Isolation of an endophytic diazotroph,Klebsiella oxytoca,from sweet potato stems in Japan

Abstract

The population density of endophytic bacteria in the stem of field-grown sweet potato cultivars (Beniotome [BO], Koganesengan [KS], and Shiroyutaka [SYD in Miyakonojo, Miyazaki, Japan, ranged from 10² to 10⁴ cells g^?1 fresh weight sample using a semi-solid nitrogen-free medium. Eleven strains were isolated from the stems and two isolates, BO-1 and BO-5, showed a positive reaction in the acetylene reduction activity (ARA) test. BO-1 and BO-5 were isolated from cv. Beniotome in September 1999 and 2000, respectively. Morpho-physiological characterization of these isolates revealed that BO-1 and BO-5 showed a similar colony color in potato sucrose agar slants, produced bubbles in a modified semi-solid medium, acidified the medium, and displayed similar characteristics using the API 20NE rapid diagnostic kit. Partial sequence analysis of 16S rRNA from BO-1 revealed a 100% similarity (491 bp) to that of Klebsiella oxytoca. The other 9 isolates showed a negative reaction in the ARA test, slightly acidified or did not acidify the medium. Partial sequence analysis of the 16S rRNA revealed that the isolate SY-2 corresponded to Methylobacterium sp. (99.3% similarity for 1,241 bp), BO-3 to Pantoea agglomerans (99.1% similarity for 469 bp), and BO-8 to Sphingomonas sanguinis (98.8% similarity for 419 bp).     

Change in T4-type bacteriophage communities during the composting process of rice straw: Estimation from the major capsid gene (g23) sequences

The present study examined T4-type phage communities in rice straw (RS) under the composting process by analyzing the composition of the major capsid gene ( g23 ) of T4-type bacteriophages. The g23 clones were obtained from RS throughout the composting process from RS materials to composting RS in the curing stage (for 124 days). Most of the g23 clones were phylogenetically closely related to those in rice field soils and rice field floodwaters, and Paddy Groups II and III appeared to characterize the g23 genes in the composting RS. The diversity of g23 genes in the composting RS was highest in the RS material (day 0 after the onset of composting) and in the early thermophilic stage (day 7), and decreased markedly in the middle and curing stages. This change was in contrast to that of the bacterial community, which showed higher diversity in the middle and curing stages. There was no specific clone that characterized any stage during the composting process. These findings indicate that the phage community is not the major controlling agent in determining eubacterial succession and that the thermophilic stage in the composting process efficiently annihilated T4-type phages in the composting pile.     

Potential chemopreventive properties of extract from baked sweet potato (Ipomoea batatas Lam. Cv. Koganesengan)

The extract from baked sweet potato (Ipomoea batatas Lam. cv. Koganesengan) showed potential cancer-preventing effects. The extract was partially fractionated to four fractions (I, II-a II-b, and III) by Sephadex G-25 gel chromatography. The cytotoxicity against human myelocytic leukemia HL-60 cells, the suppression of TPA-induced transformation in mouse skin JB6 C141 cells, the apoptosis inducing activity in HL-60 cells, and the scavenging capacity against DPPH radical were tested on the four fractions. Fractions II-a and III showed markedly strong radical scavenging effects on the DPPH radical, coinciding with the high content of total phenolic compounds in the fractions. Both of these fractions suppressed strongly the proliferation of HL-60 cells with apoptosis induction in a dose-dependent manner. Moreover, the two fractions markedly blocked TPA-induced cell transformation in the JB6 cell line. Taken together, these data suggest that the water extract from baked sweet potato had potential chemopreventive properties.     

Identification and morphological characteristics of dental neonatal line in sika deer (Cervus nippon)

The dental neonatal line of the sika deer (Cervus nippon) was identified experimentally using chronological labeling methods. In the enamel, prominent dark lines were observed under transmitted light, and the number of increments between the dark line and labeling line was almost consistent with the day-age at the time of labeling injection. Therefore, we identified the dark line as the enamel neonatal line. In the dentin, the bright line was observed under polarized light. Since the bright line corresponded to the enamel neonatal line, we recognized the bright line as the dentin neonatal line. Neonatal lines intersected with the enamel-dentin junction at approximately one-third cervical in the first molar. Using these features, it would make possible to distinguish the neonatal line in wild sika deer.     

A mitotic septin scaffold required for Mammalian chromosome congression and segregation

Coordination of cytokinesis with chromosome congression and segregation is critical for proper cell division, but the mechanism is unknown. Here, septins, a conserved family of polymerizing guanosine triphosphate-binding proteins, localized to the metaphase plate during mitosis. Septin depletion resulted in chromosome loss from the metaphase plate, lack of chromosome segregation and spindle elongation, and incomplete cytokinesis upon delayed mitotic exit. These defects correlated with loss of the mitotic motor and the checkpoint regulator centromere-associated protein E (CENP-E) from the kinetochores of congressing chromosomes. Mammalian septins may thus form a mitotic scaffold for CENP-E and other effectors to coordinate cytokinesis with chromosome congression and segregation.     

