Asian Pacific Journal of Tropical Biomedicine

977-981

Authors: Shimaa Abd El-Salam El-Sayed, Mohamed Abdo Rizk, Mohamed Alaa Terkawi, Ahmed Mousa4 , El Said El Shirbini El Said, Gehad Elsayed, Mohamed Fouda , Naoaki Yokoyama , Ikuo Igarashi

Objective: To use two diagnostic antigens belonging to the frequently associated in Theileria domain, Theileria equi (T. equi) protein 82 (Te 82) and T. equi 104 kDa microneme-rhoptry antigen precursor (Te 43), to diagnose T. equi infection in horses as compared with equi merozoite antigen-2 (EMA-2). Methods: In the current study, we applied a cocktail-ELISA containing two antigens (EMA-2 + Te 82) to diagnose T. equi infection either in experimentally infected horses or in field infection. Results: Our findings have revealed that a cocktail formula of EMA-2 + Te 82 provided a more practical and sensitive diagnostic candidate for diagnosing T. equi infection in horses as compared with Te 82 or Te 43 alone. Conclusions: The ELISA technique using a cocktail formula of EMA-2 + Te 82 offers a practical and sensitive diagnostic tool for diagnosing T. equi infection in horses and using of this promising cocktail formula will be applicable for epidemiological surveys and will help control the infection in horses.

987-991

Authors: Maryam Safarpour Dehkordi, Abbas Doosti, Asghar Arshi

Objective: To construct a novel plasmid as Salmonella enterica serovar typhimurium (S. typhimurium) sipC gene knockouts candidate. Methods: In this research, 50 upstream and 30 downstream regions of S. typhimurium sipC gene and kanamycin gene were PCR amplified. Each of these DNA fragment was cloned into pGEM T-easy vector. The construct was confirmed by PCR and restriction digest. Results: PCR amplified 320, 206 and 835 bp DNA fragments were subcloned into pET- 32 vector resulting with a plasmid called pET-32-sipC up-kan- sip C down. Conclusions: The new plasmid (pET-32-sipC up-kan- sip C down) is useful for genetic engineering and for future manipulation of S. typhimurium sipC gene.

996-1004

Authors: De-Hu Chen, Rui-Qi Lu, Xiao-Chun Ni, Ju-Gang Wu, Shou-Lian Wang, Bo-Jian Jiang, Ji-Wei Yu

Objective: To investigate the influence of CD133+ expression on patients' survival and resistance of CD133+ cells to anti-tumor agents in gastric cancer (GC). Methods: Influence of CD133 expression on prognosis was analyzed employing samples from patients with GC. GC cell lines were utilized to separate CD133+ and CD133− subpopulations by immunomagnetic separation and to analyze the biological features of two subpopulations in vitro and in vivo, especially in resistant to anti-tumor reagents and its apoptotic mechanism. Results: The lower CD133+ group showed a significantly better survival compared with the higher CD133+ group. The highest content of CD133+ subpopulations for KATO-III cells had stronger proliferative ability than CD133− subpopulations. A single CD133+ cell was capable of generating new cell colony and the tumorigenicity rate in nude mice was 100% for CD133+ clonal spheres or for CD133+ cells, but 0% for CD133− cells. Furthermore, the higher expression levels of Oct-4, Sox-2, Musashi-1 and ABCG2 in CD133+ clonal spheres were identified compared with CD133+ cells or CD133− cells. Under the treatment of anti-tumor reagents, CD133+ cells had lower suppression rates compared with CD133− cells while lower level of Bcl-2 and higher level of Bax were found in CD133+ cells compared with CD133− cells. Conclusions: The patients with lower CD133+ expression had a better survival. Enriched CD133+ cells in clonal sphere shared the ability to be self-renewable, proliferative, tumorigenic and resistant to anti-tumor agents as probably regulated by Bcl-2 and Bax.

