Phylogenic outline of the vertebrate olfactory system is summarized in the present review. In the fish and the birds, the olfactory system consists only of the olfactory epithelium (OE) and the olfactory bulb (B). In the amphibians, reptiles and mammals, the olfactory system is subdivided into the main olfactory and the vomeronasal olfactory systems, and the former consists of the OE and the main olfactory bulb (MOB), while the latter the vomeronasal organ (VNO) and the accessory olfactory bulb (AOB). The subdivision of the olfactory system into the main and the vomeronasal olfactory systems may partly be induced by the difference between paraphyletic groups and monophyletic groups in the phylogeny of vertebrates.
Obstructive jaundice causes multiple malfunctions in various organs including the pancreas. To establish how such malfunctions occur, we experimentally induced obstructive jaundice through bile duct ligation (BDL) using rats, measured serum bilirubin, amylase and insulin levels, and examined histological, immunohistochemical and cytological changes in the pancreas at 3 days, 1 week, and 4 weeks after the BDL. Morphometrical analysis was also conducted. Serum amylase levels steeply increased at 3 days, and then decreased at 1 and 4 weeks after the BDL to lower than the control level. In contrast, the number of zymogen granules decreased at 3 days after the BDL, then increased and eventually surpassed the control group at 4 weeks after the BDL. On the other hand, serum insulin levels dramatically decreased at 3 days after the BDL but recovered to a level close to that of the control group at 1 week after the BDL. At 4 weeks after the BDL, however, the serum insulin levels again showed a marked decline. Slight decrease in insulin immunoreactivity and number of insulin granules were observed at 4 weeks after the BDL. Cholecystokinin receptors (CCK-R) were expressed in both acinar and islet cells; their immunoreactivity significantly decreased in the acinar cells at 4 weeks after the BDL. Our results suggest that CCK may play a role in regulating changes in the pancreas after obstructive jaundice.
To clarify the fundamental regulation mechanism against indigenous bacterial proliferation in the alimentary tract, we immunohistochemically examined the localization of 4 bactericidal peptides (BP) in the rat digestive exocrine glands. In the upper alimentary tract, lysozyme was detected in the gustatory, extraorbital lacrimal and parotid glands. Secretory phospholipase A2 (sPLA2) was detected in the extraorbital lacrimal glands. β-defensin1 was detected in the gustatory and extraorbital lacrimal glands. β-defensin2 was detected in the Harderian glands. In the stomach, β-defensins were detected in the gastric superficial epithelial cells. In the small and large intestines, only lysozyme and sPLA2 were detected in the Paneth cells. In the cecum, all 4 BP were detected in the middle to apical portions of the crypts, and only sPLA2 was detected in the basal portion. No BP were localized in other exocrine glands associated with the alimentary tract. In addition, all 4 BP were also detected in the columnar epithelial cells of the apical portions of intestinal villi. In the intestinal superficial epithelial cells, lysozyme and β-defensins were detected in the ascending colon, whereas only β-defensin1 was detected in the descending colon and rectum. These results suggest that BP are mainly secreted from exocrine tissues in the initial portion of the digestive tract and play a role in host defense against indigenous bacteria throughout the digestive tract. Part of the BP in the chyme might be absorbed by the epithelium at the most inner sites of mucosae in the small and large intestines.
The epidemiological information has obtained on avian influenza virus (AIV) in eastern Hokkaido, Japan, where AIV surveillance has not been performed. Cloacal or fecal samples obtained from migratory water birds were screened for AIV both by real-time reverse transcriptase polymerase chain reaction to detect the influenza A virus matrix (M) gene and by egg inoculation. Between 2007 and 2009, a total of 2,488 samples were collected from various avian species in Abashiri, Kushiro, Nemuro and Tokachi districts of eastern Hokkaido. AIVs were isolated from 18 of those samples (0.7%). No AIV was isolated from the 1,449 samples collected in Abashiri, Kushiro and Nemuro districts, although 6 were positive for the M gene by RRT-PCR. In contrast, 52 (5.0%) of the 1,039 samples collected from ducks in Tokachi district were M gene positive; AIVs were isolated from 18 of those samples (1.7%). The isolates included H3N5 (1 isolate), H3N6 (1), H3N8 (9), H4N2 (1), H4N6 (2), H6N5 (1), H6N8 (1), and H11N3 (2) subtypes. H3N5 and H11N3 subtypes have not been frequently isolated, and our study is the first to report H3N5 and the second to report H11N3 in Japan. Phylogenetic analysis revealed that the M genes of all isolates belonged to the Eurasian lineage.
