Bufuralol 1'-hydroxylation was inhibited slightly by KTZ. The mean Ki values of KTZ were 10.6+/-6.0, 170+/-2.5, and 0.180+/-0.131 microM for 7-ethoxyresorufin O-deethylation, tolbutamide methylhydroxylation, and midazolam 4-hydroxylation, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: In dogs, KTZ at a therapeutic dose may change the pharmacokinetics of CYP3A12 substrates as a result of inhibition of their biotransformation. Furthermore, no influence of KTZ on the pharmacokinetics of CYP1A1/2, CYP2C21, and CYP2D15 substrates are likely. In clinical practice, adverse drug effects may develop when KTZ is administered concomitantly with a drug that is primarily metabolized by CYP3A12.     

Delignification of cell walls of <Emphasis Type="Italic">Chamaecyparis obtusa</Emphasis> during alkaline nitrobenzene oxidation

To clarify the behavior of whole lignins in wood cell walls during alkaline nitrobenzene oxidation, the delignification process from cell walls in normal and compression woods of Chamaecyparis obtusa Endl. (Cupressaceae) was observed using ultraviolet and transmission electron microscopies. The lignin content conspicuously decreased to around 10% after 35min in normal wood. The lignin content in compression wood finally leveled off at aroumd 10% after 50min. In gel filtration of oxidation products in ethyl acetate, a high molecular weight fraction was prominent in extracts from the early stage of the reaction. As the oxidation progressed, the high molecular weight fraction became less prominent in both normal and compression wood. Changes in the weights of cell wall residues during reaction indicated that approximately half of the components other than lignin were also removed from the cell walls. This shows that the majority of lignin with relatively high molecular weight is removed from the cell walls together with polysaccharides in the early stage of the reaction and that further oxidative degradation occurs in solution in later stages. Only a small amount of the lignin with low molecular weight could be analyzed by gas chromatography.Parts of this report were presented at the 47th (Kochi, April 1997) and 48th (Shizuoka, April 1998) Annual Meetings of the Japan Wood Research Society, and at the Lignin Symposium, Sapporo, October 1997     

Na?ve-like conversion enhances the difference in innate in vitro differentiation capacity between rabbit ES cells and iPS cells

Quality evaluation of pluripotent stem cells using appropriate animal models needs to be improved for human regenerative medicine. Previously, we demonstrated that although the in vitro neural differentiating capacity of rabbit induced pluripotent stem cells (iPSCs) can be mitigated by improving their baseline level of pluripotency, i.e., by converting them into the so-called “naïve-like” state, the effect after such conversion of rabbit embryonic stem cells (ESCs) remains to be elucidated. Here we found that naïve-like conversion enhanced the differences in innate in vitro differentiation capacity between ESCs and iPSCs. Naïve-like rabbit ESCs exhibited several features indicating pluripotency, including the capacity for teratoma formation. They differentiated into mature oligodendrocytes much more effectively (3.3–7.2 times) than naïve-like iPSCs. This suggests an inherent variation in differentiation potential in vitro among PSC lines. When naïve-like ESCs were injected into preimplantation rabbit embryos, although they contributed efficiently to forming the inner cell mass of blastocysts, no chimeric pups were obtained. Thus, in vitro neural differentiation following naïve-like conversion is a promising option for determining the quality of PSCs without the need to demonstrate chimeric contribution. These results provide an opportunity to evaluate which pluripotent stem cells or treatments are best suited for therapeutic use.     