Anaplasma marginale infection in a Japanese Black cow 13 years after eradication of Rhipicephalus (Boophilus) microplus in Okinawa, Japan

In October 2007, a 15-year-old Japanese Black cow on Ishigaki Island, Okinawa, Japan, was diagnosed with Anaplasma marginale infection based on clinical symptoms, blood examination, smear observation, 16S rRNA and groEL gene sequence analysis, and the result of a CF test. The cow was introduced into the farm from mainland Japan as a calf in 1993, one year before the eradication of Rhipicephalus (Boophilus) microplus, the main vector of A. marginale in Okinawa Prefecture. It is possible that the cow was first infected with A. marginale as a calf in Ishigaki Island and had been persistently infected since then. This is the first reported clinical case of A. marginale infection of cattle since the eradication of R. microplus in Okinawa Prefecture. Additional analysis of major surface protein 1α amino acid sequences revealed that the A. marginale Okinawa strain presented four new repeat forms which were not seen in other strains. This indicates that the Okinawa strain may be a unique geographical variant of A. marginale.     

Anti‐tumour activity of oncolytic reovirus against canine histiocytic sarcoma cells

Canine histiocytic sarcoma is an aggressive, fatal neoplastic disease with a poor prognosis. Lomustine is generally accepted as the first‐line systemic therapy, although this compound does not provide complete regression. Therefore, research into a novel approach against canine histiocytic sarcoma is needed. However, anti‐tumour effects of oncolytic therapy using reovirus against histiocytic sarcoma are unknown. Here, we showed that reovirus has oncolytic activity in canine histiocytic sarcoma cell lines in vitro and in vivo. We found that reovirus can replicate and induce caspase‐dependent apoptosis in canine histiocytic sarcoma cell lines. A single intra‐tumoural injection of reovirus completely suppressed the growth of subcutaneously grafted tumours in NOD/SCID mice. Additionally, we demonstrated that susceptibility to reovirus‐induced cell death was attributable to the extent of expression of type I interferons induced by reovirus infection in vitro. In conclusion, oncolytic reovirus appears to be an effective treatment option for histiocytic sarcoma, and therefore warrants further investigation in early clinical trials.     

Effect of supplementation of rice bran and fumarate alone or in combination on in vitro rumen fermentation,methanogenesis and methanogens

This study investigated the effect of fumarate (FUM) and rice bran (RB), alone and together, on in vitro rumen fermentation, methanogenesis and methanogens. In vitro incubation was performed with six media that were either unsupplemented (control) or supplemented with 10% RB, 5 mmol/L FUM, 10% RB + 5 mmol/L FUM, 10 mmol/L FUM, or 10% RB + 10 mmol/L FUM. Methane (CH₄) production, dry matter digestibility, CH₄ per digested dry matter, total short‐chain fatty acid (SCFA) production, proportion of SCFA, acetate : proprionate ratio, production of NH₃‐N, and population density of rumen microbes were determined. Supplementation with 10% RB + 10 mmol/L FUM yielded a 36% decrease in CH₄ production compared to the control. Supplementation of FUM, in the presence or absence of RB, provided increases in total SCFA production and propionate proportion up to 61% and 31%, respectively. Total bacteria, methanogens and protozoa populations were significantly (P < 0.05) decreased with the 10% RB + 10 mmol/L FUM supplementation. The effect of anti‐methanogenesis of FUM was enhanced by the addition of RB. Notably, the CH₄ production attenuation was achieved by 10% RB + 10 mmol/L FUM without reduction of digestibility or of ruminal fermentation.     

Dough and Baking Properties of High‐Amylose and Waxy Wheat Flours

The dough properties and baking qualities of a novel high‐amylose wheat flour (HAWF) and a waxy wheat flour (WWF) (both Triticum aestivum L.) were investigated by comparing them with common wheat flours. HAWF and WWF had more dietary fiber than Chinese Spring flour (CSF), a nonwaxy wheat flour. Also, HAWF contained larger amounts of lipids and proteins than WWF and CSF. There were significant differences in the amylose and amylopectin contents among all samples tested. Farinograph data showed water absorptions of HAWF and WWF were significantly higher than that of CSF, and both flours showed poorer flour qualities than CSF. The dough of WWF was weaker and less stable than that of CSF, whereas HAWF produced a harder and more viscous dough than CSF. Differential scanning calorimetry data showed that starch in HAWF dough gelatinized at a lower temperature in the baking process than the starches in doughs of WWF and CSF. The starch in a WWF suspension had a larger enthalpy of gelatinization than those in HAWF and CSF suspensions. Amylograph data showed that the WWF starch gelatinized faster and had a higher viscosity than that in CSF. The loaves made from WWF and CSF were significantly larger than the loaves made from HAWF. However, the appearance of bread baked with WWF and HAWF was inferior to the appearance of bread baked with CSF. Bread made with WWF became softer than the bread made with CSF after storage, and reheating was more effective in refreshing WWF bread than CSF bread. Moreover, clear differences in dough and bread samples were revealed by scanning electron microscopy. These differences might have some effect on dough and baking qualities.