A case of nontraumatic gas gangrene in a common marmoset (Callithrix jacchus)

The common marmoset is widely used in neuroscience and regenerative medicine research. However, information concerning common marmoset disorders, particularly infectious diseases, is scarce. Here, we report a case of a female common marmoset that died suddenly due to gas gangrene. The animal presented with gaseous abdominal distention at postmortem, and Clostridium perfringens type A was isolated from several tissues. Vacuoles, a Gram-positive bacteremia and intravascular hemolysis were observed microscopically in the muscles, liver and lungs. On the basis of these findings, we diagnosed nontraumatic gas gangrene caused by Clostridium perfringens type A in this common marmoset.     

Chordoma of the thoracic vertebrae in a Bengal tiger (Panthera tigris tigris)

A 19-year-old female Bengal tiger (Panthera tigris tigris) was presented with hind limb weakness, ataxia and respiratory distress. Computed tomography revealed a mass between the left side of the T7 vertebra and the base of the left 7th rib. The tiger then died, and necropsy was performed. Grossly, the vertebral mass was 6 × 5.7 × 3 cm, and invaded the adjacent vertebral bone and compressed the T7 spinal cord. Histologically, the mass was composed of large, clear, vacuolated and polygonal cells with osteochondral matrix. Cellular and nuclear atypia were moderate. The vacuolated cells stained positively for cytokeratin and vimentin and negatively for S-100. Based on these findings, the present case was diagnosed as a vertebral chordoma; the first report in a tiger.     

A Real-Time PCR for Detection and Quantification of Mycoplasma ovipneumoniae

A real-time PCR for detection and quantification of M. ovipneumoniae was developed using 9 recently sequenced M. ovipneumoniae genomes and primers targeting a putative adhesin gene p113. The assay proved to be specific and sensitive (with a detection limit of 22 genomic DNA) and could quantify M. ovipneumoniae DNA over a wide linear range, from 2.2 × 10² to 2.2 × 10⁷ genomes.     

Spontaneous Mammary Adenocarcinoma in a Twelve-week-old Female Sprague-Dawley Rat

Spontaneous mammary adenocarcinoma was observed in a 12-week-old female SD rat. A movable mass in the right cervical region was found at 11 weeks of age, and the rat was sacrificed the following week. The mass was located in the vicinity of the right salivary gland and measured 38 mm × 26 mm × 16 mm in gross size. It was a firm whitish mass, with a cut surface that was also whitish in appearance. Histopathologically, neoplastic cells formed glandular structures that contained secreted eosinophilic material. Ultrastructurally, similar secreted material and lipid droplets were in the cytoplasm of the neoplastic cells. Immunohistochemically, the neoplastic cells were positive for cytokeratin 8, cytokeratin 18 and estrogen receptor α. Based on these findings, the tumor was diagnosed as a mammary gland adenocarcinoma, and we therefore conclude that this tumor type can occur spontaneously in female SD rats as young as 12 weeks of age.     

Spontaneous Adenosquamous Carcinoma with Rapid Growth and EMT-like Changes in the Mammary Gland of a Young Adult Female BALB/c Mouse

This study histopathologically and immunohistochemically investigated a spontaneously occurring single mass subcutaneously located in the left lower abdomen of a female BALB/cAJcl−nu/+ mouse at 10 weeks of age. The mass was about 20 × 15 × 10 mm in size after formalin fixation; nevertheless, it was not detected by clinical observations at 9 weeks of age. H&E staining revealed the tumor origin was epithelial and probably arose from the mammary gland, and the tumor cells demonstrated a squamous, acinar or polyhedral/basal pattern. A cell kinetics analysis revealed that many of the tumor cells of the squamous, acinar or polyhedral/basal component were positive for PCNA and cyclin D1, although there were a few of TUNEL-positive tumor cells in all of the components. An epithelial/mesenchymal analysis demonstrated that most of the tumor cells of the squamous and acinar components contained keratin and E-cadherin; however, most of the tumor cells of the polyhedral/basal component were less or very weakly positive for these markers. The tumor cells of the squamous component were negative for vimentin and SMA; however, many of the tumor cells of the polyhedral/basal component exhibited vimentin. In addition, expression of SMA was confirmed in some tumor cells of the acinar and basal components. Based on the microscopic and immunohistochemical characterizations, the tumor was diagnosed to be adenosquamous carcinoma that originated from the mammary gland with rapid growth, and the tumor cells demonstrated epithelial-mesenchymal transition-like changes.     

