Asian Pacific Journal of Tropical Biomedicine

Issue 7,2014 Table of Contents

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  • 1  Antimicrobial activity against periodontopathogenic bacteria, antioxidant and cytotoxic effects of various extracts from endemic Thermopsis turcica
    Elif Burcu Bali Leyla A??k Gül?in Akca Meral Sarper Mualla P?nar El?i Ferit Avcu Mecit Vural
    2014(7):505-514. DOI: 10.12980/APJTB.4.2014APJTB-2013-0010
    [Abstract](19) [HTML](0) [PDF 814.59 K](80)
    Abstract:
    Objective: To investigate the in vitro antimicrobial potential of Thermopsis turcica Kit Tan, Vural & Küçüködük against periodontopathogenic bacteria, its antioxidant activity and cytotoxic effect on various cancer cell lines. Methods: In vitro antimicrobial activities of ethanol, methanol, ethyl acetate (EtAc), n-hexane and water extracts of Thermopsis turcica herb against periodontopathogenic bacteria, Aggregatibacter actinomycetemcomitans ATCC 29523 and Porphyromonas gingivalis ATCC 33277 were tested by agar well diffusion, minimum inhibitory concentration (MIC) and minimal bactericidal concentration (MBC). Antioxidant properties of the extracts were evaluated by 1,1-diphenyl-2-picryl-hydrazyl radical scavenging activity and β-carotene bleaching methods. Amounts of phenolic contents of the extracts were also analysed by using the Folin-Ciocalteu reagent. Additionally, cytotoxic activity of the extracts on androgen-insensitive prostate cancer, androgen-sensitive prostate cancer, chronic myelogenous leukemia and acute promyelocytic leukemia human cancer cell lines were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Human gingival fibroblast cells were used as a control. Results: Our data showed that EtAc extract had the highest antimicrobial effect on Aggregatibacter actinomycetemcomitans (MIC: 1.562 mg/mL, MBC: 3.124 mg/mL) and Porphyromonas gingivalis (MIC: 0.781 mg/ mL, MBC: 1.562 mg/mL). In antioxidant assays, EtAc extract exhibited also the highest radical scavenging activity [IC50=(30.0±0.3) μg/mL] and the highest inhibition [(74.35±0.30)%] against lineloic acide oxidation. The amount of phenolic content of it was also the highest [(162.5±1.2) μg/mg gallic acid]. In cytotoxic assay, only ethanol [IC50=(80.00±1.21) μg/mL] and EtAc extract [IC50=(70.0±0.9) μg/mL] were toxic on acute promyelocytic leukemia cells at 20-100 μg/mL (P<0.05). However, no toxic effect was observed on human gingival fibroblast cells. Conclusions: According to our findings, owing to its antioxidant and cytotoxic potential, EtAc extract might include anticancer agents for acute promyelocytic leukemia.
    2  Proteomics analysis of antimalarial targets of Garcinia mangostana Linn.
    Wanna Chaijaroenkul Artitiya Thiengsusuk Kanchana Rungsihirunrat Stephen Andrew Ward Kesara Na-Bangchang
    2014(7):515-519. DOI: 10.12980/APJTB.4.2014APJTB-2014-0043
    [Abstract](24) [HTML](0) [PDF 478.75 K](77)
    Abstract:
    Objective: To investigate possible protein targets for antimalarial activity of Garcinia mangostana Linn. (G. mangostana) (pericarp) in 3D7 Plasmodium falciparum clone using 2-dimensional electrophoresis and liquid chromatography mass-spectrometry (LC/MS/MS). Methods: 3D7 Plasmodium falciparum was exposed to the crude ethanolic extract of G. mangostana Linn. (pericarp) at the concentrations of 12µg/mL (IC50 level: concentration that inhibits parasite growth by 50%) and 30 µg/mL (IC90 level: concentration that inhibits parasite growth by 90%) for 12 h. Parasite proteins were separated by 2-dimensional electrophoresis and identified by LC/MS/MS. Results: At the IC50 concentration, about 82% of the expressed parasite proteins were matched with the control (non-exposed), while at the IC90 concentration, only 15% matched proteins were found. The selected protein spots from parasite exposed to the plant extract at the concentration of 12 µg/mL were identified as enzymes that play role in glycolysis pathway, i.e., phosphoglycerate mutase putative, L-lactate dehydrogenase/glyceraldehyde-3-phosphate dehydrogenase, and fructose-bisphosphate aldolase/phosphoglycerate kinase. The proteosome was found in parasite exposed to 30 µg/mL of the extract. Conclusions: Results suggest that proteins involved in the glycolysis pathway may be the targets for antimalarial activity of G. mangostana Linn. (pericarp).
