Development of a high-throughput field phenotyping rover optimized for size-limited breeding fields as open-source hardware

Phenotyping is a critical process in plant breeding, especially when there is an increasing demand for streamlining a selection process in a breeding program. Since manual phenotyping has limited efficiency, high-throughput phenotyping methods are recently popularized owing to progress in sensor and image processing technologies. However, in a size-limited breeding field, which is common in Japan and other Asian countries, it is challenging to introduce large machinery in the field or fly unmanned aerial vehicles over the field. In this study, we developed a ground-based high-throughput field phenotyping rover that could be easily introduced to a field regardless of the scale and location of the field even without special facilities. We also made the field rover open-source hardware, making its system available to public for easy modification, so that anyone can build one for their own use at a low cost. The trial run of the field rover revealed that it allowed the collection of detailed remote-sensing images of plants and quantitative analyses based on the images. The results suggest that the field rover developed in this study could allow efficient phenotyping of plants especially in a small breeding field.     

Survival and Fluctuation in Density of Myrothecium roridum in Mulberry Field Soil

The density of Myrothecium roridum increased in field soil from June to October and decreased from February to April in 1996–1998. The fluctuation in density of M. roridum in the surface soil was affected by the average temperature. In the greenhouse, mulberry leaves were infected by conidia that splashed from soil artificially infested with conidia of M. roridum at 10³ or more conidia/g soil. Disease incidence on mulberry increased when soil was amended with the fallen leaves and when mulberries were planted densely in soil artificially infested with a high conidial density of M. roridum. Received 9 December 1999/ Accepted in revised form 21 July 2000     

<Emphasis Type="Italic">Pseudomonas syringae </Emphasis>Strains Are Classified into Five Groups by Comparing DMA Homology at the <Emphasis Type="Italic">hrp </Emphasis>Neighboring Regions

Previously, we classified Pseudomonas syringae strains into at least three groups (I, II and U) by comparing DNA homology at the hrp cluster and its neighboring regions (Inoue and Takikawa 1999). However, heterogeneous strains remained in the undetermined group (group U). We further classify group U, using pvs. syringae and coronafaciens as references. Comparison of restriction sites for regions of each pathovar revealed distinct differences. By using probes from the two pathovars, comparisons of DNA homology at the regions separated two additional distinct groups (III and IV) from group U. Therefore, P. syringae strains are classified into at least five groups. Received 4 November 1999/ Accepted in revised form 27 January 2000     

Gibberella ear rot of corn caused by <Emphasis Type="Italic">Fusarium asiaticum</Emphasis> in Japan

Ear rot with white or pink mold was found on corn (Zea mays L.). A species of Fusarium, not registered previously as a pathogen causing Gibberella ear rot of corn in Japan, was isolated from the rotted ear. The isolates, identified as F. asiaticum based on morphological characteristics and nucleotide sequences, caused white or pink mold on corn ear after inoculation. Moreover, the 3-acetyl deoxynivalenol chemotype and the nivalenol chemotype were found in the isolates. We propose to include F. asiaticum as one of the pathogens causing Gibberella ear rot of corn in Japan.     

Rapid genotyping with DNA micro-arrays for high-density linkage mapping and QTL mapping in common buckwheat (Fagopyrum esculentum Moench)

For genetic studies and genomics-assisted breeding, particularly of minor crops, a genotyping system that does not require a priori genomic information is preferable. Here, we demonstrated the potential of a novel array-based genotyping system for the rapid construction of high-density linkage map and quantitative trait loci (QTL) mapping. By using the system, we successfully constructed an accurate, high-density linkage map for common buckwheat (Fagopyrum esculentum Moench); the map was composed of 756 loci and included 8,884 markers. The number of linkage groups converged to eight, which is the basic number of chromosomes in common buckwheat. The sizes of the linkage groups of the P1 and P2 maps were 773.8 and 800.4 cM, respectively. The average interval between adjacent loci was 2.13 cM. The linkage map constructed here will be useful for the analysis of other common buckwheat populations. We also performed QTL mapping for main stem length and detected four QTL. It took 37 days to process 178 samples from DNA extraction to genotyping, indicating the system enables genotyping of genome-wide markers for a few hundred buckwheat plants before the plants mature. The novel system will be useful for genomics-assisted breeding in minor crops without a priori genomic information.     

Density-dependent selfing and its effects on seed performance in a tropical canopy tree species, Shorea acuminata (Dipterocarpaceae)

In the conservation and management practices of natural forests, sound reproduction and regeneration form the basis of the maintenance and viability of the tree populations. To obtain and serve biological information for sustainable forest management, we investigated reproductive biology and inbreeding depression in seeds of an important dipterocarp tree species, Shorea acuminata (Dipterocarpaceae), by both field and laboratory experiments. Results of parental analysis of immature and mature seeds showed that selfing rates varied greatly, from 7.6 to 88.4% among eight mother trees, and the mean overall selfing rate was 38.3%. Observed outcrossing events within a 40-ha study plot were predominantly (76.5%) short-distance events with a mating distance (md) ≤ 100 m. Since the selfing rate sharply decreased with increase in the number of flowering conspecifics (i.e., individuals of the same species) within a 100-m radius from the mother trees, the local density of flowering conspecifics appears to be the key factor determining the outcrossing rate in S. acuminata. However, the extremely high selfing rate (88.4%) observed for one tree could not be simply explained by the low local density of flowering conspecifics. Instead, differences in its flowering phenology (its flowering peaked ca. a week earlier than most of the other examined individuals) may have severely limited its receipt of pollen from other conspecifics, and thus promoted selfing. Since there were no significant differences in the proportion of selfed progeny between immature and mature seed stages, there was no evidence of selective abortion of selfed seeds during seed development. However, the seed mass of outcrossed progeny was heavier than that of selfed progeny, and heavier seeds showed higher success rates at germination and seedling establishment. These results suggest that inbreeding depression resulted in reductions in seed mass and may reduce the fitness of selfed seeds in S. acuminata. In addition, the outcrossing rate of S. acuminata was more sensitive to low local conspecific flowering-tree densities than that of a sympatric bee-pollinated dipterocarp species with greater pollination distances. These results suggest that the management of local adult-tree densities is important for avoiding selfing and inbreeding depression in future generations, especially in a species like S. acuminata with predominantly short-distance pollination.     

