Development of a SYBR green real-time polymerase chain reaction assay for quantitative detection of Babesia gibsoni (Asian genotype) DNA.

A real-time fluorogenic polymerase chain reaction (PCR) assay based on SYBR green that allows for sensitive, reproducible, and accurate quantification of Babesia gibsoni (Asian genotype). DNA from peripheral blood of infected dogs was developed. Standard curves were created by plotting the input amount of a standard template, constructed with plasmid DNA containing 182 base pairs (bp) of the p18 gene, against threshold cycle numbers. The curves showed a wide dynamic range (1,000,000-fold input) and high correlation values (>0.99). The PCR amplification efficacy of the standard template was similar to that of intact genomic DNA obtained from peripheral blood with B. gibsoni infection. The detection limit of the assay was 9 parasites/microl of blood with B. gibsoni infection. The intra-assay and interassay coefficients of variation of the threshold cycles ranged from 0.70% to 1.89% and from 1.18% to 1.92%, respectively. This assay system was found to be reproducible and accurate for the quantification of parasite DNA in experimentally infected dogs and far more sensitive than traditional microscopic examination.     

Prediction of cellular radiosensitivity from DNA damage induced by gamma-rays and carbon ion irradiation in canine tumor cells

Diseases of companion animals are shifting from infectious diseases to neoplasms (cancer), and since radiation therapy is one of the effective choices available for cancer treatment, the application of radiotherapy in veterinary medicine is likely to increase. However tumor tissues have different radiosensitivities, and therefore it is important to determine the intrinsic radiosensitivity of tumors in individual patients in advance of radiotherapy. We have studied the relationship between the surviving cell fraction measured by a clonogenic assay and DNA double strand breaks detected by a comet assay under neutral conditions in three canine tumor cell lines, after gamma-ray and carbon ion irradiation. In all the cell lines, cell death assessed by the clonogenic assay was much higher following irradiation with carbon ions than with gamma-rays. The initial and residual (4 hr) DNA damage due to gamma-ray and carbon ion irradiation were higher in a radiosensitive cell line than in a radioresistant cell line. The surviving cell fraction at 2 Gy (SF2) showed a tendency for correlation with both the initial and residual DNA damage. In particular, the residual damage per Gy was significantly correlated with SF2, regardless of the type of radiation. This indicates that cellular radiosensitivity can be predicted by detection of radiation-induced residual DNA damage.     

Ethylene-enhanced chlorophyllase activity during degreening of Citrus unshiu Marc.

Ethylene induces the degreening of the peel of Citrus unshiu Marc. and simultaneously the increase of chlorophyllase activity, indicating that ethylene accelerates the chlorophyll destruction through the enhancement of chlorophyllase activity. Cycloheximide inhibited the ethylene-enhanced chlorophyll degradation and repressed the enhancement of the chlorophyllase activity, while chloramphenicol did not affect the chlorophyll degradation nor the chlorophyllase activity. Based on these results, the role of ethylene in the chlorophyllase activation and the chloroplast senescence is discussed.     

F-wave conduction velocity, persistence, and amplitude for the tibial nerve in clinically normal cats

OBJECTIVE: To establish a method of F-wave evaluation and to determine normative values of F-wave parameters, including F-wave conduction velocity, persistence, and amplitude for the tibial nerve in cats. ANIMALS: 30 clinically normal cats. PROCEDURES: F-waves elicited in the interosseous muscles by stimulation of the tibial nerve were recorded, and linear regression analyses of the shortest latency versus the length of the tibial nerve and the limb length were performed. F-wave persistence was calculated by dividing the number of recorded F-waves by the number of stimuli. RESULTS: The correlation coefficient between F-wave latency and nerve length was 0.92, and that between F-wave latency and limb length was 0.58. Mean +/- SD F-wave conduction velocity of the tibial nerve was calculated to be 97.1 +/- 5.0 m/s. Linear regression analysis yielded the regression equation as follows: F-wave latency (milliseconds) = 2.60 + (0.02 x nerve length [mm]). Mean F-wave persistence and amplitude were 98.7 +/- 2.3% and 1.01 +/- 0.62 mV, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that nerve length should be used for nerve conduction studies of F-waves in felids. The regression equation for F-wave latency, conduction velocity, persistence, and amplitude may contribute to the diagnosis of nervous system diseases or injury in cats, such as trauma to the spinal cord or diabetic neuropathy.     