Spontaneous extraskeletal osteosarcoma with various histological growth patterns in the abdominal wall of an ICR mouse

Extraskeletal osteosarcoma is extremely rare in mice. This case report demonstrates a spontaneous murine extraskeletal osteosarcoma that exhibited various histological growth patterns in an ICR mouse. At necropsy, the tumor mass was located in the abdominal wall and was 45 × 30 × 25 mm in size. Histopathologically, the tumor showed the following four growth patterns: a solid pattern of polygonal cells embedded in an osteoid eosinophilic matrix with calcification, an irregular sheet pattern of short spindle cells accompanying some eosinophilic multinucleated cells, a fascicular pattern of spindle cells and a cystic pattern lined by short spindle cells. Immunohistochemically, most of the tumor cells were positive for vimentin, proliferating cell nuclear antigen and osterix. The multinucleated cells mentioned above were desmin positive and were regarded as regenerative striated muscles but not tumor cells. Since no clear continuity with normal bone tissues was observed, the tumor was diagnosed as an “extraskeletal osteosarcoma.”     

Hypertrophic gastropathy associated with gastric sarcoma in a dog

A 14-y-old spayed female Labrador Retriever was presented with an 8-mo history of chronic vomiting. Abdominal ultrasound and gastrointestinal endoscopy revealed a mass protruding into the gastric lumen, with cytologic features suggestive of sarcoma. A partial gastrectomy was performed; the gastric body and antrum were thickened, with a cerebriform appearance of the mucosal surface. Histologic examination revealed a submucosal neoplastic proliferation of fusiform cells variably arranged in irregular bundles and scattered whorls. Fusiform cells strongly reacted to antibodies against vimentin, S100, and neuron-specific enolase; glial fibrillary acidic protein was moderately and multifocally expressed. Pancytokeratin, KIT, α–smooth muscle actin, and desmin were nonreactive. Histologic and immunohistochemical findings suggested a diagnosis of gastric sarcoma with features referable to a non-GIST (gastrointestinal stromal tumor), non–smooth muscle NIMT (non-angiogenic, non-lymphogenic intestinal mesenchymal tumor). The overlying gastric mucosa was thickened by elongated and dilated gastric glands, predominantly lined by intensely periodic acid-Schiff–stained mucous cells. This altered mucosal architecture was suggestive of Ménétrier-like disease. Although this disease has been hypothesized to predispose to gastric adenocarcinoma in dogs, an association with gastric sarcoma has not been documented previously in the veterinary literature, to our knowledge.     

Intraoperative Bleeding in Dogs from Grenada Seroreactive to Anaplasma platys and Ehrlichia canis

Background

Frequent exposure of Grenadian dogs to Rhipicephalus sanguineus results in Anaplasma platys, and Ehrlichia canis seroreactivity. During elective surgeries, substantial intraoperative hemorrhage occurs in some seroreactive dogs.

Objectives

To assess hemostatic parameters and bleeding tendencies as well as prevalence of PCR positivity in apparently healthy A. platys and E. canis seroreactive and seronegative free‐roaming dogs from Grenada.

Animals

Forty‐seven elective surgery dogs allocated to 4 groups: Seronegative control (n = 12), A. platys (n = 10), E. canis (n = 14) and A. platys, and E. canis (n = 11) seroreactive.