1010-1017

Authors: Supranee Upanan, Kanjana Pangjit, Chairat Uthaipibull, Suthat Fucharoen, Andrew T. McKie, Somdet Srichairatanakool

Objective To evaluate the efficacy of deferiprone (DFP), 1-(N-acetyl-6-aminohexyl)-3-hydroxy-2-methylpyridin-4-one (CM1) or green tea extract (GTE) in enhancing expression of hepatic hepcidin1 (Hamp1) mRNA and relieving iron overload in β-globin knockout thalassemic mice. Methods The β-globin knockout thalassemic mice were fed with a ferrocene-supplemented diet for 2 months and oral administration of deionized water, DFP (50 mg/kg), CM1 (50 mg/kg), GTE (50 mg epigallocatechin 3-gallate equivalent/kg), GTE along with DFP (50 mg/kg), and GTE along with CM1 (50 mg/kg) every day for 3 months. Levels of hepatic Hamp1 mRNA, plasma non-transferrin bound iron, plasma alanine aminotransferase activity and tissue iron content were determined. Results All chelation treatments could reduce plasma non-transferrin bound iron concentrations. Additionally, hepatic Hamp1 mRNA expression was significantly up-regulated in the mice in a GTE + DFP combined treatment, correlating with a decrease in the plasma alanine aminotransferase activity and tissue iron deposition. Conclusions The GTE + DFP treatment could ameliorate iron overload and liver oxidative damage in non-transfusion dependent β-thalassemic mice, by chelating toxic iron in plasma and tissues, and increasing hepcidin expression to inhibit duodenal iron absorption and iron release from hepatocytes and macrophages in the spleen. There is probably an advantage in giving GTE with DFP when treating patients with iron overload.

1027-1032

Authors: Yosephine Sri Wulan Manuhara, Alfinda Novi Kristanti, Edy Setiti Wida Utami, Arif Yachya

Objective To increase biomass and saponin production in hairy root culture of Talinum paniculatum Gaertn. (T. paniculatum) in balloon-type bubble bioreactor (BTBB). Methods Hairy roots which were collected from leaf explants of T. paniculatum were infected by Agrobacterium rhizogenes strain LB510. The hairy roots were cultivated at 400 mL Murashige and Skoog liquid medium without growth regulator (MS0) in 1 000 mL BTBB. Each BTBB had 2 g hairy roots as initial inoculum and these cultures were treated with various concentrations of sucrose (3%, 4%, 5%, 6% w/v) and potassium nitrate (0.5, 1.0, 1.5 and 2.0 strength of MS medium). Cultures were maintained for 14 days. Fresh and dry weights of hairy roots at the end of culture were investigated. Results Various concentrations of sucrose influenced the biomass accumulation of hairy roots. Maximum biomass was reached by MS medium supplemented with 6% sucrose and it was approximately threefold higher than control. Culture supplemented with potassium nitrate at 2.0 strength of MS0 could increase biomass accumulation of hairy roots until 0.14 g dry weight and it was almost threefold higher than control. However, the maximum saponin content was obtained by MS medium supplemented with 5% sucrose and 2.0 strength potassium nitrate of MS. Conclusions Based on this research, those conditions can be used to produce biomass and saponin of hairy root of T. paniculatum in the large scale.

1037-1041

Authors: Ourlad Alzeus G. Tantengco, Sonia D. Jacinto

Objective To evaluate the cytotoxic activities of Premna odorata (P. odorata) leaves and bark, Artocarpus camansi (A. camansi) and Gliricidia sepium against selected human cancer cell lines by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Methods The crude extracts of P. odorata, A. camansi and Gliricidia sepium were subjected to liquid–liquid partitioning by using hexane and ethyl acetate to separate compounds based on their polarity. The fractions were tested for their cytotoxic activity against human colon cancer cell line (HCT116), breast cancer cell line (MCF-7), lung adenocarcinoma cell line (A549) and Chinese hamster ovary cell line (AA8) by using MTT assay. Results Based on the standard values of toxicity set by the study of Suffness and Pezzuto, P. odorata leaves and P. odorata bark hexane fractions and A. camansi leaves were all considered highly cytotoxic against the selected human cancer cell lines. P. odorata bark hexane extract exhibited the highest selectivity index for HCT116, MCF-7 and A549 cancer cell lines. Conclusions The results obtained indicated that P. odorata leaves and bark and A. camansi leaves have excellent cytotoxic activity and warrant further studies to isolate novel compounds for chemotherapeutic use.