Sulfate--a candidate for the missing essential factor that is required for the formation of protein haze in white wine

Protein haze formation in white wine is dependent on the presence of both wine protein and other unknown wine components, termed factor(s) X. The ability to reconstitute protein haze upon heating artificial model wine solutions (500 mg/L thaumatin, 12% ethanol, 4 g/L tartaric acid) to which candidate components were added was employed to identify factor(s) X. No protein haze was formed in the absence of additives. The individual or combined addition of caffeic acid, caftaric acid, epicatechin, epigallocatechin-O-gallate, gallic acid, or ferulic acid at typical white wine concentrations did not generate protein haze. However, PVPP fining of commercial wines resulted in a reduction in protein haze, suggesting that phenolic compounds may play a modulating role in haze formation. To elucidate the nature of the unknown factor(s) wine was fractionated and fractions were back-added to model wine and tested for their essentiality. Wine fractions were generated by ultrafiltration, reverse-phase chromatography, and mixed-mode anion-exchange and reverse-phase chromatography. The only purified fraction containing the essential component(s) was free of phenolic compounds, and analysis by mass spectrometry identified sulfate anion as the dominant component. Reconstitution with KHSO4 using either commercially available thaumatin or wine proteins confirmed the role of sulfate in wine protein haze formation. The two main wine proteins, thaumatin-like protein and chitinase, differed in their haze response in model wines containing sulfate. Other common wine anions, acetate, chloride, citrate, phosphate, and tartrate, and wine cations, Fe(2+/3+) and Cu(+/2+), when added at typical white wine concentrations were not found to be essential for protein haze formation.     

Growth changes in bighand thornyhead <Emphasis Type="Italic">Sebastolobus macrochir</Emphasis> off the Pacific coast of northern Honshu,Japan

ABSTRACT:     The biomass of bighand thornyhead Sebastolobus macrochir was increased by the high recruitment success of the 1999–2002 year classes off the Pacific coast of northern Honshu, Japan. In this study, the growth of bighand thornyhead was examined over a 9-year period from 1996 to 2004 in this area. The growth of the 1999 year class and the 2000–2002 year classes was reduced at 3 and 2 years old, respectively, while the 1999–2002 year classes were smaller than the 1993–1998 year classes. In 2-, 3- and 4-year-old fish, the relationship between abundance and mean standard length was expressed by negative linear regressions, while fish became smaller when abundance of the year class was larger. Mean bottom temperatures were stable at depths of 350–900 m; variations in water temperature were small in the main distribution area of bighand thornyhead. We discuss the factors affecting the growth of bighand thornyhead via changes in the demersal fish community and feeding habits.     

Studies on haemophilus infection in swine I. Application of the latex agglutination test to the diagnosis of Haemophilus pleuropneumoniae (H. Parahaemolyticus) infection

A latex agglutination test was evaluated as a method for the detection and titration of antibodies against swine Haemophilus infection and it was found that the test is applicable to the etiologic diagnosis of Haemophilus infection in swine. In swine infected experimentally with H. pleuropneumoniae (H. parahaemolyticus), the micromethod of agglutination using latex particles coated with antigens of H. pleuropneumoniae was found to be comparable agar-gel immunodiffusion and complement-fixation tests, which have previously been used for the etiologic diagnosis of the disease. Antibody titers determined by the latex agglutination test corrlated well with those determined by the other serological tests. The latex agglutination test tended to detect antibodies earlier than any of the other tests. By the latex agglutination test, weak cross-reactions were observed among different serotypes of H. pleuropneumoniae, whereas no cross-reaction was demonstrated between H, pleurophneumoniae and H. parasuis. The latex agglutination test was found to be simple and useful for the serological survey of swine Haemophilus infection, especially when dealing with a large number of samples.     

Multiplex polymerase chain reaction for distinguishing Taylorella equigenitalis from Taylorella equigenitalis-like organisms.

It is difficult to distinguish isolates of Taylorella equigenitalis, the cause of contagious equine metritis, from a T. equigenitalis-like organism isolated from asymptomatic donkeys and horses. Although T. equigenitalis is responsible for a severe, contagious disease of the reproductive tract of equids, the T. equigenitalis-like organism, although contagious, does not appear to produce disease. Because of the economic consequences of correctly distinguishing isolates of these 2 microorganisms, a polymerase chain reaction (PCR)-based assay was developed that will distinguish isolates of T. equigenitalis from the T. equigenitalis-like microorganism. The primers used in the PCR assay were designed to amplify unique regions of the gene encoding the 16S ribosomal RNA.