    3  The presence of eucalyptol in Artemisia australis validates its use in traditional Hawaiian medicine
    David Zant Daniel A. Gubler
    2014(7):520-522. DOI: 10.12980/APJTB.4.2014C1112
    [Abstract](29) [HTML](0) [PDF 279.53 K](86)
    Abstract:
    Objective: To identify the major organic compounds of Artemisia australis (A. australis), a plant used in traditional Hawaiian medicine for the treatment of asthma. Methods: The dichloromethane extract of A. australis was analyzed by gas chromatography-mass spectroscopy and major compounds were identified by a National Institute of Standards and Technology library search and confirmed by peak enhancement. Results: echnology library search and confirmed by peak enhancement. The major chemical components of A. australis include eucalyptol, borneol, and caryophyllene. Conclusions: The presence and biological activity of eucalyptol correlate very well with the usage of this plant in traditional Hawaiian medicine.
    4  Effects of lactic acid bacteria isolated from fermented mustard on lowering cholesterol
    Shu Chen Wang Chen Kai Chang Shu Chang Chan Jiunn Shiuh Shieh Chih Kwang Chiu Pin-Der Duh
    2014(7):523-528. DOI: 10.12980/APJTB.4.201414B54
    [Abstract](24) [HTML](0) [PDF 259.47 K](80)
    Abstract:
    Objective: To evaluate the ability of lactic acid bacteria (LAB) strains isolated from fermented mustard to lower the cholesterol in vitro. Methods: The ability of 50 LAB strains isolated from fermented mustard on lowering cholesterol in vitro was determined by modified o-phtshalaldehyde method. The LAB isolates were analyzed for their resistance to acid and bile salt. Strains with lowering cholesterol activity, were determined adherence to Caco-2 cells. Results: Strain B0007, B0006 and B0022 assimilated more cholesterol than BCRC10474 and BCRC 17010. The isolated strains showed tolerance to pH 3.0 for 3 h despite variations in the degree of viability and bile-tolerant strains, with more than 108 CFU/mL after incubation for 24 h at 1% oxigall in MRS. In addition, strain B0007 and B0022 identified as Lactobacillus plantarum with 16S rDNA sequences were able to adhere to the Caco-2 cell lines. Conclusions: These strains B0007 and B0022 may be potential functional sources for cholesterol-lowering activities as well as adhering to Caco-2 cell lines.
    5  Jeju seaweeds suppress lipopolysaccharide-stimulated proinflammatory response in RAW 264.7 murine macrophages
    Eun-Jin Yang Ji-Young Moon Sang Suk Kim Kyong-Wol Yang Wook Jae Lee Nam Ho Lee Chang-Gu Hyun
    2014(7):529-537. DOI: 10.12980/APJTB.4.2014C1099
    [Abstract](25) [HTML](0) [PDF 1.35 M](81)
    Abstract:
    Objective: To investigate the anti-inflammatory effects of Jeju seaweeds on macrophage RAW 264.7 cells under lipopolysaccharide (LPS) stimulation. Methods: Ethyl acetate fractions were prepared from five different types of Jeju seaweeds, Dictyopteris divaricata (D. divaricata), Dictyopteris prolifera (D. prolifera), Prionitis cornea (P. cornea), Grateloupia lanceolata (G. lanceolata), and Grateloupia filicina (G. filicina). They were screened for inhibitory effects on proinflammatory mediators and cytokines such as nitric oxide (NO), prostaglandin E2, tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6). Results: Our results revealed that D. divaricata, D. prolifera, P. cornea, G. lanceolata, and G. filicina potently inhibited LPS-stimulated NO production (IC50 values were 18.0, 38.36, 38.43, 32.81 and 37.14 µg/mL, respectively). Consistent with these findings, D. divaricata, D. prolifera, P. cornea, and G. filicina also reduced the LPS-induced and prostaglandin E2 production in a concentration-dependent manner. Expectedly, they suppressed the expression of inducible NO synthase and cyclooxygenase-2 at the protein level in a dose-dependent manner in the RAW 264.7 cells, as determined by western blotting. In addition, the levels of TNF-α and IL-6, released into the medium, were also reduced by D. divaricata, D. prolifera, P. cornea, G. lanceolata, and G. filicina in a dose-dependent manner (IC50 values for TNF-α were 16.11, 28.21, 84.27, 45.52 and 74.75 µg/mL, respectively; IC50 values for IL-6 were 37.35, 80.08, 103.28, 62.53 and 84.28 µg/mL, respectively). The total phlorotannin content was measured by the Folin-Ciocalteu method and expressed as phloroglucinol equivalents. The content was 92.0 µg/mg for D. divaricata, 151.8 µg/ mg for D. prolifera, 57.2 µg/mg for P. cornea, 53.0 µg/mg for G. lanceolata, and 40.2 µg/mg for G. filicina. Conclusions: Thus, these findings suggest that Jeju seaweed extracts have potential therapeutic applications for inflammatory responses.
