Effect of ethanol extract from Chrysanthemum cinerariifolium leaves on Ki-67 proliferation and dysplasia severity in a rat model of oral squamous cell carcinoma

Background:Oral squamous cell carcinoma (OSCC) is a malignant tumor that can rapidly infiltrate the oral epithelial tissue and cause high mortality worldwide because the available therapies are less effective. Chrysanthemum cinerariifolium leaf contains secondary metabolites as anti-inflammatory, antioxidant, anticancer, and antimutagenic. Aims:The study aimed to analyze the ethanolic extract of C. cinerariifolium leaf in reducing proliferation (Ki-67) and the degree of dysplasia in OSCC rats. Methods:This study used male Sprague Dawley induced by 7,12-dimethylbenz(a)anthracene (DMBA) 0.5% and divided into five treatment groups, namely positive control/C+ (sick), negative control/C- (healthy), and treatment group induced with DMBA and given extract C. cinerariifolium leaf with successive doses of T1, T2, and T3 (50, 100, and 200 mg/kg bw). The oral epithelium was stained with hematoxylin and eosin and immunohistochemically stained with a Ki-67 monoclonal antibody. The statistical analysis utilizes the one-way analysis of variance test. Results:The results showed that T1 at a dose of 200 mg/kg bw could significantly reduce Ki-67 expression and the degree of oral epithelial dysplasia (OED; p < 0.05) close to healthy controls. Conclusion:The conclusion shows that C. cinerariifolium leaf extract can be a therapy against OSCC by decreasing cell proliferation and the degree of OED.     

A 13-week subchronic toxicity study of 2-(l-menthoxy)ethanol in F344 rats

2-(l-Menthoxy)ethanol has been frequently employed as a flavoring agent; however, data regarding 2-(l-menthoxy)ethanol toxicity remain limited. We performed a 13-week subchronic toxicity study of 2-(l-menthoxy)ethanol in male and female F344 rats, with doses of 0, 15, 60, or 250 mg/kg body weight (BW)/day orally administered by gavage using corn oil as the vehicle. No significant toxicological changes in general condition, body weight, or food intake were observed in any groups. The hematological assessment showed decreased hemoglobin, hematocrit, mean corpuscular volume, and mean corpuscular hemoglobin and increased platelet count in the male 250 mg/kg group. Serum biochemistry revealed elevated total cholesterol in the 250 mg/kg group of male and female rats, reduced triglyceride in the female 250 mg/kg group, and increased total protein in the male 250 mg/kg group, indicating effects on lipid metabolism and protein synthesis. For organ weights, absolute and relative weights of the liver and adrenal glands were increased in the 250 mg/kg group of both sexes and the male 250 mg/kg group, respectively. Histopathological analysis showed chronic nephropathy in the male 15 mg/kg or higher groups, with increased absolute and relative kidney weights, as well as elevated serum creatinine, in the male 60 and 250 mg/kg groups. However, eosinophilic granules containing α_2u-globulin were identified in proximal tubules, suggesting α_2u-globulin nephropathy specific to male rats and without toxicological significance. These results indicated that the no-observed-adverse-effect level of 2-(l-menthoxy)ethanol was 60 mg/kg BW/day for both sexes.     

Hyperferritinemia is associated with short survival time in dogs with multicentric lymphoma

In the present study, we examined the relationship between serum ferritin concentration before treatment and survival time in dogs with multicentric lymphoma. Eighteen dogs with multicentric lymphoma were enrolled in the study. When the dogs were classified into high and low ferritin groups on the basis of their serum ferritin concentration (3,000 ng/ml cut-off value), the median survival time of dogs with high concentrations (≥3,000 ng/ml, n=7) was 40 days, whereas it was 360 days among dogs with low concentrations (<3,000 ng/ml, n=11). This difference was statistically significant (P=0.001). This finding suggests that the initial high level of serum ferritin indicates short survival time in dogs with multicentric lymphoma. Large-scale research is necessary to confirm this finding.     

