Our previous studies and others have revealed detailed characteristics of the mucosal nerve network in the small intestine, but much remains unknown about the corresponding network in the large intestine. We herein investigated regional differences in the expression of neurochemical markers, the nerve network structure, and the cells in contact with nerve fibers by histological analysis using both immunohistochemistry and serial block-face scanning electron microscopy (SBF-SEM). Immunohistochemistry revealed that immunopositive structures for protein gene product 9.5, vasoactive intestinal peptide (VIP), calretinin and vesicular acetylcholine transporter were more prevalent in the lamina propria of the ascending colon than the cecum and descending colon (DC). There was no significant difference in the frequency of most neurochemical markers between the cecum and DC, but the frequencies of VIP⁺ structures were higher in the cecum than in the DC. SBF-SEM analysis showed that the nerve network structure was more developed on the luminal side of the DC than the cecum. The cells that nerve fibers abundantly contacted were subepithelial and lamina propria fibroblast-like cells and macrophages. In addition, nerve fibers in the cecum were in more frequent contact with immune cells such as macrophages and plasma cells than nerve fibers in the DC. Thus, the present histological analysis suggested that the mucosal nerve network in the large intestine possessed both regional universality and various specificities, and revealed the intimate relationship between the nerve network and immune cells, especially in the cecum.

Coronary artery anomalies that can cause sudden cardiac death have been described in some mammals. However, few studies have investigated coronary anomalies and coronary artery branching patterns in cattle. Therefore, understanding of bovine coronary arteries is incomplete. The aim of this study was to clarify anatomic variants in coronary arteries of Japanese Black cattle. The gross anatomy of the coronary arteries was examined in the hearts of 100 cattle. A total of 14 of 100 hearts exhibited confirmed coronary anomalies in the left main trunk. These 14 hearts were classified into 5 types potentially associated with cardiac ischemia. Regarding the coronary branches, the subsinuosal interventricular branch originated from the left circumflex branch (LCX) in 98 hearts and from the right coronary artery in 2 hearts. The origin of the sinoatrial nodal branch was classified into 3 types. In 99 hearts, the atrioventricular nodal branch originated from the LCX at the heart crux. This study characterized the prevalence of coronary anomalies, branching patterns of coronary arteries, and coronary artery anatomic variants in Japanese Black cattle.

This study aimed to examine the antimicrobial susceptibility of bovine mastitis pathogens in Japan and develop criteria for testing antimicrobial susceptibility using the simplified agar disk diffusion (ADD) method that is currently being used in clinical practice. Milk samples from 1,349 dairy cows with clinical mastitis were collected and cultured. The minimum inhibitory concentrations (MICs) of the antimicrobials were determined for 504 strains of 28 bacteria. Of the gram-positive bacteria, most Staphylococcus spp. were susceptible to penicillin G (PCG), kanamycin (KM), oxytetracycline (OTC), cefazolin (CEZ), pirlimycin, enrofloxacin, and marbofloxacin. Streptococcus spp. and Trueperella pyogenes showed resistance to OTC and KM. Most gram-negative bacteria were resistant to OTC and CEZ and particularly susceptible to fluoroquinolones. To develop the criteria for a disk diffusion test of the simplified ADD method, the relationships between MICs and diameters of inhibition zones (DIZs) were analyzed and compared with the conventional method. The susceptibility breakpoints of several antimicrobials were lower for both gram-positive and gram-negative bacteria. Particularly for gram-positive bacteria, the application of the new criteria lowers the breakpoint for PCG, suggesting that the use of PCG instead of CEZ may increase. The results suggest that use of these criteria for the simplified ADD method may lead to appropriate antimicrobial choice and consequently the appropriate use of antimicrobials in clinical practice.