    6  Antioxidant potential of Rumex vesicarius L.: in vitro approach
    Tajdar Husain Khan Majid Ahmad Ganaie Nasir Ali Siddiqui Aftab Alam Mohd Nazam Ansari
    2014(7):538-544. DOI: 10.12980/APJTB.4.2014C1168
    [Abstract](29) [HTML](0) [PDF 530.39 K](77)
    Abstract:
    Objective: To assess in-vitro antioxidant activity of different fraction and perform high performance thin layer chromatography fingerprint analysis of most active fraction of Rumex vesicarius L. (R. vesicarius). Methods: In the present study, acetone, ethyl acetate, n-butanol, and methanol extracts of R. vesicarius were evaluated for radical scavenging activity by studying the inhibition of the level of lipid peroxidation induced by Fe(++)/ascorbate, DNA sugar damage, scavenging of hydrogen peroxide, diphenylphosphine DPPH radical scavenging activity, total phenolic content, total flavonoids content and total proanthocyanidin. High performance thin layer chromatography finger print profiling of R. vesicarius L. was also done. Results: Lipid peroxidation induced by the iron/ascorbate system, hydrogen peroxide, diphenylphosphine and DNA sugar damage were inhibited by the addition of different extract of R. vesicarius. Among them, methanolic extract showed maximum efficacy. The methanolic extract showed the highest total phenolic, total flavonoids and total proanthocyanidin contents. Conclusions: The results suggest that the extracts can be a vital source of phytochemical antioxidants.
    7  Cytotoxicity screening of Melastoma malabathricum extracts on human breast cancer cell lines in vitro
    Nurfariza Ahmad Roslen Nur Aizura Mat Alewi Hadji Ahamada Mohammad Syaiful Bahari Abdull Rasad
    2014(7):545-548. DOI: 10.12980/APJTB.4.2014C658
    [Abstract](19) [HTML](0) [PDF 490.40 K](88)
    Abstract:
    Objective: To screen the cytotoxic activity of Melastoma malabathricum (M. malabathricum) against human breast cancer cell line (MCF-7) in vitro. Methods: A three steps extraction protocol using n-hexane, chloroform and methanol as the solvents systems was carried out on leaves, stems and flowers of M. malabathricum. Dimethyl sulfoxide was used in extracts dilution and serial dilutions were conducted to obtain five different extract concentrations (100 µg/mL, 50 µg/mL, 25 µg/mL, 12.5 µg/mL and 6.25 µg/mL). The evaluation of cell growth was determined using methylene blue assay. Results: Methanol extract from the leaves showed significant anticancer activity against MCF-7 cell lines with the IC50 value of 7.14 µg/ml while methanol and chloroform extract from the flowers exhibited a moderate activity towards MCF-7 cell line with the IC50 value of 33.63 µg/mL and 45.76 µg/mL respectively after 72 h of treatment. Conclusions: The extracts from leaves and flowers of M. malabathricum showed promising anticancer activity toward human breast cancer cell lines with the lowest IC50 at 7.14 µg/mL while the extracts from stems showed less growth inhibition activity.