Erysipelothrix rhusiopathiae is pathogenic for humans, many domestic animals and wild birds, but infectious cases with clinical symptoms in cats have not been reported. E. rhusiopathiae was recovered from a 4-month Russian blue breed cat with a very poor body condition score of 1 (BCS: 1/5). The isolate was typed as serotype 2b. Mice experimentally infected with the clinical isolate of E. rhusiopathiae through subcutaneous or intraperitoneal routes survived, and the organism was recovered from the spleen and synovial and pericardial fluids. Cats experimentally inoculated with the isolate either orally or subcutaneously survived but commonly exhibited depression and emaciation together with localized erythemal lesion of the skin accompanied by purulent ocular discharge. On hematological analysis, the number of total white blood cells was high compared with that in normal cats. Histological examination revealed congestion and moderate inflammation with focal necrosis. This observation may provide insight on E. rhusiopathiae infection in cats with the possible epidemiological significance and implications as a potential source of infection to other animals and humans.
In total, 211 isolates of A. pleuropneumoniae were collected from pigs with hemorrhagic pneumonia at slaughterhouses during 2002-2007. Serotypes, antimicrobial susceptibility and minimum inhibitory concentration (MIC) values were determined for each isolate of A. pleuropneumoniae to 10 antimicrobial agents. Serovar 1 of A. pleuropneumoniae was predominant in Taiwan in 138 of the 211 isolates, followed by serovars 2 and 5. More than 90% of collected isolates were sensitive to ceftiofur, cephalothin, and chloramphenical. However, lincospectin and gentamicin were relatively less susceptible with sensitivities of only 2.4 and 5.7%, respectively. Additionally, ceftiofur had the highest in vitro activity with an MIC50 of 2.2 μg/ml, followed by cephalothin (2.7 μg/ml) and chloramphenicol (7.9 μg/ml). Lincospectin had the least activity with MIC50 and MIC90 values of 73.9 and 114.5 μg/ml, respectively. The data indicate that ceftiofur and cephalothin were extremely active against A. pleuropneumoniae and with minimum MIC values. These drugs are suitable for controlling and treating hemorrhagic pleuropneumonia outbreaks in swine.
Chlamydophila psittaci is the causative agent of human psittacosis and avian chlamydiosis. This zoonotic pathogen is frequently transmitted from infected birds to humans. Therefore proper and rapid detection of C. psittaci in birds is important to control this disease. We developed a method for detecting C. psittaci by using SYBR Green Real-time PCR based on targeting the cysteine-rich protein gene (envB) of C. psittaci. This one step procedure was highly sensitive and rapid for detection and quantification of C. psittaci from fecal samples. This assay was also able to detect other zoonotic Chlamydophila species such as C. abortus and C. felis. The assay is well suited for use as a routine detection method in veterinary medicine.
We examined antimicrobial susceptibility and efflux systems in laboratory-derived mutants of Salmonella enterica serovar Choleraesuis selected by culture on fluoroquinolone-containing plates. The mutants exhibited decreased susceptibilities to quinolones and several other antimicrobials. Mutations in the gyrA gene were not always found in the mutants. Accumulation assays revealed that intracellular enrofloxacin concentrations were significantly lower in the mutants compared with parent isolates. Increased expression of acrB mRNA can explain the decreased susceptibilities to several antimicrobials but not in the case of carbonyl cyanide m-chlorophenylhydrazone (CCCP). Decreased susceptibility to CCCP may result from the increased expression of emrA mRNA. These results suggest that the enhancement of multiple efflux pumps is responsible for decreased susceptibilities to several antimicrobials in the laboratory-derived mutants.