Methods

Preoperatively, hemostasis was assessed by platelet count, prothrombin time, activated partial thromboplastin time, and buccal mucosal bleeding time. Intra‐ and postoperative bleeding scores were subjectively assigned. Blood, spleen, bone marrow, and lymph node aspirates were tested by PCR.

Results

Bleeding scores in dogs coseroreactive for A. platys and E. canis were higher (P = .015) than those of seronegative dogs. A. platys DNA was amplified from 7/21 (33%) A. platys seroreactive dogs and from 1 E. canis seroreactive dog; E. canis DNA was amplified from 21/25 (84%) E. canis seroreactive dogs. E. canis DNA was amplified most often from blood, whereas A. platys DNA was amplified most often from bone marrow.

Conclusions and Clinical Importance

Apparently healthy, free‐roaming dogs coseropositive for A. platys and E. canis may have increased intraoperative bleeding tendencies despite normal hemostatic parameters. Future investigations should explore the potential for vascular injury as a cause for bleeding in these dogs. Improved tick control is needed for dogs in Grenada.     

In Vitro Fertilization and Development of Porcine Oocytes Matured in Follicular Fluid

This study was conducted to assess the fertilization and development of porcine oocytes matured in a solo follicular fluid (pFF) using different in vitro culture systems and insemination periods. Cumulus-oocyte complexes (COCs), follicular cells (FCs), and pFF were collected from the follicles of ovaries. The pFF was used as a maturation medium (MpFF) after supplementation with follicle stimulating hormone (FSH) and antibiotics. The COCs were matured in a 15 ml test tube containing 3.5 ml of MpFF with FCs (5.2 × 10⁶ cells/ml; rotating culture system) or 2 ml of MpFF without FCs in a 35-mm petri dish (static culture system) for 44 to 48 h. After maturation culture, oocytes were co-incubated with frozen-thawed spermatozoa for 5 h and then cultured for 7 days. The total mean rates of sperm penetration, normal fertilization, male pronucleus (MPN) formation, cleavage, and development to the blastocyst stage of oocytes after insemination were significantly higher (P<0.01) in the rotating culture system than in the static culture system. In conclusion, compared with the static culture system, the rotating culture system is adequate for the production of developmentally competent porcine oocytes when MpFF is used as a maturation medium.     

Addition of D-penicillamine,hypotaurine, and epinephrine (PHE) mixture to IVF medium maintains motility and longevity of bovine sperm and enhances stable production of blastocysts in vitro

The present study aimed to establish an efficient system for bovine embryo production by in vitro fertilization (IVF) that can achieve stable normal fertilization and blastocyst developmental rates in any bull without optimization of the sperm concentration in IVF medium. We examined the effects of a PHE mixture (20 μM D-penicillamine, 10 μM hypotaurine and 1 μM epinephrine), theophylline (2.5 mM), and sperm concentration (1, 2 or 5 × 10⁶ cells/ml) on fertilization and blastocyst developmental rates. High cleavage rates (78.3 to 92.4%) and blastocyst developmental rates (31.9 to 62.0%) at day 7 were obtained in the presence of PHE and theophylline in IVF medium with a sperm concentration of 2 × 10⁶ cells/ml using sperm from 9 bulls. In addition, the synergistic effect of PHE and theophylline on normal fertilization (2 pronuclei) was clarified at 12 h after IVF with a sperm concentration of 1 × 10⁶ cells/ml. Moreover, high linearity, high flagellar beat cross frequency, and low amplitude of lateral head of motile sperm were found by computer-assisted sperm analysis. In conclusion, the combination of the PHE mixture and theophylline synergistically accelerates sperm motility and sperm penetration of bovine oocytes. Theophylline activates sperm motility with increasing intracellular cAMP. However, PHE prevents an excessive increase of cAMP and maintains sperm motility without hyperactivation. When the combination of PHE and theophylline is added to IVF medium at a sperm concentration of 2 × 10⁶ cells/ml, we can achieve stable normal fertilization and blastocyst development in any bull.     

