Mutations in feline immunodeficiency (FIV) virus envelope gene V3-V5 regions in FIV-infected cats

The envelope (Env) gene V3-V5 regions of the feline immunodeficiency virus (FIV) encode the neutralizing epitopes. Since mutations in these regions induce resistance to viral neutralizing antibodies, they may influence the effects of vaccines. To examine the in vivo mutation rate in these regions, we cloned cDNA for the Env gene V3-V5 regions from the PBMC of experimentally FIV-infected cats, and compared the deduced amino acid sequences. Blood or plasma from an FIV Shizuoka strain-infected cat was inoculated into a second group of SPF cats, and their blood or plasma was inoculated into the third group. The amino acid sequence encoded by the viral gene of the first cat was compared with those encoded by the viral genes of a total of eight cats in the second and third groups (two and six cats, respectively). The amino acid sequences in two cats in the second and third groups were 100% homologous and in one cat in the third group was 98.3% homologous to that in the first infected cat. Five cats had the same sequence, which was 97.8% homologous to that in the first infected cat. Three kittens, born 2 months after the inoculation of the FIV Aomori-2 strain into the mother cat, were anti-FIV negative at 4 weeks after birth, but became seropositive at 33 weeks after birth, confirming FIV infection. Comparison of the encoded amino acid sequences of the viral gene in two cats at 48 weeks after birth showed 100% homology to that of the virus inoculated into the mother cat, and the remaining one cat had a single residue substitution, resulting in 99.4% homology. These results suggest that the FIV Env gene V3-V5 regions are stably maintained for at least 1-2 years after infection.     

Glucose transport and glycolytic enzyme activities in erythrocytes of two-year-old thoroughbreds undergoing training exercise

d-Glucose transport and cytosolic enzyme activities were measured in erythrocytes from 2-year-old thoroughbreds under continuous training exercise (race horses) and compared with those from untrained horses of various ages (sires, mares and untrained 2-year-old thoroughbreds). The activities of the glucose transport and glycolytic enzymes, hexokinase and pyruvate kinase, in the race horses' erythrocytes were elevated to 2–3.5 times above those of untrained horses. There were no significant differences in plasma glucose, triglyceride or IRI concentrations between the horses in training and untrained horses. The increases in glucose transport and glycolytic enzyme activities in their erythrocytes are considered to reflect an increased metabolic activity in the race horses resulting from the training exercises.Abbreviations D-GT d-glucose transport - EDTA ethylenediaminetetraacetic acid - ELISA enzyme-linked immunosorbent assay - G6PD glucose-6-phosphate dehydrogenase - HK hexokinase - IRI immunoreactive insulin - PBS phosphate-buffered saline - PK pyruvate kinase     

Differentiation of feline immunodeficiency virus vaccination, infection, or vaccination and infection in cats

BACKGROUND: Serodiagnosis of feline immunodeficiency virus (FIV) is complicated by the use of a formalin-inactivated whole-virus FIV vaccine. Cats respond to immunization with antibodies indistinguishable from those produced during natural infection by currently available diagnostic tests, which are unable to distinguish cats that are vaccinated against FIV, infected with FIV, or both. HYPOTHESIS: An enzyme-linked immunosorbent assay (ELISA) detecting antibodies against formalin-treated FIV whole virus and untreated transmembrane peptide will distinguish uninfected from infected cats, regardless of vaccination status. ANIMALS: Blood samples were evaluated from uninfected unvaccinated cats (n = 73 samples), uninfected FIV-vaccinated cats (n = 89), and FIV-infected cats (n = 102, including 3 from cats that were also vaccinated). METHODS: The true status of each sample was determined by virus isolation. Plasma samples were tested for FIV antibodies by a commercial FIV diagnostic assay and an experimental discriminant ELISA. RESULTS: All samples from uninfected cats were correctly identified by the discriminant ELISA (specificity 100%). Of the samples collected from FIV-infected cats, 99 were correctly identified as FIV-infected (sensitivity 97.1%). CONCLUSIONS AND CLINICAL IMPORTANCE: With the exception of viral isolation, the discriminant ELISA is the most reliable assay for diagnosis of FIV. A practical strategy for the diagnosis of FIV infection would be to use existing commercial FIV antibody assays as screening tests. Negative results with commercial assays are highly reliable predictors for lack of infection. Positive results can be confirmed with the discriminant ELISA. If the discriminant ELISA is negative, the cat is probably vaccinated against FIV but not infected. Positive results are likely to represent infection.     

