Asian Pacific Journal of Tropical Biomedicine

730–734

Authors: Aioub Sofizadeh, Hamideh Edalat, Mohammad Reza Abai, Ahmad Ali Hanafi-Bojd

Objective: To determine fauna and some ecological aspects of Anopheles mosquitoes in northeast of Iran. Methods: In this descriptive study, 3 villages in Kalaleh County were selected in different geographical zones. Anopheles mosquitoes were collected biweekly from May to October using standard dipping method for larvae, and hand catch, total catch, artificial pit shelter as well as night-biting collections on human and animal baits for adults. Results: Totally 399 larvae and 2602 adults of Anopheles mosquitoes were collected and identified as 2 species: Anopheles superpictus s.l. (An. superpictus s.l.) and Anopheles maculipennis s.l. The dominant species was An. superpictus s.l. (92.1%). Activity of these mosquitoes found to be started from middle of May and extended till September with two peaks of activity in July and August. Conclusions: An. superpictus s.l. as one of the main malaria vectors in Iran as well as some other parts of the world is the dominant species in the study area. This species has high potential for transmission and possibility of establishing a transmission cycle with low abundance. Other species, Anopheles maculipennis s.l. also has introduced as a malaria vector in northern parts of Iran. As this Anopheles is a complex species, genetic studies are recommended to determine the members of this complex in the study area.

740–744

Authors: Rosella Camere-Colarossi, Gabriela Ulloa-Urizar, Dyanne Medina-Flores, Stefany Caballero-García, Frank Mayta-Tovalino, Juana del Valle-Mendoza

Objective: To evaluate the antibacterial and cytotoxic effect of Myrciaria dubia (Camu camu) (M. dubia) methanol extract, against Streptococcus mutans (ATCC 25175) (S. mutans) and Streptococcus sanguinis (ATCC 10556) (S. sanguinis). Methods: Two methanol extracts of M. dubia were prepared in vitro, from the seeds and pulp. Ten independent tests were prepared for each type of extract, using 0.12% chlorhexidine solution as positive control. Agar diffusion test was used by preparing wells with the experimental solutions cultivated in anaerobic conditions for 48 h at 37
C. Meanwhile, the minimum inhibitory concentration and the cytotoxic effect over MDCK cell line was found. Results: A higher antibacterial effect was observed with the methanol seed extract with an inhibitory halo of (21.36 ± 6.35) mm and (19.21 ± 5.18) mm against S. mutans and S. sanguinis, respectively. The methanol extract of the pulp had an effect of (16.20 ± 2.08) mm and (19.34 ± 2.90) mm, respectively. The minimum inhibitory concentration of the pulp extract was 62.5 mg/mL for both strains, whereas for the seed antibacterial activity was observed even at low concentrations. The CC50 of the seeds extract was at a higher concentration than 800 mg/mL and 524.37 mg/mL for the pulp extract. Conclusions: The experimental findings demonstrated the antibacterial effect of the methanol extract of M. dubia against S. mutans and S. sanguinis. These extracts were not cytotoxic at high concentrations.

751–754

Authors: Azis Saifudin, Alfian Mahardika Forentin, Arini Fadhilah, Kuswandi Tirtodiharjo, Witri Dyah Melani, Devita Widyasari, Tri Agus Saroso

Objective: To search an herbal material, capable of inhibiting plaque producing bacteria Streptococcus mutans. Methods: Twenty materials comprising 10 flowers and 10 rhizomes were extracted with 70% ethanol. Their activity was then examined at a concentration of 10% (w/v) against Streptococcus mutans in vitro on Mueller–Hinton media. Erythromycin (Oxoid, 20 mg disc) was used as a positive control. Meanwhile, to establish a fingerprint guide for authentication or quality control, the most potent material was further analyzed regarding its chemical constituents by means of reversed phase-high performance liquid chromatography (HPLC) and thin-layer chromatography (TLC). Results: Of the tested samples, Sesbania grandiflora (S. grandiflora) flower and Costus speciosus rhizome extracts showed the most potent activity with inhibited zone diameters of 18.5 and 14.8 mm, respectively. On the other hand, other extract plants showed a diameter zone in the range of 0.5–10.6 mm or being inactive (diameter = 0 mm). The activity of S. grandiflora was comparable to that of erythromycin (diameter = 18.0 mm). The best separation was achieved on HPLC system with acetonitrile-water with a ratio of 2:8, and a flow rate at 0.5 mL/min. TLC, meanwhile, was featured on chloroform– methanol (8.5:1.5) as a mobile system. Conclusions: S. grandiflora flower is a promising material to be developed as the active ingredient of anti-plaque toothpaste as well as mouthwash solution. The developed HPLC and TLC system can be used for a further standard in its material authentication as well as for a fingerprinting of quality control during the manufacturing process.