1046–1050

Authors: Erna Susanti, Ciptati, Retty Ratnawati, Aulanni'am, Achmad Rudijanto

Objective To identify the bioactive compounds in catechins isolation and its components from green tea GMB-4 clone. Methods Green tea GMB-4 clones were extracted with distilled water at 90 °C. Samples were eluted into the column with 10% ethanol. Subsequently, the column was eluted with 95% ethanol and evaporated separately. Green tea extract was identified by thin layer chromatography. Catechins were separated by the stationary phase in column chromatography using polyamide with 10% ethanol eluent and 95% ethanol. The results of isolations were analyzed by high performance liquid chromatographic (HPLC) and LC-MS/MS. Analysis of catechins by HPLC was done by external standard. Results Fraction from 10% ethanol showed that four major peaks at retention time of 1.663, 2.367, 2.950 and 4.890, indicated the presence of four catechins components including catechin, epicatechins, gallocatechin and epigallocatechin. Whereas, fraction from 95% ethanol showed two main peaks at retention time of 5.167 and 9.82, which indicated the presence of epigallocatechin gallate (EGCG) and epicatechin gallate (ECG). EGCG (m/z 459), epigallocatechin (m/z 307), ECG (m/z 443), and epicatechin (m/z 291) were isolated and separated successfully using HPLC and LC-MS/MS. Conclusions The HPLC and LC-MS/MS methods were successfully tuned for the qualitative analysis of green tea extract with EGCG and ECG. Four major catechins were separated and identified by LC-MS/MS, such as EGCG, epigallocatechin, ECG and epicatechin. The result of HPLC analysis showed that EGCG and ECG were main components from catechins isolation of green tea GMB-4 clone.

1055-1059

Authors: Aymen Mudawe Nurain, Naser Eldin Bilal, Mutasim Elhadi Ibrahim

Objective To determine the prevalence and antimicrobial resistance rates of nosocomial pathogens isolated from cancer patients and hospital environments. Methods A descriptive cross-sectional study was conducted between December 2010 to May 2013 at Radiation and Isotopes Centre of Khartoum, Sudan. A total of 1 503 samples (505 clinical and 998 environmental) were examined. Isolates were identified, and their antimicrobial susceptibility was determined using standard laboratory procedures. Results Out of 505 clinical samples, nosocomial pathogens were found as 48.1%. Among hospital environment samples, bacterial contaminants were detected in 29.7% of samples. The main microorganisms recovered from cancer patients were Proteus spp. (23.5%), Escherichia coli (22.2%), Pseudomonas aeruginosa (P. aeruginosa) (21.0%) and Staphylococcus aureus (20.2%). The most frequent isolates from hospital environments were Bacillus spp. (50.0%), Staphylococcus aureus (14.2%) and P. aeruginosa (11.5%). The proportions of resistance among Gram-negative pathogens from cancer patients were high for ampicillin, cefotaxime, ceftazidime and ceftriaxone. Moderate resistance rates were recorded to ciprofloxacin, such as 51.0% for P. aeruginosa, 21.7% for Klebsiella pneumoniae and 55.5% for Escherichia coli. Except Klebsiella, there were no significant differences (P ≥ 0.05) of resistance rates between Gram-negative isolates from cancer patients to those from the hospital environments. The proportions of extended-spectrum β-lactamase producing isolates from cancer patients were not differ significantly (P = 0.763) from those collected from the hospital environments (49.2%; 91/185 vs. 47%; 32/68). Conclusions The prevalence of nosocomial infection among cancer patients was high (48.1%) with the increasing of antimicrobial resistance rates. Hospital environments are potential reservoirs for nosocomial infections, which calls for intervention program to reduce environmental transmission of pathogens.