    8  In vitro cytotoxicity of Indonesian stingless bee products against human cancer cell lines
    Paula M. Kustiawan Songchan Puthong Enos T. Arung Chanpen Chanchao
    2014(7):549-556. DOI: 10.12980/APJTB.4.2014APJTB-2013-0039
    [Abstract](26) [HTML](0) [PDF 608.82 K](80)
    Abstract:
    Objective: To screen crude extracts of propolis, bee pollen and honey from four stingless bee species [Trigona incisa (T. incisa)], Timia apicalis, Trigona fusco-balteata and Trigona fuscibasis) native to East Kalimantan, Indonesia for cytotoxic activity against five human cancer cell lines (HepG2, SW620, ChaGo-Ⅰ, KATO-Ⅲ and BT474). Methods: All samples were extracted with methanol, and then subpartitioned with n-hexane and ethyl acetate. Each crude extract was screened at 20 µg/mL for in vitro cytotoxicity against the cell lines using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. In addition, four previously shown bioactive components from propolis (apigenin, caffeic acid phenyl ester, kaempferol and naringenin) and two chemotherapeutic drugs (doxorubicin and 5-fluorouracil) were used to evaluate the sensitivity of the cell lines. Results: Overall, crude extracts from propolis and honey had higher cytotoxic activities than bee pollen, but the activity was dependent upon the extraction solvent, bee species and cell line. Propolis extracts from T. incisa and Timia apicalis showed the highest and lowest cytotoxic activity, respectively. Only the HepG2 cell line was broadly sensitive to the honey extracts. For pure compounds, doxorubicin was the most cytotoxic, the four propolis compounds the least, but the ChaGo-Ⅰ cell line was sensitive to kaempferol at 10 µg/mL and KATO-Ⅲ was sensitive to kaempferol and apigenin at 10 µg/mL. All pure compounds were effective against the BT474 cell line. Conclusions: Propolis from T. incisa and Trigona fusco-balteata contain an in vitro cytotoxic activity against human cancer cell lines. Further study is required, including the isolation and characterization of the active antiproliferative agent(s).
    9  Modified technique of Wolbachia removal from Malaysian Aedes albopictus
    Sylvia Joanne Indra Vythilingam Nava Yugavathy Jonathan Inbaraj Doss
    2014(7):557-560. DOI: 10.12980/APJTB.4.2014APJTB-2014-0020
    [Abstract](23) [HTML](0) [PDF 305.84 K](76)
    Abstract:
    Objective: To develop an artificial and modified Wolbachia removal technique using tetracycline from naturally Wolbachia infected Aedes albopictus (Ae. albopictus) so as to be able to produce generations of Wolbachia free offsprings. Methods: In this study, seven different tetracycline treatment methods were conducted to obtain the best removal method. Four methods focused on larvae tetracycline treatment, one method on both larvae and adult tetracycline treatment and the last two methods on adult mosquito sucrose treatment. Results: All larval tetracycline treatments resulted in either high larvae mortality, sterile F0 adult mosquitoes or unsuccessful Wolbachia removal. Treatment of both larvae and adults resulted in reduced larvae mortality, successful Wolbachia removal but slow mosquito fecundity. As for the adult treatment, 1.0 mg/mL as previously published was not able to completely remove Wolbachia in F1 generation whereas 1.25 mg/mL successfully removed Wolbachia from F1 and F2 mosquitoes in 2 weeks. Conclusions: This method is different from the previously published methods as it provides an improved Wolbachia removal technique from Ae. albopictus with high egg hatchability, low larvae mortality, increased fecundity and better Wolbachia removal rate.