The dissemination of mcr-harboring Enterobacteriaceae, e.g., Escherichia fergusonii, with resistance to colistin via animal products is a public health concern. In our previous study, E. fergusonii harboring the mcr gene were isolated from 11 pigs and 43 chickens. To understand the spread of mcr-harboring E. fergusonii in Okinawa, Japan, and to gain further insights into how they can be controlled, an antimicrobial susceptibility testing, pulsed-field gel electrophoresis (PFGE), a conjugation test for the transferability of mcr-harboring plasmids, and PCR-based replicon typing (PBRT) were performed using the 54 strains. According to the disk-diffusion and broth microdilution methods, 9 of the 11 strains from pigs and 9 of the 43 strains from chickens had multidrug resistance (MDR). The broth microdilution method showed that all strains were resistant to colistin, and the minimum inhibitory concentration of colistin was 4–16 μg/mL. PFGE suggested identical PFGE types were being transmitted within one pig farm, within one chicken farm, and among several chicken farms. These findings showed that some mcr-harboring E. fergusonii in Okinawa exhibited MDR, and these had spread within farms and between farms. In the mcr gene conjugation test and PBRT, a type IncI2 plasmid replicon was detected in all mcr-1-harboring transconjugants. Therefore, evidence suggests that the IncI2 plasmid is probably involved in the transmission of the mcr-1 gene. It is important to monitor the antimicrobial resistance profile and dissemination of the IncI2 plasmid in mcr-harboring E. fergusonii.

We describe phenotypic and genetic characterization of an atypical Japanese Actinobacillus pleuropneumoniae isolate OT761. Nucleotide sequence analysis revealed that gene clusters involved in capsular polysaccharide and O-polysaccharide (O-PS) biosynthesis of the isolate were nearly identical to those of serovar 2 reference strain. The main difference found between the O-PS loci is the shortening of 31 amino acids from the C terminus of WcaJ in the atypical isolate due to a 93 bp deletion at the 3' end of wcaJ gene. Immunoblot analysis revealed that this isolate could not produce O-PS. Taken together, our results showed that the C-terminal domain of the A. pleuropneumoniae WcaJ plays a critical role in enzyme function of WcaJ involved in the biosynthesis of O-PS.

Intestinal lymphangiectasia (IL) is a common complication in dogs. This study analyzed intestinal microbiota using 16S rRNA amplicon analysis as candidate factors that strongly influence the small intestinal lymphatic vessels in dogs with and without IL. Twelve dogs were included, of which six were diagnosed with lymphoplasmacytic enteritis, four with small-cell lymphoma, and two with large-cell lymphoma. Seven of these dogs had IL, whereas five did not. First, the microbial diversity analyzed by Faith pd index was significantly decreased in dogs with IL compared to dogs without IL. Then, the relative amounts of each bacterial taxa were compared between dogs with and without IL using Linear discriminant analysis effect size analysis. At the genus level, the Ruminococcus gnavus group significantly increased in dogs with IL compared to dogs without IL. A total of four genera, including Ruminococcus torques group and Faecalibacterium, which produce butyrate, significantly decreased in dogs with IL. This study showed decreased intestinal bacterial diversity and several alterations of intestinal microbiota, including a decrease in butyrate-producing bacteria in dogs with IL, compared to dogs without IL.

This study aimed to prospectively assess nasopharyngeal luminal changes in brachycephalic dogs without respiratory signs using cervical radiography. Forty brachycephalic dogs without cardiopulmonary diseases were included. The nasopharyngeal luminal change was calculated on inspiratory and expiratory cervical lateral radiographs. The median nasopharyngeal luminal change was 21.6% (range, 0.3–85.6%). In five dogs, a nasopharyngeal luminal change of >50% was identified. There was no correlation between nasopharyngeal luminal changes and age, sex, body weight, or body condition score. These results suggest that brachycephalic dogs without cardiopulmonary diseases may be over-diagnosed with partial pharyngeal collapse. Further studies comparing nasopharyngeal luminal changes between clinically healthy brachycephalic dogs and dogs with respiratory signs are warranted.