Immunohistochemical expression of HER - 2 and Ki - 67 in granulosa cell tumor in bitches

Granulosa cell tumour, an ovarian neoplasm of stromal origin, is an important tumour related to oestrogenic dominance syndrome and cystic endometrial hyperplasia–pyometra complex. In order to analyse ovarian tumour´s malignant potential, immunohistochemical markers can be used, such as anti-HER2 and anti-Ki-67. The aim of this study was to evaluate the expression of immunohistochemical markers HER-2 and Ki-67 in granulosa cell tumour from bitches´ ovaries. In HER-2 immunomarker analysis using the HercepTest^® method, most tumours were classified as 2+ (moderate labelling). Concerning Ki-67 immunomarker, only one case was described as having a high proliferative index. An association was found between immunostained cell percentage by anti-HER-2 antibodies and high pleomorphism, represented by the pattern of follicular/trabecular tumour arrangement. There was no correlation between anti-Ki-67 and anti-HER-2 antibody immunostaining intensities, probably due to only one case with a high Ki-67 index. With an effective protocol for HER-2 and Ki-67 immunohistochemical identification in granulosa cell tumours in bitches, it was possible to characterize this neoplasm proliferation profile.     

Trace amounts of antibiotic exacerbated diarrhea and systemic inflammation of weaned pigs infected with a pathogenic Escherichia coli

The experiment was conducted to investigate the effects of trace amounts of antibiotic on growth performance, diarrhea, systemic immunity, and intestinal health of weaned pigs experimentally infected with an enterotoxigenic Escherichia coli. Weaned pigs (n = 34, 6.88 ± 1.03 kg body weight [BW]) were individually housed in disease containment rooms and randomly allotted to one of the three dietary treatments: nursery basal diet (CON) and two additional diets supplemented with 0.5 or 50 mg/kg carbadox to the nursery basal diet (TRA or REC), respectively. The experiment lasted 18 d with 7 d before and 11 d after the first E. coli inoculation. The E. coli F18 inoculum was orally provided to all pigs with a dose of 10¹⁰ colony-forming unit (CFU)/3 mL for three consecutive days. Fecal and blood samples were collected on day 0 before inoculation and days 2, 5, 8, and 11 postinoculation (PI) to test the percentage of β-hemolytic coliforms in total coliforms and complete blood cell count, respectively. Sixteen pigs were euthanized on day 5 PI, whereas the remaining pigs were euthanized at the end of the experiment to collect the jejunal and ileal mucosa and mesenteric lymph node for gene expression and bacterial translocation, respectively. Pigs in REC had greater (P < 0.05) final BW and lower (P < 0.05) overall frequency of diarrhea compared with pigs in the CON and TRA groups. Pigs in TRA had the lowest (P < 0.05) average daily gain and feed efficiency from day 0 to 5 PI, highest (P < 0.05) percentage of β-hemolytic coliforms in fecal samples on days 2 and 5 PI, and greatest (P < 0.05) bacterial colonies in mesenteric lymph nodes on day 11 PI compared with pigs in the CON and REC groups. Pigs in TRA had the greatest (P < 0.05) neutrophils on day 5 PI and higher (P < 0.05) white blood cell counts and lymphocytes than other groups on day 11 PI. Pigs in TRA had the greatest (P < 0.05) serum C-reactive protein on days 2 and 5 PI and serum tumor necrosis factor-α on day 5 PI, compared with pigs in the CON and REC groups. Pigs fed REC had increased (P < 0.05) mRNA expression of zona occludens-1 (ZO-1) and occludin (OCDN) and reduced (P < 0.05) interleukin-1 beta (IL1B), interleukin-6 (IL6), and tumor necrosis factor-alpha (TNFA) in ileal mucosa on day 5 PI, compared with the CON, whereas TRA upregulated (P < 0.05) mRNA expression of IL1B, IL6, and cyclooxygenase-2 (COX2) in the ileal mucosa on day 11 PI, compared with the REC. In conclusion, trace amounts of antibiotic may exacerbate the detrimental effects of E. coli infection on pig performance by increasing diarrhea and systemic inflammation of weanling pigs.     