Saponin is the generic name of steroid or triterpene glycosides, and the capacities of some saponins to stimulate both Th1 immune response and production of cytotoxic T cells are useful as vaccine components against intracellular pathogens. Because saponins have been found commonly in starfish, we assessed the potential of starfish, Asterias amurensis and Asterina pectinifera, as adjuvant sources. Crude starfish saponins had hemolytic activities (EC50=10 to 100 μg/ml) and thin layer chromatography indicated heterogeneity of their constituents. When starfish saponis were subcutaneously injected into mice with ovalbumin (OVA), OVA-specific IgG, especially IgG2a instead of IgG1 was produced in mouse blood, suggesting starfish saponins stimulated Th1 type immunity and they were potential sources of new adjuvants.
We performed this study in order to establish an effective, simple and safe treatment for chytridiomycosis. The subjects were 12 amphibians (11 anurans of 4 different species and 1 urodela) diagnosed with chytridiomycosis by clinical signs and a PCR test. A 0.01% aqueous solution of the antifungal agent itraconazole was used to treat the subjects, and we evaluated the efficacy of treatment by 3 methods: clinical signs, direct microscopy and a nested PCR test. A 10-min immersion in a 0.01% aqueous solution of itraconazole every other day for a total of 7 treatments resulted in an improvement of clinical signs in 11 of the 12 cases. Specifically, we observed an abatement of increased sloughing and disappearance of zoosporangia by direct microscopy. DNA fragments of Batrachochytrium dendrobatidis were not detected by a PCR test at the end of treatment, nor were they detected after treatment (20-57 days following treatment; average, 34.4 days). No recurrence was observed 12 months after the end of treatment, nor did we observe any obvious side effects from itraconazole. Therefore, we recommend this as a treatment method for chytridiomycosis and as an elimination technique for use in captive amphibians.
CpG oligodeoxynucleotides (CpG-ODNs) are ligands for toll-like receptor 9 (TLR9), signaling of which plays a role in innate immunity by inducing T helper 1 (TH1)-cell responses and pro-inflammatory cytokine production. The activation of TLR9 signaling is considered to be effective for the therapy of cancer, infectious diseases, and allergies and preclinical studies using CpG-ODNs have been performed in dogs and humans. In order to investigate the precise mechanisms responsible for the effect of CpG-ODNs in dogs, we examined their role in cell proliferation and cytokine gene expression in canine B cells. Canine B cells were collected by a magnetic cell isolation method using anti-CD21 antibody. Flow cytometric analysis for the intracellular CD79α revealed the purity of canine B cells to be as high as 90.2 ± 2.1%. Transcription of TLR2, TLR4, and TLR9 mRNA on canine CD21+ cells was confirmed by reverse-transcript polymerase chain reaction (RT-PCR). CpG-ODNs induced dose-dependent proliferation of canine CD21+ cells (P<0.05 compared with control-ODNs) detected by BrdU incorporation. Quantification of IL-6, IL-10, and IL-12p40 mRNA transcription on canine CD21+ cells revealed that CpG-ODNs enhanced IL-6 mRNA transcription but not IL-10 and IL-12p40 mRNA transcription (P<0.05 compared with control-ODNs). These responses to CpG-ODNs in the canine B cells indicated that CpG-ODNs would be useful as an immunological adjuvant for vaccine in dogs.
The present study evaluated the effects of calcitriol dissolved in an oleaginous vehicle (calcitriol-OLE) on changes in plasma calcitriol, calcium and bone metabolic markers in nonpregnant, nonlactating Holstein cows. Five cows were treated intramuscularly or subcutaneously with calcitriol-OLE and oleaginous vehicle alone using a 5 × 5 Latin square design. Additionally, cows were also treated intravenously with calcitriol dissolved in an aqueous vehicle (calcitriol-AQU) for comparison. The plasma calcitriol concentrations after intramuscular and subcutaneous calcitriol-OLE administrations peaked at 24 and 12 hr, respectively, remained significantly elevated until day 3, returned to the respective control levels on day 5 and decreased significantly on day 7. In cows given intravenous calcitriol-AQU, the calcitriol levels decreased with linearity on day 1. The plasma calcium levels rose from 12 hr post-dose and peaked on day 2 for both preparations and in all three administration routes. Significantly increased calcium levels continued until day 5 in the intramuscular and intravenous routes and day 7 in the subcutaneous route. The plasma osteocalcin concentrations significantly increased from day 3 for calcitriol-OLE and from day 5 for calcitriol-AQU, whereas the bone resorption markers, tartrate-resistant acid phosphatase isoform 5b and hydroxyproline, decreased during this time. These results suggest that either intramuscular or subcutaneous injection of calcitriol-OLE extends and maintains supraphysiological calcitriol levels in the plasma and prolongs hypercalcemia. Moreover, exogenous calcitriol in normocalcemic cows increases the plasma osteocalcin concentration and decreases the plasma levels of bone resorption markers probably due to hypercalcemia.