Inflammatory bowel disease (IBD) is classified into two types: Crohn’s disease and ulcerative colitis. In IBD, the imbalance between the pro-inflammatory and anti-inflammatory cytokines prevents recovery from the inflammatory state, resulting in chronic inflammation in the colon. The mitotic spindle positioning protein (MISP) is localized to the apical membrane in the colon. In this study, we observed increased expression of MISP in the intestinal epithelial cells in dextran sulfate sodium (DSS)-induced colitis in mice. MISP-deficient mice receiving DSS showed significant exacerbation of colitis (e.g., weight loss, loss of the crypts). The intestinal epithelial cells of the MISP-deficient mice showed a trend towards decreased cell proliferation after DSS treatment. Reverse transcription followed by quantitative polymerase chain reaction revealed that the expression levels of Tgfb1, an anti-inflammatory cytokine, were significantly reduced in the colon of MISP-deficient mice compared with the wild-type mice regardless of DSS treatment. These findings indicate that MISP may play a role in the recovery of the colon after inflammation through its anti-inflammatory and proliferative activities, suggesting that MISP may be a new therapeutic target for IBD.

The detailed reproductive physiology of cockatoos based on gonadal hormone dynamics is unclear. In this study, we aimed to investigate ovarian activity by monitoring urofecal sex steroid hormone profiles in a captive female white cockatoo (Cacatua alba) and to noninvasively reveal basic reproductive physiology by comparing the hormone profiles with the laying dates, body mass changes, and molt progress. Urofeces were collected regularly for approximately 4 years from one female that frequently laid unfertilized eggs under single-rearing conditions. Urofecal progesterone (P₄) and estradiol-17β (E₂) were measured by enzyme immunoassay. In addition, body mass and the number of fallen feathers were measured periodically. The urofecal P₄ concentration peaked at an average of 17.7 days after the start of the rise in urofecal E₂ concentration, and egg laying began on the day after the peak urofecal P₄ concentration. The clutch size was usually two eggs, with an average interval of 4.5 days between eggs in each egg-laying cycle. There was a significant correlation between the dynamics of E₂ concentration in urofeces and body mass. The results strongly suggest that E₂ and P₄ reflect the follicle growth and ovulation status, respectively, and that noninvasive monitoring of hormone dynamics using urofeces can accurately capture ovarian activity in the white cockatoo. Furthermore, changes in body mass can predict follicular growth, and reproduction and molt are antagonistic.

Amitriptyline intoxication is caused by its suicidal or accidental overdose. In the present study, by intravenously injecting 1.5 or 3.0 mg/kg amitriptyline into bullfrogs, we actually revealed that amitriptyline causes the widening of QRS complexes in electrocardiogram (ECG). In simultaneous recordings of the cardiac action potential, amitriptyline decreased the slope of phase 0 in the action potential, indicating the inhibition of the inward sodium currents during this phase. The following treatment with sodium bicarbonate quickly restored the widened QRS complexes in the ECG, demonstrating the counteraction with the sodium channel blockade caused by amitriptyline. The dual recordings of ECG waveforms and the action potential in cardiomyocytes enabled us to demonstrate the mechanisms of characteristic ECG abnormalities caused by amitriptyline intoxication.

An intact, male mixed-breed cat approximately 3 months of age was referred to Iwate University Veterinary Teaching Hospital with dysuria that developed after a traffic accident. At the initial visit, a cystostomy tube was placed as a temporal urinary diversion. Antegrade urethrography revealed severe stenosis of the intrapelvic urethra. A prepubic urethrostomy (PPU) with formation of a convex urethral stoma was performed 22 days after the first visit. Twenty-four months after surgery, the cat could urinate smoothly in a standing position, although a small amount of intermittent leakage was observed immediately after getting up. There was no evidence of peristomal dermatitis. Therefore, the formation of a convex urethral stoma may effectively prevent peristomal dermatitis in cats who undergo PPU.