Lack of hepatocarcinogenicity of 2,2’-[1,2-ethanediylbis(oxymethylene)]bis-oxirane, 3-hydroxy-2-naphthoic acid,and acetoacetanilide in a medium-term rat liver bioassay

The carcinogenicity of 2,2’-[1,2-ethanediylbis(oxymethylene)]bis-oxirane (ethylene glycol diglycidyl ether; EGDE), 3-hydroxy-2-naphthoic acid (HNA), and acetoacetanilide (AAA) was investigated using a medium-term rat liver bioassay for an occupational safety assessment. F344 male rats were administered a single intraperitoneal injection of diethylnitrosamine (200 mg/kg body weight (bw)/day) and then starting 2 weeks later, they received EGDE at 6, 20, and 60 mg/kg bw/day, HNA at 20, 60, and 200 mg/kg bw/day, or AAA at 60, 200, and 600 mg/kg bw/day by oral gavage for 6 weeks. The animals in the positive control group received phenobarbital sodium solution (PB, 25 mg/kg bw/day) by oral gavage and those in the negative control group received a vehicle (water/corn oil) during the administration period of test substances in this model. All animals were subjected to two-thirds partial hepatectomy at week 3 and euthanized at week 8. Neither the number nor the area of hepatocellular foci positive for glutathione S-transferase placental form (GST-P) increased in any of the EGDE, HNA, or AAA treated groups. However, the number and area of GST-P-positive foci significantly increased in the positive control group treated with PB. The results indicate that EGDE, HNA, and AAA lack hepatocarcinogenicity in rats.     

Modifications of azoxymethane-induced carcinogenesis and 90-day oral toxicities of 2-tetradecylcyclobutanone as a radiolytic product of stearic acid in F344 rats

A 90-day oral toxicity test in rats was performed to evaluate the toxicity of 2-tetradecylcyclobutanone (2-tDCB), a unique radiolytic product of stearic acid. Six-week-old male and female F344 rats (n=15/group) were given 2-tDCB at concentrations of 0, 12, 60 and 300 ppm in a powder diet for 13 weeks. Slight dose-dependent increases in serum total protein and albumin in male rats were found, but these changes were not considered to be a toxic effect. The fasting, but not non-fasting, blood glucose levels of the male rats in the 300 ppm group and female rats in the 60 and 300 ppm groups were lower than those of the controls. Gas chromatography-mass spectrometry analysis showed dose-dependent accumulation of 2-tDCB in adipose tissue, notably in males. Next, we performed an azoxymethane (AOM)-induced two-stage carcinogenesis study. After injection of 6-week-old male F344 rats (n=30/group) once a week for 3 weeks, the animals received 2-tDCB at concentrations of 0, 10, 50 and 250 ppm in a powder diet for 25 weeks. The incidences of colon tumors for the 2-tDCB dosages were 34%, 45%, 40% and 37%, respectively, and were not statistically significant. These data suggest that 2-tDCB shows no toxic or tumor-modifying effects under the present conditions, and that the no-observed-adverse-effect level for 2-tDCB is 300 ppm in both sexes, equivalent to 15.5 mg/kg b.w./day in males and 16.5 mg/kg b.w./day in females.     

Dietary carbohydrate modifies the density of L cells in the chicken ileum

Glucagon-like peptides (GLPs) are secreted from intestinal L cells and stimulate various physiological functions in the gastrointestinal tract. The secretion of GLPs is influenced by macronutrient ingestion. This study aims to clarify the effects of dietary carbohydrate (CHO) on L cells in the chicken ileum. Six-week-old, male White Leghorn chickens were divided into three groups: control, low-CHO and CHO-free, with five chickens in each group. Paraffin sections were made from the proximal and distal ileum of each animal and subjected to immunohistochemistry for GLP-1 and GLP-2 peptides and in situ hybridization for proglucagon (PG) mRNA. A significant reduction of GLP-1- and GLP-2-immunoreactive cells was observed in the two experimental groups compared with that in the control. A reduction of cells expressing PG mRNA was observed in the proximal and distal ileum of the CHO-free group compared with that in the control. The ratio of GLP-1-immunoreactive cells showing Ki-67 immunoreactivity was significantly lower in the distal ileum of the CHO-free group than that in the control group. These data suggest that dietary CHO is an effective stimulator for modifying L cell density in the chicken ileum.     

LC-MS/MS measurement of ampicillin residue in swine tissues at 5 days after in-feed administration

We assessed ampicillin (ABPC) concentrations of kidney, muscle and intestine after a 5–day withdrawal period in 2 male and a female young Large White pigs fed the diet containing ABPC (ABPC medicated feed, 24 mg/kg/day) for a week. The ABPC residues were measured with liquid chromatography–tandem mass spectrometry, and the mean recoveries and quantitation limits ranged from 91.8 to 97.2% and from 0.1 to 0.12 ng/g, respectively. The residual ABPC concentrations were ≤1.18 ng/g for the muscle, ≤0.53 ng/g for the kidney and ≤1.93 ng/g for the intestine, suggesting below the Japanese provisional maximum residue limits. These results reveal that the analytical method is developed for residual ABPC and that the withdrawal period is appropriate.     