Classification of the spermatogenic cycle,seasonal changes of seminiferous tubule morphology and estimation of the breeding season of the large Japanese field mouse (Apodemus speciosus) in Toyama and Aomori prefectures, Japan

The large Japanese field mouse, Apodemus speciosus, is a potential indicator of environmental stress, but this function has not been confirmed by histological studies. Since environmental stress affects the reproductive function of mice, we determined the reproductive characteristics of this species at two locations: Toyama (36°35ʹN, 137°24ʹE) and Aomori (40°35ʹN, 140°57ʹE). Mice were captured during May–November (n=119) and July–November (n=146) at these locations, respectively. We classified the breeding season from the numbers of pregnant females and young, in addition to the spermatogenic cycle and seasonal changes in seminiferous tubule morphology of males. Testicular weight was measured, and seminiferous tubule morphology was examined histologically. Fourteen stages were found in the seminiferous epithelium cycle based on acrosome formation and spermatid head morphology. At both locations, the breeding season peaked from late summer to early autumn and possibly in spring. Spermatogenic activity was classified into 4 periods from June to November: resting around June and October–November; resumptive around July; active around August; and degenerative around September. During the resting period, the seminiferous tubules consisted of Sertoli cells, spermatogonia and spermatocytes. Spermatogenesis began during the resumptive period, and spermatids were observed. During the active period, active spermatogenesis and a broad lumen were observed. During the degenerative period, spermatogenesis ended, and Sertoli cells, spermatogonia, spermatocytes and degenerating exfoliated round spermatids were observed. This study provides scientific information about the testicular histopathological evaluations of the large Japanese field mouse for its use as an index species of environmental pollution.     

Anethole inhibits growth of recently emerged multidrug resistant toxigenic Vibrio cholerae O1 El Tor variant strains in vitro

To search natural compounds having inhibitory effect on bacterial growth is important, particularly in view of growing multidrug resistant (MDR) strains of bacterial pathogens. Like other bacterial pathogens, MDR Vibrio cholerae, the causative agent of diarrheal disease cholera, is becoming a great concern. As an approach of searching new antimicrobial agents, here, we show that anethole, a well-studied natural component of sweet fennel and star anise seeds, could potentially inhibit the growth of MDR O1 El Tor biotype, the ongoing 7th cholera pandemic variant strains of toxigenic V. cholerae. The minimum inhibitory concentration (MIC) of anethole against diverse O1 El Tor biotype strains is evaluated as 200 µg/ml. Moreover, the effect of anethole is bactericidal and exerts rapid-killing action on V. cholerae cells. This study is the first report which demonstrates that anethole, purified from natural compound, is a potent inhibitor of growth of toxigenic V. cholerae. Our data suggest that anethole could be a potential antimicrobial drug candidate, particularly against MDR V. cholerae mediated infections.     

A Spontaneous Epithelial-Myoepithelial Carcinoma of the Submandibular Gland in a Sprague-Dawley Rat

The present report describes a rare case of spontaneous tumor of the salivary gland in a male Sprague-Dawley rat. The clinically confirmed mass rapidly developed in the cervical region between 19 and 21 weeks of age, and the animal was subsequently euthanized. At necropsy, a well-circumscribed nodule approximately 7 × 6 cm in diameter was found at the site of the salivary gland. The cut surface of the nodule was lobulated and soft and had a pinkish tan fish-flesh appearance. One large cyst (approximately 3 × 2 cm in size) containing reddish fluid was also present in the nodule. Histopathologically, the tumor, with a partially lobulated structure, was surrounded by a thin fibrous capsule. The majority of tumor cells formed a diffuse solid sheet structure that mainly consisted of small ovoid or spindle-shaped cells. In the tumor periphery, some cells were arranged in nest-like structures. Small duct-like structures lined with a monolayer of cuboidal epithelial cells resembling an intercalated duct or large polygonal clear cells with a myoepithelial component were also observed. Mitotic figures and necrotic foci were frequently observed in solid areas. Immunohistochemically, the tumor cells were positive for cytokeratin, epithelial membrane antigen, vimentin, p63, α-smooth muscle actin and calponin. The cells were negative for calcitonin, synaptophysin and chromogranin A. On the basis of these findings, the tumor was diagnosed as an epithelial-myoepithelial carcinoma originating from the luminal epithelial cells and myoepithelial cells in the submandibular gland.     

