Quantification of Mirafiori lettuce big‐vein virus and its vector,Olpidium virulentus,from soil using real‐time PCR

Big vein disease of lettuce (Lactuca sativa) is an economically important disease transmitted through soil by Olpidium virulentus, and has occurred in most production areas worldwide. The disease is assumed to be caused by Mirafiori lettuce big‐vein virus (MiLBVV). To understand the dynamics of the virus and its vector, MiLBVV and O. virulentus were directly detected in soil. DNA and RNA were extracted from 5 g soil using a bead beating method, followed by purification using adsorption to a column. Detection and quantification were performed using real‐time PCR and a TaqMan probe that was prepared based on the CP region of MiLBVV and the rDNA‐ITS region of O. virulentus, respectively. Furthermore, using a visual assessment of the incidence rate of big vein disease on lettuce in agricultural fields, the C_t values of MiLBVV and O. virulentus from soil were also determined using real‐time PCR. The results showed that MiLBVV concentrations in the soil were high in the field, as also determined by a visual assessment of the incidence rate of big vein disease on lettuce. However, the amount of O. virulentus in soil was not directly correlated with the incidence of MiLBVV. From these results, it is suggested that the risk of lettuce crops developing big vein disease can be estimated using an index of the amount of MiLBVV in the soil.     

Insect growth regulating activity of 1,5-disubstituted imidazoles against Bombyx mori and Oncopeltus fasciatus

Seventeen 1,5-disubstituted imidazoles possessing a variety of substituents were tested on Bombyx mori and Oncopeltus fasciatus. All compounds induced precocious metamorphosis from 4th-instar larvae of B. mori. Of the compounds tested, 1-benzyl-5-[(E)-2,6-dimethyl-1,5-heptadienyl]imidazole (KK-42) was the most active against B. mori larvae. However, none of the compounds induced precocious metamorphosis in O. fasciatus nymphs, although several showed delayed toxicity. Treated nymphs died during or immediately after the moult. A series of 1-isobutylimidazoles exhibited high activity against O. fasciatus, whereas KK-42 was inactive even at high doses. There was no apparent correlation between the ability of imidazoles to cause precocious metamorphosis in B. mori and the delayed toxicity for O. fasciatus.     

Ruminal fermentation and microbial ecology of buffaloes and cattle fed the same diet

Although buffaloes and cattle are ruminants, their digestive capabilities and rumen microbial compositions are considered to be different. The purpose of this study was to compare the rumen microbial ecology of crossbred water buffaloes and cattle that were fed the same diet. Cattle exhibited a higher fermentation rate than buffaloes. Methane production and methanogen density were lower in buffaloes. Phylogenetic analysis of Fibrobacter succinogenes‐specific 16S ribosomal RNA gene clone library showed that the diversity of groups within a species was significantly different (P < 0.05) between buffalo and cattle and most of the clones were affiliated with group 2 of the species. Population densities of F. succinogenes, Ruminococcus albus and R. flavefaciens were higher until 6 h post‐feeding in cattle; however, buffaloes exhibited different traits. The population of anaerobic fungi decreased at 3 h in cattle compared to buffaloes and was similar at 0 h and 6 h. The diversity profiles of bacteria and fungi were similar in the two species. The present study showed that the profiles of the fermentation process, microbial population and diversity were similar in crossbred water buffaloes and crossbred cattle.     

Genome‐wide association studies reveal additional related loci for fatty acid composition in a Duroc pig multigenerational population

The aim of the present study was to detect quantitative trait loci affecting fatty acid composition in back fat and intramuscular fat in a Duroc pig population comprising seventh‐generation pedigrees using genome‐wide association studies (GWAS). In total, 305 animals were genotyped using single nucleotide polymorphisms (SNPs) array and five selected SNPs from regions containing known candidate genes related to fatty acid synthesis or metabolism. In total, 24 genome‐wide significant SNP regions were detected in 12 traits, and 76 genome‐wide suggestive SNP regions were detected in 33 traits. The Sus scrofa chromosome (SSC) 7 at 10.3 Mb was significantly associated with C17:0 in intramuscular fat, while the SSC9 at 13.6 Mb was significantly associated with C14:0 in intramuscular fat. The SSC12 at 1.0 Mb was significantly associated with C14:0 in back fat and the SSC14 at 121.0 Mb was significantly associated with C18:0 in intramuscular fat. These regions not only replicated previously reported loci containing some candidate genes involved in fatty acid composition (fatty acid synthase and stearoyl‐CoA desaturase) but also included several additional related loci.     

Nucleotide sequence of the coat protein gene of Mirafiori lettuce virus

 The coat protein (CP) gene of Mirafiori lettuce virus (MiLV), a tentative member of the genus Ophiovirus was isolated and sequenced. The established sequence consists of 1514 nucleotides including one open reading frame (ORF) with 1311 nucleotides that encodes 437 amino acids with a relative molecular mass 48 543. When the ORF was expressed in Escherichia coli, the obtained protein was confirmed as CP by Western blotting using an antiserum against MiLV. Database searches showed that the CP gene of MiLV has a sequence similar to that of Citrus psorosis virus (CPsV), the type species of the genus Ophiovirus. The comparison between MiLV and CPsV CP genes revealed that the identities of the nucleotide and amino acid sequences were 46.5% and 30.9%, respectively. Received: July 29, 2002 / Accepted: October 2, 2002     

Purification and characterization of a lipolytic enzyme active at low temperature from Norwegian Typhula ishikariensis group III strain

An extracellular lipolytic enzyme active at low temperature was purified from the culture filtrate of the snow mold fungus, Typhula ishikariensis group III, strain 6-1-1. The molecular mass of enzyme was approximately 83 kDa (SDS-PAGE). The lipolytic enzyme was most active for p-nitrophenyl palmitate at 30 –C (pH 9.0). The lipolytic activity was 23.4% of the maximum at 4 –C, the temperature of culture. Thus, the isolated T. ishikariensis lipolytic enzyme is thought to represent a new member of a group of enzymes active at low temperature.     

Characterization of tetraploid plants regenerated via protoplast culture of Iris fulva and their crossability with Japanese irises

Characteristics such as flower form, size and color of outer and inner perianths, anthocyanins in outer perianths, size, color and fertility of pollen and self-fertility of diploid and tetraploid lines regenerated via protoplast culture of Iris fulva were examined and compared with those of the diploid wild line. Among these characteristics, flower form, inner and outer perianth sizes of the tetraploid lines were noticeable, because these lines had upward flower forms and bigger flowers than diploid lines. Furthermore, reciprocal crosses between diploid or tetraploid lines of I. fulva and I. ensata and those of I. fulva and I. laevigata were performed. Three seedlings were obtained from the cross of tetraploid I. fulva × diploid I. laevigata through embryo rescue. One of them was identified as the interspecific hybrid between tetraploid I. fulva and I. laevigata by flow cytometoric (FCM), cytological and molecular (RAPD) analyses. This is the first report on production of hybrids from these lines. I. fulva has unique brown flowers, and this trait could be very useful for flower color breeding of I. laevigata which lacks this color. Therefore, the hybrid of I. fulva (4×) × I. laevigata may be the best available gene source for brown color breeding of this species.