The object of this study was to evaluate hypofractionated multiportal field and two-portion (rostral and caudal portions divided by the eyelid) radiation therapy for canine nasal tumors. Sixty-three dogs underwent multiportal hypofractionated radiation therapy. The radiation field was divided into rostral and caudal portions by the eyelid. Treatments were performed four times for 57 dogs. The median irradiation dose/fraction was 8 Gy (range, 5-10 Gy); the median total dose was 32 Gy (10-40 Gy). Improvement of clinical symptoms was achieved in 53 (84.1%) of 63 cases. Median survival time was 197 days (range, 2-1,080 days). Median survival times with and without destruction of the cribriform plate before radiotherapy were 163 and 219 days, respectively. There was no significant difference between them. No other factors were related to survival according to a univariate analysis. All radiation side effects, except one, were grade I according to the VRTOG classification. It was not necessary to treat any dogs for skin side effects. One dog (1.6%) developed an oronasal fistula 1 year after completion of radiation therapy. This radiation protocol may be useful in reducing radiation side effects in dogs with cribriform plate destruction.
A six year-old intact female miniature poodle was presented with a soft mass of the forehead region. Computed tomography identified generalized frontal bone loss and a large extracranial mass, which had a low attenuation area of hemorrhagic necrosis with septation and enhancement of solid components. In magnetic resonance imaging, the mass was isointense in T1-weighted images except its fluid parts and hyperintense in T2-weighted images with lobulated by low-signal septa. Surgery was performed to remove the mass, and histopathologic examination revealed that the mass was consistent with undifferentiated pleomorphic sarcoma (malignant fibrous histiocytoma). The dog died from the rapidly recurrent mass and severe pulmonary metastasis.
Three Japanese Black cows housed with 6 other cows exhibited main clinical symptoms of severe hemoglobinuria. Hematological analyses conducted after antibiotic therapy demonstrated severe anemia, and biochemical analyses indicated both severe hemolysis and disruption of hepatic function. One of the three cows died. Based on the above analyses and observation of typical clinical symptoms, a speculative diagnosis of bacillary hemoglobinuria was made, and immediate high-dose antibiotic treatment improved the general conditions of the surviving animals. Blood samples from the other 2 cows were collected sequentially after antibacterial therapy. Clostridium haemolyticum was detected by a nested polymerase chain reaction analysis of the blood samples. The cows were diagnosed with the second recorded occurrence of bacillary hemoglobinuria in Japan.
Stem cell therapy is being special premise for various renal diseases. However, there is limited literature on localization and pathologic and functional effects of allogenic mesenchymal stem cells (MSCs) in healthy dogs. Two healthy dogs were included in this study. Canine MSCs (cMSCs) were cultured from canine bone marrow and incubated with superparamagnetic iron oxide (SPIO) for in vivo cell tracking via MR imaging. The dogs were given the MSC (3 × 106 cells) into a renal artery via femoral artery catheterization. Follow-up serial renal assessments included ultrasonography and MRI, serum chemistry, urine analysis, and renal clearance tests. The dogs were euthanized at days 8 and 35 respectively for histopathologic evaluation of kidney. Strong hypointensity in MRI was detected in the treated renal cortex the day after cMSCs infusion. However they disappeared from MR image by the 8th day. Of the serum chemistry tests, serum hepatic enzymes (ALT, AST) were significantly elevated for one week after cMSCs treatment. Histopathological findings also revealed infiltration of SPIO-containing cells into the parenchyma of kidney. On 35th day, histopathology, glomerular atrophy, tubular necrosis, and mineralization were found in the subcapsular cortex, with fibrosis of the interstitial tissues. In vivo MRI studies of stem cells were useful in determining the sequential location of stem cells in the renal parenchyma of healthy dogs. Allogenic stem cells administered via renal artery caused inflammation, tubular necrosis, mineralization, and fibrosis without functional complications.