    10  An initial study of insect succession on decomposing rabbit carrions in Harare, Zimbabwe
    Nyasha Mabika Ron Masendu Gilbert Mawera
    2014(7):561-565. DOI: 10.12980/APJTB.4.2014C1031
    [Abstract](17) [HTML](0) [PDF 230.28 K](72)
    Abstract:
    Objective: To investigate insects visiting sun exposed and shaded decomposing rabbit carcasses and to establish the relationship between insects and carcasses which may be of forensic importance in Harare. Methods: Two rabbits weighing 2.3 kg and 2.5 kg were killed by sharp blows on the head. One was exposed to the sun while the other was placed under shade. The carcasses were allowed to decompose and insects were collected twice a day for the first week and thereafter once a day up to the end of the 7 weeks. Maggots were also collected from the decomposing carcasses and reared. Results: Five dipteran families (Calliphoridae, Muscidae, Sarcophagidae, Phoridae and Drosophilidae) were identified from the sun-exposed carcass. Species collected included Lucilia cuprina (L. cuprina), Chrysomya albiceps (C. albiceps), Musca domestica, Sarcophaga sp. and Drosophila sp. Four families (Calliphoridae, Muscidae, Phoridae, Anthomyiidae) were identified from the shaded carcass. Representatives of these families included L. cuprina, C. albiceps, Musca domestica, and Hydrotaea sp. Three Coleopteran families (Histeridae, Cleridae and Dermestidae) were identified from both carcasses. The observed species were Saprinus sp., Necrobia rufipes and Dermestes sp. Formicidae (Hymenoptera) was represented by only one species (Pheidole sp.). Flies which emerged from the rearing units were L. cuprina, Lucilia sp., C. albiceps, Sarcophaga sp. and Sepsis sp.). Conclusions: Of the dipteran species collected during the study, L. cuprina and C. albiceps could be important for further forensic studies since they were collected from the carcasses and also observed from the rearing units.
    11  Knowledge, attitude, and practices related to cutaneous leishmaniasis in an endemic focus of cutaneous leishmaniasis, Southern Iran
    Bahador Sarkari Asgari Qasem Mohammad Reza Shafaf
    2014(7):566-569. DOI: 10.12980/APJTB.4.2014C744
    [Abstract](18) [HTML](0) [PDF 262.18 K](72)
    Abstract:
    Objective: To assess knowledge, attitudes, and practices (KAP) of inhabitants of an endemic area in Fars province toward cutaneous leishmaniasis (CL). Methods: The study was carried out in Lapui district in Fars province, south of Iran, one of the most important foci of CL in this province. Sample size (237 residents) was calculated based on population. House-to-house survey was done to collect the data regarding knowledge, attitudes, and practices of the inhabitants. The head of each household was interviewed by a trained staff to assess his/her KAP related to CL. A semi-structured KAP questionnaire was used for data collection. Results: Mean age of participants was 39 and more than half of the respondents were in the age group of 31-40. Males constituted 172 (72.5%) of subjects. Most of the respondents (84.3%) were literate. The majority of the study population (83%) had heard about Salak (local name for CL) and most of these respondents (91%) were aware that CL is presented with a cutaneous lesion. Nearly two-third of the participants (63.5%) stated the bite of mosquito (not specifically sandflies) for CL transmission. The respondents’ attitude regarding the treatment of CL was not satisfactory since only 48% believed that CL can be treated by medicine. A noticeable proportion of respondents (21%) believed in indigenous medicine for the treatment of CL. A small proportion of respondents (14%) stated that traditional healers are good at treating this disease. More than two-third (69%) of respondents believed that the disease is preventable although most of interviewees did not know about preventive measures. Conclusions: In this study, insufficient knowledge of community about infection nature, vector, transmission mode and preventive measures of CL, highlights the needs for a health education initiative to enhance the awareness of people about CL. This would improve inhabitants’ contribution in control program of CL in this area.
    12  Down-regulated expression of NPM1 in IMS-M2 cell line by (-)-epigallocatechin-3-gallate
    Hoang Thanh Chi Bui Thi Kim Ly Hoang Anh Vu Yuko Sato Phu Chi Dung Phan Thi Xinh
    2014(7):570-574. DOI: 10.12980/APJTB.4.2014APJTB-2014-0177
    [Abstract](20) [HTML](0) [PDF 679.26 K](79)
    Abstract:
    Objective: To investigate the inhibited effect of epigallocatechin-3-gallate (EGCG) on the expression of NPM1 in IMS-M2 cells harboring the NPM1 mutations. Methods: Cell proliferation assay was performed to test the effects of EGCG on cell growth of IMS-M2 cells harboring the NPM1 mutations. Western blot analysis were performed to test the protein expression of NPM1, AKT, those associated with apoptosis. Results: EGCG can down-regulate the expression of NPM1 in IMS-M2 cells harboring the NPM1 mutations. Moreover, EGCG also suppressed the cell proliferation and induced apoptosis in IMS- M2 cells. Conclusions: The results suggested that EGCG could be considered as a reagent for treatment of AML patients with NPM1 mutations.