Swim bladder tumor in the young adult scoliotic medaka (Oryzias latipes)

A swim bladder tumor was detected in one scoliotic medaka aged 22 weeks. The tumor was located in the dorsal abdominal cavity, with maximum dimension of 1,850 × 1,500 µm. No swim bladder lumen was identified, and the region where the swim bladder lumen would have been located, was replaced with adipose tissues. The tumor was a non-invasive, expansile, and encapsulated solid mass with a few cysts, and comprised a homogenous population of well-differentiated, densely packed, gas glandular epithelium-like cells. The tumor mass was connected to a rete mirabile that showed a hyperplastic capillary plexus; however, the tumor cells did not invade the rete mirabile, thereby revealing that the tumor was an adenoma originating from the gas glandular epithelium of the swim bladder. Since proliferative lesions in the swim bladder have been reported in some teleosts with skeletal deformations, including medaka, the occurrence of a spontaneous swim bladder tumor in teleosts is considered to be closely associated with various types of skeletal deformation, and spinal curvature in particular.     

Detection of a novel herpesvirus associated with squamous cell carcinoma in a free-ranging Blanding’s turtle

The spread of both infectious and noninfectious diseases through wildlife populations is of increasing concern. Neoplastic diseases are rarely associated with population-level impacts in wildlife; however, impacts on individual health can be severe and might reflect deteriorating environmental conditions. An adult male free-ranging Blanding’s turtle (Emydoidea blandingii) originally captured in 2005 and deemed healthy, was recaptured in 2018 with a 1 × 1.5 cm intra-oral broad-based right mandibular mass. An excisional biopsy was performed, and histopathology revealed squamous cell carcinoma (SCC). Consensus herpesvirus PCR identified a novel herpesvirus (proposed name Emydoidea herpesvirus 2 [EBHV-2]) within the tumor. EBHV-2 shares 85% sequence homology with Terrapene herpesvirus 2 (TerHV-2), a herpesvirus linked to fibropapillomas in eastern box turtles (Terrapene carolina carolina). Virus-associated fibropapillomas have been identified in multiple marine turtle species and have had debilitating effects on their populations, but to date, virus-associated SCCs are rarely reported.     

Porphyromonas gulae 41-kDa fimbriae induced osteoclast differentiation and cytokine production

Porphyromonas gulae is considered to be associated with canine periodontitis. We have previously reported that the P. gulae American Type Culture Collection (ATCC) 51700 comprised 41-kDa fimbriae. The purpose of the present study was to demonstrate the roles of 41-kDa fimbrial protein in periodontal disease. In this study, we examined the involvement of the 41-kDa fimbrial protein in osteoclast differentiation and cytokine production in murine macrophages. Furthermore, alveolar bone resorption induced by P. gulae infection in rats was evaluated. To estimate osteoclast differentiation, bone marrow cells and MC3T3-G2/PA6 cells were cultured with or without the 41-kDa fimbrial protein for 7 days. BALB/c mouse peritoneal macrophages were stimulated with the 41-kDa fimbrial protein, and the levels of interleukin (IL)-1β and tumor necrosis factor (TNF)-α production were determined by enzyme-linked immunosorbent assay. Osteoclast differentiation was significantly enhanced by treatment with the 41-kDa fimbrial protein in a dose-dependent manner. The total area of pits formed on the dentine slices with osteoclasts incubated with the 41-kDa fimbrial protein was significantly greater than that of the control. The purified 41-kDa fimbrial protein induced IL-1β and TNF-α production in BALB/c mouse peritoneal macrophages after 6 hr of incubation in a dose-dependent manner. The bone loss level in rats infected with P. gulae was significantly higher than that of the sham-infected rats. These results suggest that P. gulae 41-kDa fimbriae play important roles in the pathogenesis of periodontal disease.     