The Spontaneously Diabetic Torii (SDT) fatty rat, established by introducing the fa allele of the Zucker fatty rat into the SDT rat genome, is a new model of obesity/type 2 diabetes. The present study investigated effects of food restriction on metabolic and endocrinological function in SDT fatty rats. SDT fatty rats were pair-fed with SDT rats from 7 to 21 weeks of age. The SDT fatty rats were already hyperinsulinemic and hyperlipidemic at 7 weeks of age. After 7 weeks of age, SDT fatty rats showed age-dependently increasing serum glucose levels associated with decreasing serum insulin levels. However, in pair-fed SDT fatty rats, hyperglycemia and hyperinsulinemia were attenuated at 9 weeks of age. After 9 weeks of age, the serum insulin levels unexpectedly increased in the pair-fed SDT fatty rats. Glucose tolerance was also improved, and the pancreatic insulin contents were increased in these rats. Pancreatic islets were hypertrophied in pair-fed SDT fatty rats compared with ad lib-fed SDT fatty rats, which were comparable to SDT rats. This study showed that, in SDT fatty rats, calorie restriction by paired-feeding with SDT rats attenuated hyperglycemia and hyperinsulinemia for the first 2 weeks. Thereafter, the serum insulin levels and pancreatic insulin contents were increased, though the restriction was continued. Hypertrophic pancreatic islets were also remarkable, indicating increased beta cell proliferation. The activated pancreatic beta cell functions might be due to rapid food ingestion, a change of feeding behavior resulting form increasing the fasting period, which was indispensable for calorie restriction.
Zinc (Zn) is an essential trace element for DNA synthesis and for cell growth and differentiation. The deficiency induces a wide range of disorders including immunodeficiency. In this study, the influence of Zn deficiency to the mice infected with Babesia microti was examined, and was compared with the influence in the rats infected with B. rodhaini previously reported. Experiments of B. microti infection were conducted using Zn-deficient (ZD; allowed to eat ad libitum on the ZD diet), Zn-adequate (ZA; allowed to eat ad libitum on the ZA diet), and diet-restricted (DR; supplied 2 g/day on the ZA diet) mice. It was suggested that the Zn deficiency exacerbated the infection dynamics of the mice with B. microti by the growth retardation, the reduction of immunity and the decrease in PCV. The results in the mice supported the consequences in the rats previously reported.
Clinical grape poisoning in two dogs (a 1.6-year-old male Shih Tzu and a 5-year-old female Yorkshire Terrier) was described in the present study. Clinical signs included decreased urine output in the Shih Tzu and ataxia in the Yorkshire Terrier after grape ingestion. The Shih Tzu died 5 days post-grape ingestion, while the Yorkshire Terrier died 3 days post-grape ingestion. Erythematous serosae and mucosae, multifocal red small intestinal foci, and blood and grape seeds were identified in the intestinal lumen. Brownish-yellow crystals were bilaterally identified in the renal pelvis. The primary histological findings were acute tubular necrosis of the proximal convoluted tubules, severe necrosis, and mineralization in the renal cortical tubules. Blood urea nitrogen, creatinine, and alanine aminotransferase were increased in the dogs. Many Korean veterinary clinicians have suspected clinical grape poisoning. However, to our knowledge, grape poisoning has not been identified by pathologic and clinicopathologic basis until this writing in Korea. Education and knowledge about the risks of grape poisoning is necessary for the prevention of accidental exposures.