760–766

Authors: Amina Benabdallah, Chaabane Rahmoune, Mahieddine Boumendjel, Oumayma Aissi, Chokri Messaoud

Objective: To investigate the total phenolics, flavonoids and tannins content and the in vitro antioxidant activities of methanolic extracts of six wild Mentha species which are Mentha aquatica, Mentha arvensis, Mentha piperita, Mentha pulegium, Mentha rotundifolia and Mentha villosa. Methods: The Folin–Ciocalteu method was used to determine the total phenols content while flavonoids were estimated according to the aluminum chloride colorimetric method. To evaluate tannins content, vanillin and HCl were added to methanolic extracts. The antioxidant potential was measured by 1,1-diphenyl-2-picrylhydrazyl radical scavenging, ferrous ion chelating and the inhibition of b-carotene bleaching assays. Results: The methanol extracts of Algerian mints were rich in phenolic compounds and exhibited powerful antioxidant activity ranging from 7.5 mg/mL to 44.66 mg/mL, which varied significantly among species. Mentha aquatica stood out with efficient antioxidant ability which was correlated to the high total phenolics content, followed by Mentha arvensis and Mentha piperita with very close values, comparing to Mentha pulegium, Mentha rotundifolia and Mentha villosa with lowest values. Conclusions: These results show that methanolic extracts of Mentha species from Algeria have a great potential of polyphenols which can be used as a natural food preservative and antioxidant source.

771–777

Authors: Abd El-Moneim Mohamed Radwan Afify, Hazem Mohamed Mahmoud Hassan

Objective: To evaluate three flowers of Hibiscus rosa-sinensis (H. rosa-sinensis), Quisqualis indica (Q. indica) and Senna surattensis (S. surattensis) for their antioxidant activity by different methods in addition to total phenolic, flavonoid and pigment contents. Methods: Antioxidant activity of water, ethanol and absolute ethanol extracts of three flowers; H. rosa-sinensis, Q. indica and S. surattensis was evaluated. The antioxidant activity was assessed by 1,1-diphenyl-2-picrylhydrazyl free radical scavenging activity, ferrous chelating activity, reducing power, nitric oxide scavenging activity, hydroxyl radical scavenging activity as well as total antioxidant capacity. Total flavonoids, total phenols and total pigments including chlorophylls and carotenoids were measured for the three flowers. Results: The results showed that the highest total antioxidant capacity at concentration of 500 mg/L was found in S. surattensis as 0.479 ± 0.001. Scavenging activity of H. rosasinensis, Q. indica and S. surattensis flower extracts against 1,1-diphenyl-2- picrylhydrazyl radical showed the highest activity of (90.20 ± 0.29)% with 500 mg/L. Phytochemical screening of the three flowers extracts were carried out for alkaloids, flavonoids, saponins, tannins, steroids, glycosides, terpenoids, amino acid and mucilages. H. rosa-sinensis showed the total phenolic in water extract of (235.77 ± 14.31) mg/100 g, the other two flowers Q. indica and S. surattensis had the total phenolic in ethanol extracts of (937.70 ± 25.06) and (850.30 ± 13.81) mg/100 g, respectively. On the other hand total flavonoids were identified in absolute ethanol extracts in the three flowers [(32.83 ± 1.34), (49.24 ± 4.87) and (2.79 ± 0.23) mg/100 g, respectively]. Conclusions: The extracts in the constituents of the three flowers could be used as additives as supplement fractions in foods.

785–794

Authors: Fernanda Furlan Gonçalves Dias, Ana Lúcia Tasca Gois Ruiz, Adriana Della Torre, Helia Harumi Sato

Objective: To explore the anti-proliferative activity of purified L-asparaginase from Aspergillus oryzae CCT 3940 (A. oryzae). Methods: L-asparaginase was produced by submerged fermentation and purified to electrophoresis homogeneity by ionic exchanged chromatography in a fast protein liquid chromatographic system. The purified enzyme was characterized and used for the antiproliferative assay against nine tumor cell lines and one non-tumor cell line. Results: The free glutaminase L-asparaginase was purified 28.6 fold. L-asparaginase showed high stability under physiological condition, remaining stable in the pH range 7.0–8.0 after 1 h incubation at temperature range 30–45
C. The Km and Vmax values of purified L-asparaginase were estimated as 0.66 mmol/L and 313 IU/mL, respectively. The purified enzyme could inhibit the growth of a broad range of human tumor cell lines at the concentrations studied. Also, the enzyme from A. oryzae CCT 3940 could inhibit tumor growth of leukemia cell line (K562) with a total growth inhibition value of (3.2 ± 2.5) IU/ mL and did not inhibit the non-carcinogenic human cell line growth at the concentrations studied. Conclusions: The sensitivity of the cells lines to purified L-asparaginase from A. oryzae CCT 3940 appeared to be concentration dependent affording a more significant decrease in cell growth than that observed for the commercial L-asparaginase from Escherichia coli. The L-asparaginase from A. oryzae CCT 3940 has a high potential for pharmaceutical exploitation in the treatment of leukemia.