Reproductive traits of the Ryukyu long-furred rat (Diplothrix legata) on Okinawa-jima Island

The Ryukyu long-furred rat, Diplothrix legata, is a large rodent distributed only on Amami-ohshima Island, Tokuno-shima Island and Okinawa-jima Island, Japan. This animal is endangered as a result of deforestation, predation by introduced carnivores and mortality caused by vehicles. We performed theriogenological examinations of 32 male and 25 female Ryukyu long-furred rats carcasses collected from wild populations on northern Okinawa-jima Island from December 2005 to September 2013. Adult males had remarkably large preputial glands. Seminiferous diameter of adult was significantly small (136 ± 28 µm, n=8) from April to August. Numerous spermatozoa were observed from September through February, and seminiferous diameter was significantly large (216 ± 27 µm, n=12) during this time in adults; testes length changed in a similar pattern. These findings indicate that the mating season may occur from September through February. Size (body length) at sexual maturity was estimated to be >560 mm in both sexes. From observation of corpora lutea and placental scars, litter size was estimated to range from 2 to 12 (average=6, n=4). These results provide fundamental knowledge that will be beneficial for in situ and ex situ conservation of this rare species.     

Dietary high protein-induced diarrhea and intestinal inflammation by activation of NF-κB signaling in piglets

The present study aimed to investigate whether inflammation-associated responses in piglets are induced by high protein (HP) through activating nuclear factor kappa B (NF-κB) signaling. Sixteen piglets (35 d of age, Duroc × [Landrace × Yorkshire], weaned at d 21, initial BW = 9.70 ± 0.11 kg) were allocated to 18% and 26% CP (HP group) at random, comprising 8 replicate pens per treatment. The piglets were slaughtered to collect intestinal tissues when apparent, persistent, and stable diarrhea syndromes happened (on d 12). No significant differences were observed in their growth performance (P > 0.05), but reduction by 19.11%, 25.31%, 23.64% of ADFI, ADG, and G:F, respectively was detected in the HP group. The HP group had greater (P = 0.002) diarrhea rates. Furthermore, dietary HP had lower ileal villus height (VH; P = 0.048), ratio of villus height to crypt depth (VH/CD ratio; P = 0.016), and colonic CD (P = 0.034), as well as had the trend (P = 0.075) to reduce the ileal villus absorptive area. Moreover, HP diets significantly elevated the goblet cell numbers in the ileal villi (P = 0.016) and colonic crypts (P < 0.001) and up-regulated (P = 0.012) the mRNA expression of mucin2 (Muc2) in the ileum. In addition, HP diets increased the myeloperoxidase concentration in the ileum (P = 0.002) and colon (P = 0.007) of piglets. Dietary HP significantly down-regulated the mRNA expression of tumor necrosis factor-α (TNF-α; P < 0.001) in the ileum, induced nitric oxide synthase (iNOS; P = 0.040) and interleukin-22 (IL-22; P = 0.008) in the colon, and inclined to down-regulate interleukin-1β (IL-1β; P = 0.076) expression in the colon. The relative protein abundance of Galectin-3 (P = 0.046) in the colon and the ratio of phosphorylation NF-κB to NF-κB (p–NF–κB/NF-κB ratio) in the ileum of HP piglets were also greater (P = 0.038). These results suggest that dietary HP may cause diarrhea in piglets by activating NF-κB signaling induced intestinal inflammation.     