Association between breed,gender and age in relation to cardiovascular disorders in insured dogs in Japan

The association between breed, gender and age and cardiovascular disorders in the insured dog population in Japan was investigated, using multiple logistic regression analysis and data from 299,555 dogs insured between April 2010 and March 2011. The overall annual prevalence of cardiovascular disorder diagnosis was 2.1%. Using the Miniature Dachshund as the reference breed, Cavalier King Charles Spaniel had the highest odds of cardiovascular disorder with a ratio of 16.2 (95% confidence interval: 14.4–18.2), followed by Maltese, Pomeranian, Chihuahua and Shih Tzu. Male dogs had increased odds of 1.2 (1.1–1.3). The dogs had increased odds of having cardiovascular disorder by 1.5 times as their age increased by one year.     

Chiral response of Oryzeae and Paniceae plants in alpha-methylbenzyl-3-p-tolyl urea agar medium

The results presented here support the hypothesis that plants of the tribe Oryzeae respond enantioselectively and homogeneously to optically active 1-alpha-methylbenzyl-3-p-tolylurea (MBTU) in root growth inhibition, in contrast to Echinochloa species. The Oryzeae plants tested in this study belong to different genera (Oryza, Leersia, Chikusichloa and Zizania), to different species (O sativa, O glaberrima, O alta, O coarctata, O latifolia, O minuta, O rufipogon), to various ecospecies of Oryza (japonica, indica, japonica x indica, javanica) and to different levels of evolution [cultivated rice (O sativa and O glaberrima) and ancestral wild rice species]. In spite of their different phylogenic status and diverse sensitivity, the root growth of all members of the genus Oryza was inhibited more by R-MBTU than by S-MBTU. Zizania palustris, Z latifolia, Leersia oryzoides and Chikusichloa aquatica belonging to the tribe Oryzeae exhibited similar chiral recognition to the Oryza plants, suggesting that Oryzeae have a common chiral recognition mechanism in their response to optically active MBTUs. In contrast, Echinochloa plants (E crus-galli (L) Beauv var crus-galli and E colonum (L) Link), belonging into subfamily Panicoideae tribe Paniceae, responded in a different way, where their root growth was more sensitive to S-MBTU than to the antipodal R-MBTU. A reverse chiral response between the tribe Oryzeae and the genus Echinochloa was clearly indicated in this study. This diverse response may be relevant to Gramineae classification.     

A cell preparation of Enterococcus faecalis strain EC-12 stimulates the luminal immunoglobulin A secretion in juvenile calves

The immune system in juvenile calves is immature, so calves are susceptible to several diarrheal and respiratory diseases. Oral administration of lactic acid bacteria (LAB) is known to improve the growth performance and prevent diarrheal and respiratory diseases by stimulating the immune system in juvenile calves. Most of the immunostimulation by LAB is achieved by their cell wall components, and therefore we evaluated the immunostimulation of the cell preparation of Enterococcus faecalis strain EC-12 (EC-12) in juvenile calves in a clinical field. Twenty-nine 1-week old calves were used. Fourteen calves were administered 0.2% (w/w) of an EC-12 preparation that supplemented a milk replacer, and other calves were not supplemented. Feces and serum was collected at day 0, 7 and 49 after the administration to measure the IgA and IgG concentration. The fecal IgA concentration was increased by EC-12 administration at day 49, and the serum IgA concentration was also increased at day 7. These results suggested that oral administration of EC-12 in juvenile calves might have an immunostimulatory effect and provide earlier recovery of IgA levels in mucosal immunity.     

Establishment of embryo transfer in the musk shrew (Suncus murinus)

The present study established techniques to induce pseudopregnancy, in vitro oocyte cultures from pronuclear to 2- to 4-cell stages, and embryo transfer in musk shrews, a reflex ovulator. Offspring were subsequently obtained by transferring in vivo-developed or in vitro-cultured embryos. Female musk shrews received human chronic gonadotropin (hCG), with or without mating stimuli, from vasectomized males to produce pseudopregnant recipients. Embryos at the 2- to 4-cell stage were collected 44–48 h after mating. Another set of embryos was collected 26–27 h after mating and then cultured for 20 h from the pronuclear to 2- to 4-cell stages. Subsequently, embryos were transferred into the oviducts of pseudopregnant recipients 24 or 48 h after the induction of pseudopregnancy. Offsprings were successfully obtained from recipients that received hCG 24 h before embryo transfer, regardless of mating stimuli. These techniques may be valuable for producing transgenic musk shrews.