801–802

Authors: Ouarda Ayadi, Mohamed Gharbi, Mohamed Cherif Benchikh Elfegoun

The authors studied the impact of tropical theileriosis onset on milk yield decrease in 10 local bred cows in Skikda (Northern Algeria) during 2015 summer season. The milk yield decrease estimated weekly during two months was 2.76 L/day/cow corresponding to 31.92% of the total milk yield. This decrease corresponds to 110.5 Algerian Dinars (1.02 US$)/day/diseased cow. The relative variation of milk yield showed a dramatic decrease from 82.72% to 0.76% at Day 21 then became constant. Further studies are needed to improve these estimations of financial losses due to bovine tropical theileriosis in Algeria.

808–814

Authors: Mohamed Amine Gacem, Aminata Ould El Hadj-Khelil

Mycotoxins and their derivatives since their discoveries and until the present time are behind unspecified economic and medical damages. Aflatoxins are classified according to their physical–chemical and toxicological characters in the most dangerous row of the mycotoxins. These aflatoxins are in part responsible, of irreversible medical disasters that are not easily manageable such as cancer of the liver and kidneys, and in the other part, of losses in the stored cereal products. Based on these crucial findings, monitoring of this toxin became imperative in post-harvest food products, during storage, during transformation chain and even during the long phases of conservation. Vigilance of this toxin is delivered by detection methods using very advanced technologies to respond in the shortest possible times. In addition, the knowledge of factors supporting the biosynthesis of aflatoxins such as the temperature, moisture content, concentration of nitrogen and carbon, and the molecules responsible for the genetic control of the synthesis will be reflected later in the choice of bio-control techniques. This control is currently based on new strategies using the bioactives substances of the plants, the lactic bacteria and some strains of actinomycetes that have good inhibiting activity against aflatoxins with fewer side effects on Man. On the other hand, this brief review summarizes the results of new studies demonstrating the toxicity of the toxin, new detection methods and bio-control.

640–645

Authors: Ghada Mohamed El-Bassiony, John George Stoffolano Jr

Objective: To compare the differences in intake and excretion between Musca domestica and other three species from families Muscidae and Calliphoridae which may help explaining the significance of house fly in the transmission of pathogens. Methods: The four adult species were supplied with two concentrations of sucrose via modified capillary feeder assay system. The two sucrose concentrations were applied to one adult male/each experiment and the elimination spots were counted. Using 0.25 mol/ L sucrose + 0.25% bromophenol blue, one active non-starved male/cup was observed carefully for 1 h to record its behavior. As a growing medium used in bacterial transmission experiments, undiluted trypticase soy broth was used to feed 3-day-old females and males of Musca domestica following two different diets upon emergence and the frequency of elimination spots was estimated. Results: The two Musca species have half the weight of the two Phormia species. Comparing the volume of intake per hour, house fly took as much as the other species, all of which were larger. House fly produced twice, or more, the number of elimination spots/h than the other three species. Feeding the flies a sugar liquid diet resulted in producing more fecal spots than regurgitation spots. The male house flies produced less elimination spots/h when fed with trypticase soy broth than with the two sucrose solutions. Conclusions: House flies eliminated more than the other examined fly species and most of these elimination events were defecation which implicates the fecal route for pathogen transmission by this important vector.

651–657

Authors: Mohamed Saleh Abdelfattah, Mohammed Ismail Youssef Elmallah, Usama Wahid Hawas, Lamia Taha Abou El-Kassema, Mennat Allah Gamal Eid