The effects of supplementing <Emphasis Type="Italic">Acacia mearnsii</Emphasis> tannin extract on dairy cow dry matter intake,milk production,and methane emission in a tropical pasture

The study assessed the effect of Acacia mearnsii tannin extract supplementation grazing dairy cows on dry matter (DM) intake, enteric methane (CH₄) emission, and performance. Twelve Holstein cows were divided into two groups and subjected to two treatments that consisted of millet pasture (Pennisetum glaucum L.) plus supplementation with 6 kg of concentrate (750-g/kg ground corn and 250-g/kg soybean meal) including or excluding 120-g tannin extract. The trial design was a double reversal using three periods of 28 days each, with 21 days for the adaption period, and 7 days for sample collection. Herbage intake was measured using the n-alkane technique, and daily CH₄ emission was measured with the sulfur hexafluoride tracer gas technique. Individual total DM intake (mean = 17.1 kg/day), herbage DM intake (mean = 11.8 kg/day), and milk production (mean = 19.2 kg/day) were similar between treatments. CH₄ emission significantly decreased (32%, P < 0.05) in the animals supplemented with tannin extract, compared to non-supplemented animals. On the other hand, as proportion of DM intake or milk production, methane emission tended to decrease in tannin-supplemented animals. Supplementing dairy cows grazing a millet pasture with 120-g tannin extract reduced daily CH₄ emission without affecting animal performance.     

RNAi-mediated knockdown of INHBB increases apoptosis and inhibits steroidogenesis in mouse granulosa cells

Inhibins are members of the TGFβ superfamily and act as suppressors of follicle stimulating hormone (FSH) secretion from pituitary glands via a negative feedback mechanism to regulate folliculogenesis. In this study, the INHBB gene was knocked down by three RNAi-Ready pSIREN-RetroQ-ZsGreen vector- mediated recombinant plasmids to explore the effects of INHBB silencing on granulosa cell (GC) cell cycle, apoptosis and steroid production in vitro. Quantitative real-time polymerase chain reaction, Western blot, flow cytometry and ELISA were performed to evaluate the role of INHBB in the mouse GC cell cycle, apoptosis and steroid production in vitro. The results showed that the relative mRNA and protein expression of INHBB in mouse GCs can be significantly reduced by RNAi with pshRNA-B1, pshRNA-B2 and pshRNA-B3 plasmids, with pshRNA-B3 having the best knockdown efficiency. Downregulation of the expression of INHBB significantly arrests cells in the G1 phase of the cell cycle and increases the apoptosis rate in GCs. This was further confirmed by downregulation of the protein expressions of Cyclin D1, Cyclin E and Bcl2, while the protein expression of Bax was upregulated. In addition, specific downregulation of INHBB markedly decreased the concentration of estradiol and progesterone, which was further validated by the decrease in the mRNA levels of CYP19A1and CYP11A1. These findings suggest that inhibin βB is important in the regulation of apoptosis and cell cycle progression in granulosa cells. Furthermore, the inhibin βB subunit has a role in the regulation of steroid hormone biosynthesis. Evidence is accumulating to support the concept that inhibin βB is physiologically essential for early folliculogenesis in the mouse.     

A molecular epidemiological survey of Babesia, Hepatozoon,Ehrlichia and Anaplasma infections of dogs in Japan

Tick-borne diseases are often encountered in canine clinical practice. In the present study, a molecular epidemiological survey of dogs in Japan was conducted to understand the prevalence and geographical distribution of Babesia spp., Hepatozoon spp., Ehrlichia spp. and Anaplasma spp. Pathogen-derived DNA in blood samples obtained from 722 dogs with a history of exposure to ticks and/or fleas was examined by PCR. The prevalence of Babesia gibsoni, Babesia odocoilei-like species, Hepatozoon canis and Ehrlichia spp./Anaplasma spp. was 2.4% (16/722), 0.1% (1/722), 2.5% (18/722) and 1.5% (11/722), respectively. While B. gibsoni and Ehrlichia spp./Anaplasma spp. were detected in the western part of Japan, H. canis was detected in Tohoku area in addition to western and central parts of Japan.     