Multiple yellowish-white, cauliflower-like mass lesions on the skin of the head and back in a 4-month-old piglet were pathologically examined. These lesions had developed before the weaning period. Histologically, the cutaneous neoplasms were characterized by papillary outgrowth of connective tissue covered by thick epidermis. Hyperplasia of the epidermis was corresponded with proliferation of capillaries, lympho-plasmacytic infiltration, and proliferation of fibroblasts in the dermal stroma. There were no inclusion bodies and significant degeneration in the keratinocytes. Papillomavirus antigen and DNA were not detected in these lesions by immunohistochemistry and polymerase chain reaction, respectively. Accordingly, the fibropapillomatosis of the present case might be hamartomatous rather than infectious.
A total of 15 isolates of Salmonella enterica serovar 4,5,12:i:- obtained from diseased cows and patients in Iwate Prefecture, Japan, were characterized to clarify the genetic basis of this serovar. S. Typhimurium- specific IS200 was detected from all the isolates. A 94-kb plasmid and the spvB gene were detected from all but one of the 15 isolates. The results of PCR mapping of the fljAB operon and its flanking regions indicate that there are deletions or mutations in this chromosomal region. These data suggest that the 15 isolates are monophasic variants of S. Typhimurium. Epidemiological relationships between the isolates obtained from cattle and humans were not suspected based on the comparison of data employing plasmid profiling and pulsed-field gel electrophoresis.
Reproduction of the masked palm civet (Paguma larvata) has not been well investigated in Japan. We examined 361 female masked palm civets harvested as nuisance animals between April 2007 and March 2009 in Kanagawa Prefecture and Tokyo Metropolis. Pregnant animals and placental scars-bearing animals were found only in 12 months old and over. In these animals, the observed rate of pregnant animals was 13.7% (29/212) and that of placental scar-bearing animals was 29.2% (62/212). The number of fetuses ranged from 1-4 (average 2.8), and the principal months of parturition estimated from the crown-rump lengths of fetuses were from March to November.
Cats show repeated copulation, but changes in semen qualities and quantities with repetition of ejaculation have not previously been clarified. We collected semen 4 times consecutively from 5 cats using the artificial vagina method and observed the semen qualities and quantities. No significant changes were noted in the semen volume, frequency of abnormal sperm or incidence of immature sperm, but the number of sperm and sperm motility and viability decreased with repetition, and in particular, the number of sperm in the first semen accounted for 55.0% of the total number in the 4 consecutive ejaculations, showing a significant difference from those in the 2nd-4th semen (P<0.01).
The mean post-thaw sperm motilities of feline frozen semen prepared with 1% OEP or 3 g/ml SLS as a cryoprotective agent, in addition to 7% glycerin, were 35.0 ± 2.4 and 37.0 ± 2.5%, respectively, showing no significant difference. On unilateral intrauterine insemination (UIUI) using these semen samples at a sperm number of 40 × 106, the conception rate was 70.0% (7/10) in the OEP group and 30% (3/10) in the SLS group, showing that the rate was higher in the OEP group, but the difference was not significant. It was suggested that sperm in frozen semen showing the above qualities were transferred to the contralateral uterine horn on UIUI.
The study was designed to explore the toxic effects of arsanilic acid on piglet Sertoli cells. Sertoli cells were isolated from piglet testes using a two-step enzyme digestion followed by differential plating. Piglet Sertoli cells were cultured and classified into the following five groups: group A, the control without arsanilic acid treatment; group B, cultured with 5 μM arsanilic acid; group C, cultured with 50 μM arsanilic acid; group D, cultured with 0.5 mM arsanilic acid; and group E, cultured with 5 mM arsanilic acid. We found that Sertoli cell growth was inhibited by arsanilic acid at 0.5 mM compared with the control, group A. The oxidase activity of Sertoli cells was decreased by arsanilic acid at 0.5 mM as evidenced by the observations that arsanilic acid increased MDA content but decreased the SOD and GSH-Px activities of Sertoli cells. Moreover, 50 μM of arsanilic acid was observed to cause DNA damage in Sertoli cells. The results of our study suggest that exposure of Sertoli cells to arsanilic acid leads to induction of oxidative stress and inhibition of cell growth at a high concentration, while arsanilic acid causes DNA damage in Sertoli cells at a low concentration.