    13  Anti-diabetic effects of Caulerpa lentillifera: stimulation of insulin secretion in pancreatic β-cells and enhancement of glucose uptake in adipocytes
    Bhesh Raj Sharma Dong Young Rhyu
    2014(7):575-580. DOI: 10.12980/APJTB.4.2014APJTB-2014-0091
    [Abstract](24) [HTML](0) [PDF 673.58 K](89)
    Abstract:
    Objective: To evaluate anti-diabetic effect of Caulerpa lentillifera (C. lentillifera). Methods: The inhibitory effect of C. lentillifera extract on dipeptidyl peptidase-Ⅳ and α-glucosidase enzyme was measured in a cell free system. Then, interleukin-1β and interferon-γ induced cell death and insulin secretion were measured in rat insulinoma (RIN) cells by 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and ELISA kit, respectively. Glucose uptake and glucose transporter expression were measured by fluorometry and western blotting, using 3T3-L1 adipocytes. Results: C. lentillifera extract significantly decreased dipeptidyl peptidase-Ⅳ and α-glucosidase enzyme activities, and effectively inhibited cell death and iNOS expression in interleukin-1β and interferon-γ induced RIN cells. Furthermore, C. lentillifera extract significantly enhanced insulin secretion in RIN cells and glucose transporter expression and glucose uptake in 3T3-L1 adipocytes. Conclusions: Thus, our results suggest that C. lentillifera could be used as a potential anti-diabetic agent.
    14  Clinical, radiological and molecular diagnosis correlation in serum samples from patients with osteoarticular tuberculosis
    Guadalupe García-Elorriaga Olga Martínez-Elizondo Guillermo del Rey-Pineda César González-Bonilla
    2014(7):581-585. DOI: 10.12980/APJTB.4.201414B112
    [Abstract](17) [HTML](0) [PDF 352.41 K](74)
    Abstract:
    Objective: To assess the role of polymerase chain reaction (PCR) in serum samples, in the diagnosis of osteoarticular tuberculosis (OTB) in a setting where only clinical and imaging diagnoses determine the treatment. Methods: A total of 44 consecutive serum specimens were collected from clinically suspected OTB patients, based on clinical and radiological [X-ray or magnetic resonance imaging/computed tomography] features. They were screened by in-house nested PCR. In addition, a few specimens were examined by Gram stain, acid-fast bacilli stain, histopathology and routine bacterial culture. A total of 39 specimens were collected from patients suffering from other bone diseases of nontuberculous origin and included as negative controls. Results: Of the 44 clinically suspected OTB patients, in-house nested PCR was positive in 40 (91%) cases; PCR was negative in 38 (97%) negative controls. Sensitivity and specificity of our inhouse nested PCR was 90.9% and 97.4%, respectively. The PCR report was available within 48 h. It was possible to standardize serum PCR technique and in positive cases, a good correlation was observed in terms of an adequate treatment response. Conclusions: Nested PCR in serum samples is a rapid, highly sensitive and specific modality for OTB detection. PCR should be performed in addition to clinical evaluation, imaging studies, acid-fast bacilli staining, culture and histopathology diagnosis, if possible.
    15  Reticulo-cutaneous fistula due to the ingestion of a long metallic rod in a cow
    Azari Omid Ali Asghar Mozaffari
    2014(7):586-588. DOI: 10.12980/APJTB.4.2014B102
    [Abstract](38) [HTML](0) [PDF 409.69 K](72)
    Abstract:
    A five-year-old cow was referred to the veterinary faculty hospital for treatment of a swelling mass with a cutaneous fistula at the left lower part of the chest wall, between 7th till 9th intercostals space. Abdominal pain in palpation of the mass was observed with no abnormality in clinical symptoms. In surgical exploration, skin incision was carried out on the swelling mass and surprisingly a sinus tract with a sharp metallic rod (26 cm length) that continued to the reticulum lumen was identified. Because of unsuccessful attempting to remove of the foreign body, flank laparotomy and rumenotomy was performed. After one month, the cow led to complete recovery.

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