Automated analysis of fine‐root dynamics using a series of digital images

Information related to the growth of fine roots is important for understanding C allocation in trees and the mechanisms of C cycling in ecosystems. Observations using a camera or scanner embedded in the soil enabled us to obtain continuous images of fine‐root‐growth dynamics. However, these methods are still labor‐intensive because the image analysis has to be conducted manually. We developed an automated method for tracking movement or elongation of fine roots using a sequence of scanner images. We also show how data obtained with these methods can be used for calculating fine‐root behavior. Two A4‐size scanners were buried in a mixed forest in Japan and images were taken continuously from within the soil. We preprocessed these images by extracting the fine‐root area from the images and developed an automated calculation plug‐in we named A‐root for tracking growth movement of the tips of fine roots. A‐root and manual‐tracking results were compared using the same images. The results show the A‐root and manual‐tracking methods yielded similar levels of accuracy. The average growth rate of 17 fine roots tracked using the program was 0.16 mm h^–1. The observation of the direction of growth in fine roots showed the direction may be influenced by the original root's growth where the fine roots branched, distribution of soil particles, other roots, and the force of gravity. The A‐root analysis also suggested there may be an interaction between speed of growth and changes in direction of growing fine roots.     

Involvement of jasmonic acid signalling in bacterial wilt disease resistance induced by biocontrol agent Pythium oligandrum in tomato

When the biocontrol agent Pythium oligandrum (PO) colonizes the rhizosphere, it suppresses bacterial wilt disease in tomato (Solanum lycopersicum cv. Micro‐Tom) caused by Ralstonia solanacearum, and a homogenate of its mycelia exhibits elicitor activity, inducing an ethylene (ET)‐dependent defence response in Micro‐Tom. Since salicylic acid (SA) and jasmonic acid (JA) play an important role in plant defence responses to pathogens, the involvement of SA‐ and JA‐dependent signal transduction pathways in resistance to R. solanacearum was investigated in tomato roots treated with a mycelial homogenate of PO. Bacterial wilt disease was also suppressed in tomato cv. Moneymaker treated with the PO homogenate. However, the SA‐inducible PR‐1(P6) gene was not up‐regulated in either Micro‐Tom or Moneymaker. SA did not accumulate in homogenate‐treated roots in comparison with distilled water‐treated controls, even 24 h after inoculation. Induced resistance against R. solanacearum was not compromised in SA‐non‐accumulating NahG transgenic plants treated with the PO homogenate. On the other hand, the expression of the JA‐responsive gene for the basic PR‐6 protein was induced in both tomato cultivars treated with the PO homogenate. Furthermore, quantitative disease assays showed that the induced resistance against R. solanacearum was compromized in PO homogenate‐treated jai1‐1 mutant plants defective in JA signalling. These results indicated that the JA‐dependent signalling pathway is required for PO‐induced resistance against R. solanacearum in tomato.     

Predation by neogastropods on <Emphasis Type="Italic">Turbo cornutus</Emphasis> juveniles and other small gastropods inhabiting coralline algal turfs

To understand the effects of predation by gastropods on juveniles of the Japanese spiny turban snail Turbo cornutus, a field survey and laboratory experiments were conducted. The species compositions of the order Neogastropoda inside turfs of articulated coralline algae (ACA) on the east coast of Sagami Bay were surveyed monthly, and the dominant species inside the ACA turfs were identified. Laboratory experiments were conducted to examine the effects of predation by the dominant neogastropods on small gastropods in ACA turfs. Turban snails, which inhabit ACA turfs during their juvenile stages, were predated by two muricid gastropods, Ergalatax contractus and Thais bronni, with the predation rate of E. contractus being significantly higher than that of T. bronni. While E. contractus could also predate two other species of gastropods, Anachis misera and Cantharidus japonicus, the predation rates on these two snails were significantly lower than on T. cornutus. The observed defensive behaviors were different among the three prey species, and the defensive strategy related to the calcified operculum of T. cornutus was not efficient against predation by E. contractus. It is concluded that the predation by E. contractus represents an important factor in the mortality of juvenile turban snails inside ACA turfs.     