<Emphasis Type="Italic">Eremothecium coryli</Emphasis> and <Emphasis Type="Italic">E.</Emphasis><Emphasis Type="Italic">ashbyi</Emphasis> cause yeast spot of azuki bean

Yeast-like fungi were isolated from lesions on azuki bean (cv. Shin-Kyotodainagon) seeds that had been sucked by bean bugs in Kyoto Prefecture, Japan. On the basis of morphological and physiological characteristics and sequence data of the internal transcribed spacer (ITS) regions including the 5.8S rDNA, these yeasts were identified as Eremothecium coryli and E. ashbyi. Pathogenicity of those yeasts was confirmed by a reinoculation test. To our knowledge, this is the first report of the occurrence of yeast spot in azuki bean in Japan. The nucleotide sequence data reported are available in the GeneBank/EMBL/DDBJ database as accessions AB478291–AB478309 for E. coryli AZC1–19 and AB478310–AB478317 for E. ashbyi AZA1–8.     

Aldehyde dehydrogenase activity in cancer stem cells from canine mammary carcinoma cell lines

Increasing evidence suggests that diverse solid tumours arise from a small population of cells known as cancer stem cells or tumour-initiating cells. Cancer stem cells in several solid tumours are enriched for aldehyde dehydrogenase (ALDH) activity. High levels of ALDH activity (ALDH(high)) were detected in four cell lines derived from canine mammary carcinomas. ALDH(high) cells were enriched in a CD44(+)CD24(-) population having self-renewal capacity. Xenotransplantation into immunodeficient mice demonstrated that 1×10(4) ALDH(high) cells were sufficient for tumour formation in all injected mice, whereas 1×10(4) ALDH(low) cells failed to initiate any tumours. ALDH(high)-derived tumours contained both ALDH(+) and ALDH(-) cells, indicating that these cells had cancer stem cell-like properties.     

Hepatic enzyme activities and plasma insulin concentrations in diabetic herbivorous voles

The activities of the hepatic glycolytic enzymes glucokinase (GKase) and hexokinase (HKase) in herbivorous Microtus arvalis were very low and the hepatic fructose-1,6-diphosphatase (FDPase) activities were aomost the same as those in C57BL/6J mice. Glycosuria was observed in over 50% of voles fed on a low fibre, high concentrate diet. Voles with a high incidence of glycosuria for over 6 weeks became insulin deficient. In these diabetic voles, the hepatic GKase, HKase and FDPase activities decreased considerably as a result of diminished insulin secretion and fatty degeneration of the hepatic cells. It was considered that M. arvalis would be a useful animal model in which to study disorders of glucose utilization in herbivora.     

Morphological characterization of skin ganglion-like cells in Djungarian hamsters (Phodopus sungorus)