RNAi-mediated Knockdown of Xist Does Not Rescue the Impaired Development of Female Cloned Mouse Embryos

In mice, one of the major epigenetic errors associated with somatic cell nuclear transfer (SCNT) is ectopic expression of Xist during the preimplantation period in both sexes. We found that this aberrant Xist expression could be impeded by deletion of Xist from the putative active X chromosome in donor cells. In male clones, it was also found that prior injection of Xist-specific siRNA could significantly improve the postimplantation development of cloned embryos as a result of a significant repression of Xist at the morula stage. In this study, we examined whether the same knockdown strategy could work as well in female SCNT-derived embryos. Embryos were reconstructed with cumulus cell nuclei and injected with Xist-specific siRNA at 6–7 h after oocyte activation. RNA FISH analysis revealed that siRNA treatment successfully repressed Xist RNA at the morula stage, as shown by the significant decrease in the number of cloud-type Xist signals in the blastomere nuclei. However, blastomeres with different sizes (from “pinpoint” to “cloud”) and numbers of Xist RNA signals remained within single embryos. After implantation, the dysregulated Xist expression was normalized autonomously, as in male clones, to a state of monoallelic expression in both embryonic and extraembryonic tissues. However, at term there was no significant improvement in the survival of the siRNA-injected cloned embryos. Thus, siRNA injection was largely effective in repressing the Xist overexpression in female cloned embryos but failed to rescue them, probably because of an inability to mimic consistent monoallelic Xist expression in these embryos. This could only be achieved in female embryos by applying a gene knockout strategy rather than an siRNA approach.     

Tarantula cubensis extract alters the degree of apoptosis and mitosis in canine mammary adenocarcinomas

In the present study, 13 clinical cases of canine mammary adenocarcinoma were evaluated in order to understand the effect of Tarantula cubensis extract (TCE) on tumor tissue. Punch biopsies were taken from the tumors before treatment with TCE. Subcutaneous injections of TCE were administered three times at weekly intervals (3 mL per dog). Between days 7 and 10 after the third injection, the tumor masses were extirpated by complete unilateral mastectomy. Pre- and post-treatment tumor tissues were immunohistochemically assessed. The expression of B-cell lymphoma 2 (Bcl-2) was found to be higher in pre-treatment compared to post-treatment tissues (p < 0.01) whereas Ki-67 expression was lower in post-treatment tissues (p < 0.01). No significant differences in fibroblast growth factor or vascular endothelial growth factor expression were observed between pre- and post-treatment tissues (p > 0.05). The apoptotic index was determined to be low before treatment and increased during treatment. These results suggest that TCE may be effective for controlling the local growth of canine mammary adenocarcinoma by regulating apoptosis.     

Diazepam ameliorated myocardial ischemia-reperfusion injury via inhibition of C-C chemokine receptor type 2/tumor necrosis factor-alpha/interleukins and Bcl-2-associated X protein/caspase-3 pathways in experimental rats

Myocardial ischemia-reperfusion injury (IRI) is one of the most leading concerns for public health globally. Diazepam, a local anesthetic, has been reported for its cardioprotective potential. The present investigation aimed to evaluate the possible mechanism of action of diazepam against left anterior descending ligation-induced myocardial IRI in experimental rats. IRI was induced in healthy male rats by ligating coronary artery for 30 min and then reperfused for 60 min. The animals were pre-treated with either vehicle or diltiazem (10 mg/kg) or diazepam (1, 2.5, and 5 mg/kg) for 14 days. Compared to the IRI group, diazepam (2.5 and 5 mg/kg) markedly (P<0.05) attenuated IRI-induced alterations in cardiac function and oxido-nitrosative stress. In addition, diazepam prominently (P<0.05) improved cardiac Na⁺K⁺ATPase, Ca²⁺ATPase levels and hypoxia-inducible factor-1 alpha (HIF-1α) mRNA expression. It also significantly (P<0.05) down-regulated cardiac mRNA expressions of cardiac troponin I (cTn-I), C-C chemokine receptor type 2 (CCR2), tumor necrosis factor-alpha (TNF-α), interleukins (IL)-1β, and IL-6. In western blot analysis, IRI-induced myocardial apoptosis was reduced by diazepam treatment reflected by a marked (P<0.05) decreased in Bcl-2-associated X protein (Bax) and Caspase-3 protein expression. Diazepam also efficiently (P<0.05) improved IRI-induced histological aberration in cardiac tissue. In conclusion, diazepam exerts cardioprotective effect by inhibiting inflammatory release (CCR2, TNF-α, and ILs), oxido-nitrosative stress, and apoptosis (Bax and Caspase-3) pathway during myocardial IRI in experimental rats.     