Spontaneous Erythroid Leukemia in a 7-Week-Old Crl:CD (SD) Rat

A young male Crl:CD (SD) rat with erythroid leukemia that presented with emaciation, abdominal distension and a pale visible mucosal membrane was euthanized at 7 weeks of age. At necropsy, enlargement of liver, spleen and pancreatic lymph node was noted. Analysis of blood smear samples revealed many mono- or binucleated erythroblasts that had PAS-positive vacuoles in the cytoplasm. Histopathologically, neoplastic proliferation of atypical cells was observed in the hepatic sinusoids, splenic red pulp, bone marrow, pancreatic lymph node, kidney and lung. Neoplastic cells showed a round to spindle shape, and some neoplastic cells had deeply stained small nuclei and small cytoplasms and resembled erythroblasts. Immunohistochemically, many neoplastic cells were positive for hemoglobin. To our knowledge, this is the first report of erythroid leukemia in a rat of this age. The observed features were similar to those of pure erythroid leukemia in humans.     

Decomposition of organic matter and nutrient mineralisation in wood ant (<Emphasis Type="Italic">Formica rufa</Emphasis> group) mounds in boreal coniferous forests of different age

Wood ants (Formica rufa group) are dominating ecosystem elements of the boreal region due to their wide and abundant occurrence. They collect and concentrate organic material from the surrounding forest floor by building large above-ground mounds. These mounds have higher temperature and lower water content than the surrounding forest floor. We studied how these peculiar environmental conditions affected mass loss and carbon (C), nitrogen (N), phosphorus (P) and potassium (K) mineralisation of organic matter in boreal Norway spruce (Picea abies L. Karst.)-dominated mixed forest stands of four different age classes (5-, 30-, 60-, and 100-year-old) situated in eastern Finland using the litter bag technique. Norway spruce needle litter was incubated in inhabited and abandoned wood ant mounds as well as on the surrounding forest floor. We expected decomposition to be extremely slow due to the dryness of the mounds. Mass losses inside inhabited mounds were lower compared to the surrounding forest floor (on average 30 vs 50% after 2 years) but not as low as we expected, which might be a result of ant and microbial activity in the mounds. Decomposition in the abandoned mounds proceeded similarly as on the forest floor. Nutrient mineralisation proceeded more slowly in the ant mounds than on the surrounding forest floor. Mineralisation occurred for all studied nutrients in the ant mounds, except for N, which net amount remained stable during the years of the experiment. When wood ant mounds are abandoned and their porous and dry structure is no longer maintained by the ants, their decomposition is accelerated, and nutrients may be available for uptake by plants, although the nutrient mineralisation seems still to remain lower compared to the surrounding forest floor. However, eventually the mound material will be decomposed and nutrients mineralised, thus providing a nutrient hot spot increasing the heterogeneity of forest floor nutrient availability.     

Molecular cloning of two Haemaphysalis longicornis cathepsin L-like cysteine proteinase genes.

Immunological protection of mammalian hosts against tick infestation has been proposed as the most sustainable alternative tick control method to the current use of acaricides which has several limitations. The success of this method is dependent on the identification of key molecules for use as tick vaccine antigens. Proteolytic enzymes are involved in a wide range of cellular processes in eukaryotes such as development regulation and nutrition, thus they can be considered as good target antigens for a tick vaccine. In the present study we used primers designed based on the consensus amino acid motifs flanking the conserved active sites C25 and N175 present in all papain-like cysteine proteinases to amplify by polymerase chain reaction, sequence and characterize two Haemaphysalis longicornis tick cysteine proteinase genes. Based on the nucleotide and deduced amino acid sequences, both genes were identified as members of the cysteine proteinase gene family by presence in their sequences of consensus motifs flanking the conserved active sites C25, H150 and N175 that are present in all papain-like cysteine proteinases. Both genes are about 1.2 kb in size and show high sequence homology predominantly to invertebrate cathepsin L-like cysteine proteinases.     