Characteristic ganglion-like cell proliferation observed in the skin of Djungarian hamsters was investigated using 24 male and 24 female hamsters, 1-6 months of age, to examine the anatomic location of these ganglion-like cells and their morphologic features. One abdominal skin tumor composed of these cells and resembling proliferative fasciitis in humans was also examined. Skin ganglion-like cells were rarely observed in young animals but increased in number and extent with age, especially in males. These cells were frequently seen in the ventral and medial regions of the trunk and legs rather than in the dorsal and lateral regions. Light microscopic examination of these ganglion-like cells revealed abundant vesicular basophilic cytoplasm with delicate intracytoplasmic silver stain-positive fibrils. Ultrastructurally, these cells contained abundant rough endoplasmic reticulum and Golgi complexes with dilated cisternae; intracellular collagen fibrils were present within these cisternae. Heat shock protein 47, beta-tubulin, and androgen receptor were expressed in these cells. The morphologic features of cells of one tumor resembling human proliferative fasciitis were identical to those observed in ganglion-like cells. The results of the present study suggest that these ganglion-like cells are derived from intrinsic undifferentiated mesenchymal cells in the dermis or subcutaneous adipose tissue and that any tumor-like lesion they form should be regarded as an abnormal proliferative lesion of skin ganglion-like cells rather than as proliferative fasciitis or fibroma.     

Cytosolic ratio of malate dehyrogenase/lactate dehydrogenase activity in peripheral leukocytes of race horses with training

The activities of the enzymes involved in the malate-aspartate shuttle and m RNA expression of malate dehydrogenase (MDH), a crucial enzyme for the NADH shuttle that produces ATP in glucose metabolism in the peripheral leukocytes of horses, were measured to investigate the change in metabolic states with training. There were no significant differences in plasma glucose and immunoreactive insulin concentrations between race horses and riding horses, used as a comparable reference. The cytosolic and mitochondrial MDH activities in leukocytes of race horses were significantly higher than those of riding horses. High activities of MDH in leukocytes of race horses were confirmed by RT - PCR analysis on the total RNA extracted from the whole blood. The cytosolic ratio of MDH /lactate dehydrogenase (LDH) activity (M/L ratio) in leukocytes of race horses was significantly higher than in those of riding horses. Increase in the M/L ratio was considered to reflect elevation of energy metabolism in animal tissues. The M/L ratio may be a useful parameter to evaluate the difference in metabolic states between race horses and riding horses.     

Role of Cylindrosporium-type conidia of Mycosphaerella pomi (Pass.) Lindau: cause of Brooks fruit spot of apple, as an infection source in apple orchards

Ascospores of Mycosphaerella pomi, the pathogen of Brooks fruit spot of apple, were produced in pseudothecia on previously infected and overwintered apple leaves from late April through early August in Aomori Prefecture, Japan. In June 2003, the ascospores were germinating and producing Cylindrosporium-type conidia on apple fruit and leaf surfaces in an orchard. After ascospores were sprayed on apple leaves, Cylindrosporium-type conidia developed on the leaf surfaces. Such Cylindrosporium-type conidia caused typical symptoms of Brooks fruit spot on apple trees after inoculations. These results suggested that the Cylindrosporium-type conidia also serve as an infection source, in addition to the ascospores, for Brooks fruit spot in apple orchards.     

DNA fingerprinting of Pyricularia grisea by rep-PCR using a single primer based on the terminal inverted repeat from either of the transposable elements Pot2 and MGR586

We investigated the use of single primers complementary to sequences in the terminal inverted repeat (TIR) of either Pot2 or MGR586, transposable elements found in Pyricularia grisea, for DNA fingerprinting by repetitive-element-based polymerase chain reaction (rep-PCR). Under standard amplification conditions, rep-PCR with each single primer generated distinct fingerprint patterns among rice-infecting P. grisea isolates collected in Japan. With the Pot2-TIR primer, bands ranging in size from 0.2 to 8 kb and in number from 8 to 13 per isolate were amplified. Although fewer bands were amplified with the MGR586-TIR primer, this molecular technique should be more reliable to identify and classify P. grisea isolates by combining the data of fingerprint patterns from each TIR primer. In a cluster analysis based on DNA fingerprints from this rep-PCR with the Pot2-TIR primer, 10 reference isolates and 12 field isolates from Saga Prefecture in 2002 were separated into six clonal lineages. We also demonstrated that the 12 field isolates belonged to one clonal lineage. Thus, this rep-PCR method using the single primer Pot2-TIR will be useful for the analysis of the population structure of rice blast pathogens.