Dichlorodiphenyltrichloroethane (DDT) is an organochlorine insecticide used worldwide. Several studies have reported the toxic effects of DDT and its metabolites on steroid hormone biosynthesis; however, its environmental effects are not well understood. This study examined wild rats collected in DDT-sprayed areas of South Africa and quantified plasma metabolites using liquid chromatography quadrupole time-of-flight mass spectrometry (LC-Q-TOF-MS). Fold change analysis of the metabolome revealed the effect of DDT on bile acid biosynthesis. Gene expression of the related enzyme in rat liver samples was also quantified. Significant association was found between DDT and gene expression levels related to constitutive androstane receptor mediated enzymes, such as Cyp2b1 in rat livers. However, our results could not fully demonstrate that enzymes related to bile acid biosynthesis were strongly affected by DDT. The correlation between DDT concentration and gene expression involved in steroid hormone synthesis in testis was also evaluated; however, no significant correlation was found. The disturbance of metabolic enzymes occurred in rat liver in the target area. Our results suggest that DDT exposure affects gene expression in wild rats living in DDT-sprayed areas. Therefore, there is a need for DDT toxicity evaluation in mammals living in DDT-sprayed areas. We could not find an effective biomarker that could reflect the mechanism of DDT exposure; however, this approach can provide new insights for future research to evaluate DDT effects in sprayed areas.

Locoweeds, a type of poisonous weedare, are widely distributed throughout the world and have a significant impact on the development of herbivore animal husbandry. Swainsonine (SW), the main toxin in locoweeds, can competitively inhibit lysosomes α-mannosidase (LAM) in animal cells, resulting in α-mannosidosis. However, the specifics of the interaction between SW and LAM are still unclear. Here, we used molecular docking to predicte the interaction points between SW and LAM, built mutated lysosomes α-mannosidase (LAM^M), and analyzed its biochemical properties changes in presumption points. The Trp at the 28th position and the Tyr at the 599th position of the LAM were interaction point candidates, and the above two amino acids in Capra hircus LAM (chLAM), were successfully mutated to glycine by constructing recombinant yeast GS115/PIC9K- LAM^M. The results showed that the sensitivity of Capra hircus LAM^M (chLAM^M), to SW decreased significantly compared with wild-type LAM, the enzyme activity of LAM decreased approximately threefold, the optimum temperature of LAM^M decreased from 55°C to 50°C, the optimum pH value increased from 4.5 to 5.0, and the effects of Mn²⁺, Fe³⁺, Al³⁺, Co²⁺, Cr³⁺, and ethylenediaminetetraacetic acid (EDTA) on LAM enzyme activity before and after point mutation changed significantly. These findings help us better understanding the molecular mechanism of the interaction mechanism between SW and chLAM, and provide new reference for solving locoweeds poisoning.

Type 1 recombinant enterovirus G (EV-G), which carries the papain-like cysteine protease (PLCP) gene of torovirus between its 2C/3A regions, and type 2 recombinant EV-G, which carries the torovirus PLCP gene with its flanking regions having non-EV-G sequences in place of the viral structural genes, have been detected in pig farms in several countries. In a previous study, we collected 222 fecal samples from 77 pig farms from 2104 to 2016 and detected one type 2 recombinant EV-G genome by metagenomics sequencing. In this study, we reanalyzed the metagenomic data and detected 11 type 2 recombinant EV-G genomes. In addition, we discovered new type 2 recombinant EV-G genomes of the two strains from two pig farms samples in 2018 and 2019. Thus, we identified the genomes of 13 novel type 2 recombinant EV-Gs isolated from several pig farms in Japan. Type 2 recombinant EV-G has previously been detected only in neonatal piglets. The present findings suggest that type 2 recombinant EV-G replicates in weaning piglets and sows. The detection of type 1 recombinant EV-Gs and type 2 recombinant EV-Gs at 3-year and 2-year intervals, respectively, from the same pig farm suggests that the viruses were persistently infecting or circulating in these farms.

Biosecurity enhancement contributes to the reduction of various microbial pathogens. Mammalian orthoreoviruses (MRVs) which are increasingly recognized as potentially serious problems on swine industry were used as indicators of biosecurity enhancement on two pig farms. Twelve MRVs were detected and isolated from fecal specimens of healthy pigs collected from one of the two farms in Japan. By sequencing based on the partial S1 gene, MRV isolates were classified as MRV1 and MRV2. Additionally, the virucidal activities of disinfectants toward the isolated MRV1 were evaluated using quaternary ammonium compound (QAC) diluted 500 times with water (QAC-500), 0.17% food additive glade calcium hydroxide (FdCa(OH)₂) solution, QAC diluted with 0.17% FdCa(OH)₂ solution (Mix-500), sodium hypochlorite at 100 or 1,000 parts per million (ppm) of total chlorine (NaClO-100 or NaClO-1000, respectively). To efficiently inactivate MRV1 (≥3 log₁₀ reductions), 0.17% FdCa(OH)₂, Mix-500 and NaClO-1000 required 5 min, whereas it took 30 min for QAC-500. The number of MRV detections has decreased over time, after using Mix-500 for disinfection on the positive farm. These results suggest that different serotypes of MRVs are circulating among pigs, and that the occurrence of MRVs in the farms decreased consequent to more effective disinfection.