Determination phase at transition of gonocytes to spermatogonial stem cells improves establishment efficiency of spermatogonial stem cells in domestic cats

The development of germ cells has not been entirely documented in the cat especially the transition phase of the gonocyte to the spermatogonial stem cell (G/SSC). The aims of study were to examine testicular development and to identify the G/SSC transition in order to isolate and culture SSCs in vitro. Testes were divided into 3 groups according to donor age (I, < 4 months; II, 4–6 months; and III, > 6 months). In Exp. 1, we studied testicular development by histology, transmission electron microscopy and immunohistochemistry. In Exp. 2, we determined the expression of GFRα-1, DDX-4 and c-kit and performed flow cytometry. The SSCs isolated from groups II and III were characterized by RT-PCR and TEM (Exp. 3). Chronological changes in the G/SSC transition were demonstrated. The size, morphology and ultrastructure of SSCs were distinguishable from those of gonocytes. The results demonstrated that group II contained the highest numbers of SSCs per seminiferous cord/tubule (17.66 ± 2.20%) and GFRα-1⁺ cells (14.89 ± 5.66%) compared with the other groups. The findings coincided with an increased efficiency of SSC derivation in group II compared with group III (74.33 ± 2.64% vs. 23.33 ± 2.23%). The colonies expressed mRNA for GFRA1, ZBTB16, RET and POU5F1. Our study found that the G/SSC transition occurs at 4–6 months of age. This period is useful for isolation and improves the establishment efficiency of cat SSCs in vitro.     

Effect of Saccharomyces cerevisiae on alfalfa nutrient degradation characteristics and rumen microbial populations of steers fed diets with different concentrate-to-forage ratios

Live yeast (Saccharomyces cerevisiae) constitutes an effective additive for animal production; its probiotic effect may be related to the concentrate-to-forage ratio (CTFR). The objective of this study was to assess the effects of S. cerevisiae (SC) on fiber degradation and rumen microbial populations in steers fed diets with different levels of dietary concentrate. Ten Simmental × Local crossbred steers (450 ± 50 kg BW) were assigned to a control group or an SC group. Both groups were fed the same basal diet but the SC group received SC supplementation (8 × 10⁹ cfu/h/d through the ruminal fistula) following a two-period crossover design. Each period consisted of four phases, each of which lasted 17 d: 10 d for dietary adaptation, 6 d for degradation study, and 1 d for rumen sample collection. From the 1^st to the 4^th phase, steers were fed in a stepwise fashion with increasing CTFRs, i.e., 30:70, 50:50, 70:30, and 90:10. The kinetics of dry matter and fiber degradation of alfalfa pellets were evaluated; the rumen microbial populations were detected using real-time PCR. The results revealed no significant (P > 0.05) interactions between dietary CTFR and SC for most parameters. Dietary CTFR had a significant effect (P < 0.01) on degradation characteristics of alfalfa pellets and the copies of rumen microorganism; the increasing concentrate level resulted in linear, quadratic or cubic variation trend for these parameters. SC supplementation significantly (P < 0.05) affected dry matter (DM) and neutral detergent fiber (NDF) degradation rates (c_DM, c_NDF) and NDF effective degradability (ED_NDF). Compared with the control group, there was an increasing trend of rumen fungi and protozoa in SC group (P < 0.1); copies of total bacteria in SC group were significantly higher (P < 0.05). Additionally, percentage of Ruminobacter amylophilus was significantly lower (P < 0.05) but percentage of Selenomonas ruminantium was significantly higher (P < 0.05) in the SC group. In a word, dietary CTFR had a significant effect on degradation characteristics of forage and rumen microbial population. S. cerevisiae had positive effects on DM and NDF degradation rate or effective degradability of forage; S. cerevisiae increased rumen total bacteria, fungi, protozoa, and lactate-utilizing bacteria but reduced starch-degrading and lactate-producing bacteria.