Effects of active egg white product/ Clostridium butyricum Miyairi 588 additive on peripheral leukocyte populations in periparturient dairy cows

The leukocyte populations of periparturient dairy cows were analyzed after administration of active egg white/Clostridium butyricum Miyairi additive. Sixty-eight Holstein milking cows were divided into 3 groups. Group A was administered active egg white product (AEWP)/Clostridium butyricum Miyairi 588 (Miyairi 588) additive (n=23). Group B was administered Miyairi 588 only (n=23), and Group C was the control group (n=22). The challenged groups were administered 100 g of AEWP + Miyairi 588, or Miyairi 588 alone, daily for 60 days from 1 month before until 1 month after paturition. Blood samples were collected from all groups three times (1 month before, 1 week after and 1 month after parturition) for analysis of the peripheral leukocyte population. The results showed significantly higher numbers of CD4+ cells in Group A compared with Group C 1 week after paturition. AEWP/Miyairi 588 additive may enhance the number of CD4+ T cells in periparturient dairy cows.     

Effect of dietary administration of bananas on immunocytes in F1 hybrid calves

The effect of dietary administration of bananas on immunocytes in calves was investigated. Twenty Fl hybrid calves were used in this study (treated group n=10, control group n=10). Banana (2 g/kg BW) was administered to the calves for 5 days. Leukocyte subsets were examined on days 0, 5, 10, and 15. The numbers CD3+, (CD3+)CD45R-, and (CD3+)TcR+ cells significantly increased between day 0 and day 5 in the treated group (P<0.01), and were significantly higher on day 5 in the treated group relative to the control group (P<0.05). These data showed that feeding banana to calves increased T-lymphocytes, suggesting it might be possible to enhance protective functions against infections.     

Prevalence of equine proliferative enteropathy in Hidaka district,Hokkaido, over five seasons

Equine proliferative enteropathy (EPE) is an equine infectious disease that can lead to severe weight loss and hyperplasia of the intestinal mucosa due to infection with Lawsonia intracellularis. In this study, we investigated the prevalence of EPE in a major Thoroughbred breeding area: Hidaka district, Hokkaido, Japan. Of the 252 symptomatic horses that we tested, 192 EPE cases (76.2%), including 8 fatal cases, were confirmed from April 2015 to March 2020 by etiological and/or serological investigation. Most of the EPE cases were observed in foals (88.5%), with fewer cases in yearlings (7.3%) and adults (4.2%). Asymptomatic infection was observed in 62.9% of the horses kept with affected horses. These results suggest that EPE is an enzootic disease in Hidaka district.     

New cylindrocladium diseases of strawberry and coral bells caused by <Emphasis Type="Italic">Cylindrocladium canadense</Emphasis>

Based on inoculation experiments and morphological and molecular phylogenetic studies, Cylindrocladium damping-off of strawberry and Cylindrocladium blight of coral bells in Japan were demonstrated to be caused by Cylindrocladium canadense. This is the first report of a disease on strawberry and coral bells caused by this fungus.     

The behavior of deuterium-labeled monolignol and monolignol glucosides in lignin biosynthesis in angiosperms

To examine the behavior of monolignol and monolignol glucosides in lignin biosynthesis, pentadeutero[9-D(2), 3-OCD(3)]coniferyl alcohol and pentadeutero[9-D(2), 3-OCD(3)]coniferin were synthesized and fed to growing Eucalyptus camaldulensis and Magnolia kobus. The differences in the incorporation patterns of these labeled precursors were studied using gas chromatography-mass spectrometry (GC-MS). Both precursors were incorporated into lignin, but the labeled coniferyl alcohol was incorporated more directly, resulting in a high proportion of pentadeutero-labeled guaiacyl and syringyl units in newly formed xylem, while labeled coniferin tended to be incorporated in lignin as tetradeutero units, especially in syringyl lignin in both trees. However, the incorporation efficiencies of the precursors into syringyl lignin were higher in Magnolia than in Eucalyptus, and the ratios of tetradeutero to pentadeutero in guaiacyl lignin were lower in Magnolia than in Eucalyptus when the trees were fed coniferin.