Somatic cell and innate immune responses in mammary glands of lactating cows to intramammary infusion of Bifidobacterium breve at pre-drying off period

Intramammary infusion of Bifidobacterium breve (B. breve)-induced somatic cell (SC) counts, chemiluminescent response (CL), lactoferrin (LF) concentrations and mastitis-causing pathogens from quarters with subclinical mastitis were measured to evaluate innate immune response of mammary glands in dairy cows at 3 to 4 weeks before drying off. SC counts in 7 quarters of 7 control cows and 5 quarters of 6 cows with mastitis increased markedly on day 1 and SC values in control cows were significantly (P<0.05) increased and returned to pre-infusion levels on day 5 after B. breve-infusion. CL values in both groups increased markedly on day 1 and then decreased after B. breve-infusion; however, CL values in cows with mastitis did not return to normal levels on day 5 and at postpartum. The CL values were highly correlated with their SC counts in milk from both groups. LF concentrations increased toward day 3 after B. breve-infusion and were higher in cows with mastitis. B. breve-infusion eliminated 16.6% (1/6) of pathogens from 6 quarters with chronic subclinical mastitis. B. breve-induced SC responses in quarters from 3 cows with mastitis showed characteristic patterns of recovery, persistent and new infections. B. breve-induced SC counts in quarters from the cows in the pre-drying off were lower (25.7–70.6%) than those of the cows in mid-lactation. The intrinsic innate immune response in cows on pre-drying off may be decreased and appears to be insufficient to eliminate pathogens from mammary gland in the pre-drying off.     

Protective effects of silymarin on fumonisin B1-induced hepatotoxicity in mice

The present study was conducted to investigate the effect of silymarin on experimental liver toxication induced by Fumonisin B₁ (FB₁) in BALB/c mice. The mice were divided into six groups (n = 15). Group 1 served as the control. Group 2 was the silymarin control (100 mg/kg by gavage). Groups 3 and 4 were treated with FB₁ (Group 3, 1.5 mg/kg FB₁, intraperitoneally; and Group 4, 4.5 mg/kg FB₁). Group 5 received FB₁ (1.5 mg/kg) and silymarin (100 mg/kg), and Group 6 was given a higher dose of FB₁ (4.5 mg/kg FB₁) with silymarin (100 mg/kg). Silymarin treatment significantly decreased (p < 0.0001) the apoptotic rate. FB₁ administration significantly increased (p < 0.0001) proliferating cell nuclear antigen and Ki-67 expression. Furthermore, FB₁ elevated the levels of caspase-8 and tumor necrosis factor-alpha mediators while silymarin significantly reduced (p < 0.0001) the expression of these factors. Vascular endothelial growth factor (VEGF) and fibroblast growth factor-2 (FGF-2) expressions were significantly elevated in Group 4 (p < 0.0001). Silymarin administration alleviated increased VEGF and FGF-2 expression levels (p < 0.0001). In conclusion, silymarin ameliorated toxic liver damage caused by FB₁ in BALB/c mice.     

Efficacy of levamisole alone and in combination with mebendazole against Gongylonema pulchrum infection in rabbits

Gongylonema pulchrum is an important parasite of captive primates. Twelve rabbits were infected with 30 third-stage larvae of G. pulchrum. At 4–7 months post-infection, animals were administered levamisole at a single dose of 12 mg/kg, levamisole at 8 mg/kg three times at 2-day intervals, levamisole at a single dose of 8 mg/kg after administration of mebendazole at 70 mg/kg for 3 days or 8 ml of distilled water for 3 days (control). Necropsy at 14 days after treatment revealed that single and multiple dosages of levamisole reduced nematode burdens by 68.4% and 89.5%, respectively. The combined regimen of mebendazole and levamisole exhibited high efficacy for treating G. pulchrum located widely within the upper digestive tract, with a reduction of 98.2%. These results suggest that this combined chemotherapy treatment may be effective against G. pulchrum infection, including buccal and lingual gongylonemiasis in primates.