In the prostate gland of the raccoon (Procyon lotor), the morphological appearance of the epithelial cells, such as basal and luminal cells, and the expressions of p63, androgen receptor (AR), and proliferating cell nuclear antigen (PCNA) were examined histologically and immunohistochemically to clarify their seasonal dynamics throughout the year. In this study, the regression with luminal cell defluxion and the regeneration process of the prostatic glandular epithelium was revealed in the seasons with declined spermatogenesis (June to August). The expression of p63 was observed only in the basal cells. AR immunoreactivity in the luminal cells was shown in the developed and regenerating (close to developed) prostates, whereas the basal cells exhibited AR immunoreactivity all year round. PCNA expression was rare in epithelial cells of the developed prostate gland. In the regressed gland, the basal cells demonstrated proliferative ability, whereas PCNA of the luminal cells appeared for the first time in the regenerating phase. This study is the first to clarify the regression with luminal cell defluxion and restoration and the seasonal dynamics of AR expression and proliferative activity in the prostate gland of seasonal breeders.

Lameness has a significant impact not only on the economy but also on elephant welfare. Several gait characteristics are altered to compensate for the discomfort. The traditional approach to detecting lameness has relied on mahout and veterinarian observation. The study aimed to determine how lameness affected the variability of an elephant’s gait by using a three-dimensional inertial measurement unit (IMU) with Wi-Fi sensors. Five elephants with lameness, as determined by an experienced veterinarian and two, non-lamed elephants, aged between 58–80 years were included in the study. Gait biomechanics including limb segment motion, obtained from individually gyrometric- and accelero-based parameters and demonstrated as a graphical pattern showing changes in absolute rotation angle over time. The result revealed some character changes in gait kinematics parameters, but it was heterogeneity with an inconclusive pattern. The interlimb coordination could be a part of maintaining the actual locomotion pattern, or it could be a result of the mild degree of lameness for which all of the clients have compensated. This study introduces a new objective method for quantifying gait changes caused by lameness, additional research is required to make this objective more clinically applicable.

A 14-year-old, male sugar glider presented with lethargy, anorexia, diarrhea, and paralysis of the hind limbs, and ultrasonography showed possible liver dysfunction. Some medications were administered, but the animal died 10 months after the first presentation, and a necropsy was performed. Histopathologically, hepatocellular degeneration and necrosis, severe deposition of hemosiderin in Kupffer cells and hepatocytes, bridging fibrosis, and regenerative nodules were observed in the liver. Variably amounts of hemosiderin deposition was observed in the heart, lungs, spleen, and kidney. These findings led to the diagnosis of hemochromatosis. No sugar glider cases with hemochromatosis have been reported. The pathological characteristics of hemochromatosis in this species were documented for the first time. The pathogenesis of hemochromatosis in animals remains unclear, but it has been suggested that some commercially available food for sugar gliders containing excessive amounts of iron and vitamin C may induce the disease.

The approved Japanese measurement method of circulating alkaline phosphatase (ALP) has changed from that of the Japan Society of Clinical Chemistry (JSCC) to that of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC). We measured the serum levels of total ALP (t-ALP) and those of the isoenzymes ALP2 and ALP3 in 50 Asian elephant (Elephas maximus) specimens using both methods. The activities determined by the IFCC method were roughly one-third lower than those determined by the JSCC method. We present conversion formulae. Our results enable comparisons of historical and current data on serum ALP activities in endangered, zoo-managed Asian elephants.