Relationship between retinoic acid receptor alpha gene expression and growth-inhibitory effect of all-trans retinoic acid on canine tumor cells

Retinoids show antitumor effects on human acute promyelocytic leukemia and other tumors via retinoid receptors. In dogs, the role of retinoid receptors in inhibiting tumor development remains unclear. To evaluate the correlation between the degree of expression of retinoic acid receptor alpha (RARalpha) mRNA and the antiproliferative effects of all-trans retinoic acid (ATRA) treatments, expression analysis of RARalpha mRNA and cell growth inhibition assay were performed on 17 established canine tumor cell lines, including 6 mammary gland tumor (MGT) cell lines, 3 osteosarcoma cell lines, 5 melanoma cell lines, and 3 mast cell tumor (MCT) cell lines. Among the cell lines investigated, all 3 MCT cell lines showed high expression of RARalpha, and the most effective cell growth inhibition was observed in ATRA-treated MCT cell lines. However, remarkable antiproliferative effects of ATRA treatments were not observed on other tumor cell lines with moderate or low RARalpha mRNA expression. As a result of the relationship between RARalpha mRNA expression and ATRA treatment with regression analysis, statistically significant correlation was suggested. Furthermore, real-time quantitative polymerase chain reaction analysis of RARalpha was performed on MCT tissue samples of dogs with spontaneous disease, and 5 of 9 tissues showed high expression. These results suggest that ATRA may be an effective antitumor agent for MCT in dogs, and that prior measurement of expression of RARalpha mRNA may be a good indicator of the effectiveness of ATRA treatment.     

Relationships between the age and blood test results or body sizes in Noma horses

The objective of this study is to analyze the relationships between the age and blood test results or body sizes in Noma horses by using the results of periodical health examination. Out of 45 hematological or physical items examined, statistically significant, but loose correlations were observed in 14 items. Red blood cell count, activities of aspartate aminotransferase, alkaline phosphatase, and creatinine kinase, concentrations of calcium and inorganic phosphorus decreased with aging. Conversely, mean corpuscular volume, mean corpuscular hemoglobin, lipase activity, γ-globulin and chloride concentrations, body height, chest circumference and cannon bone circumference increased with aging. The changes in a few items seemed unique to Noma horse. However, most age-related changes found in this study might be considered as a common trend in horse breeds rather than distinctive characteristic in Noma horse.     

Detection of the meq gene in the T cell subsets from chickens infected with Marek's disease virus serotype 1

The meq gene was thought to be only detected in Marek's disease virus serotype 1 (MDV 1) including a very virulent strain, Md5, while L-meq, in which a 180-bp sequence is inserted into the meq open reading frame, is found in other strains of MDV 1, such as CVI 988/R6. However, both meq and L-meq were previously detected by PCR in chickens infected with MDV 1, suggesting that MDV 1 may consists of at least two subpopulations, one with meq, the other with L-meq. To further analyze these subpopulations, we analyzed the time course changes in distribution of these subpopulations among T cell subsets from chickens infected with MDV 1. Both meq and L-meq were detected in CD4+ and CD8+ T cells infected with strain Md5 or CVI 988/R6. The shift in MDV subpopulations from one displaying meq to the other displaying L-meq and/or the conversion from meq to L-meq occurred mainly in the CD8+ T cell subset from Md5-infected chickens. PCR products corresponding to L-meq rather than meq were frequently amplified from the CD8+ T cell subset from CVI 988/R 6 -infected chickens. These results suggest that a dominant subpopulation of MDV 1 changes depending on the T cell subsets, and that L-meq is dominantly present in the CD8+ T cells which play a role in the clearance of pathogenic agents.     

Cloning and characterization of cDNA encoding a prohibitin-like protein from Theileria orientalis

A cDNA clone encoding a prohibitin-like protein (Toprh) was isolated from a piroplasm cDNA library of Theileria orientalis and its nucleotide sequence was determined. An open reading frame, encoding a polypeptide of 278 amino acid residues, was found in Toprh cDNA sequence. An intron of 89 bp was identified when this cDNA clone was compared with the Toprh gene in the genome of T. orientalis. The deduced amino acid sequence of Toprh shares 93.8, 93.1 and 69.1% identities with the prohibitins of T. parva (from chromosome 1), T. annulata (from chromosome 1), and Plasmodium falciparum, (from chromosome 10), respectively. By Western blot analysis, Toprh was found to be expressed in the piroplasm stage of the parasites.