Objective: To isolate and evaluate the cytotoxic activity of different actinomycetes species isolated from the Red Sea coast in Sharm el-Sheikh, Egypt. Methods: Forty actinomycetes strains were isolated from different sediments and seawater samples collected from the Red Sea coast in Egypt. Actinomycetes were recognized by morphological and microscopic examinations. Cell viability and cytotoxicity induced by the crude extracts on breast cancer cell lines MDA-MB-231 were assessed using methylene blue assay. The strains with promising cytotoxic activity were identified by sequencing and amplifying the 16S rRNA genes. The antibacterial activities of the crude extracts were performed using Kirby–Bauer disc diffusion method. Results: The results indicated that five ethyl acetate extracts exhibited cytotoxicity towards breast cancer cell lines MDA-MB-231. The highest cytotoxic activity was found for the ethyl acetate extracts of EGY2 and EGY39. The isolate EGY3 was identified as a new Streptomyces species, while the actinomycete EGY22 was found to be a member of the genus Nocardiopsis sp. The crude extract of the isolate EGY8 showed slightly high antimicrobial activity against different test microorganisms. Conclusions: The results of the present study reveal that marine sediments of the Red Sea are a potent source of novel species of actinomycetes. The isolates may be useful in discovery of novel bioactive compounds and an important step in the development of microbial natural product research. 1. Introduction Breast cancer is one of the leading causes of cancer death among women

665–670

Authors: Moussa Compaore´, Roland Nag-Tiˆ ero Meda´ , Sahabi Bakasso, Laurian Vlase, Martin Kiendrebeogo

Objective: To assess the antioxidant and anti-inflammatory activities as well as to determine the flavonoids and phenolic acids content of active fractions. Methods: Two medicinal plant samples were extracted successively in Soxhlet apparatus with n-hexane, dichloromethane, acetonitrile, ethyl acetate, methanol and n-butanol. Five methods were used to evaluate the antioxidant activity. Anti-inflammatory activity was done through the inhibition of the cyclooxygenase enzymes (COX-1 and COX-2). Polyphenolic compounds were analyzed by using a spectrophotometrical and high performance liquid chromatography-mass spectrometry (HPLC-MS) methods. Results: The data showed that the stem leaves extracts of Commiphora africana and Loeseneriella africana possessed significant in vitro antioxidant and anti-inflammatory activities. Polar extracts had radical scavenging effects and they reduced iron (III). The prostaglandin production was significantly stopped by acetonitrile and methanol extracts. These biological activities were supported by some bioactive compounds quantified by using the HPLC-MS. p-Coumaric acid, ferulic acid, isoquercitrin, quercitrin, quercetin, rutin, kaempferol and apigenin were the most metabolites quantified. Conclusions: The present study may explain the effectiveness of plants in traditional medicine of Burkina Faso, singularly Commiphora africana and Loeseneriella africana. The next investigation was to sub-fractionate the methanol fraction in order to isolate new antioxidant and/or anti-inflammatory compounds.

677–681

Authors: Christiane Takayama, Felipe Meira de-Faria, Ana Cristina Alves de Almeida, Ricardo Jose Dunder´ 2, Luis Paulo Manzo2, Eduardo Augusto Rabelo Socca, Leonia Maria Batista, Marcos Jose Salvador´ , Alba Regina Monteiro Souza-Brito, Anderson Luiz-Ferreira

Objective: To evaluate the antioxidant activity of the essential oil obtained from Rosmarinus officinalis (R. officinalis) in ethanol-induced gastric ulcer model in vivo. Methods: The antioxidant properties of the essential oil obtained from R. officinalis were evaluated against gastric injury induced by absolute ethanol. Gastric tissues were prepared to enzymatic assays. The levels of glutathione, lipid peroxides, and the activities of glutathione peroxidase, superoxide dismutase and myeloperoxidase were measured. Results: Ethanol produced severe hemorrhagic lesions in the stomach with ulcerative lesion of (140.2 ± 37.2) mm2. In animals pretreated with essential oil of R. officinalis (50 mg/kg, p.o.), a significant inhibition of mucosal injury of (21.2 ± 7.1) mm2 (84% inhibition) was observed. The essential oil of R. officinalis protected the gastric mucosa probably by modulating the activities of the enzymes (superoxide dismutase and glutathione peroxidase) and increasing or maintaining the levels of glutathione. In addition, lipid peroxides levels were reduced. The essential oil of R. officinalis was analyzed by gas chromatography–mass spectrometer and the main constituents were cineole (28.5%), camphor (27.7%) and alpha-pinene (21.3%). Conclusions: We suggest that the monoterpenes present in the essential oil obtained from R. officinalis may be among the active principles responsible for the antioxidant activity shown by essential oil of R. officinalis.

686–691

Authors: Guntur Berlian, Olivia Mayasari Tandrasasmita, Raymond Rubianto Tjandrawinata

Objective: To verify that Proliverenol has a potential ability in protecting cells from ethanol-induced hepatotoxicity. Methods: Activity of Proliverenol against ethanol-induced apoptosis was evaluated at mRNA and protein levels in HepG2 cell exposed to Proliverenol for 1 and 3 h. Results: Proliverenol conferred hepatoprotective activity through increasing cell survival up to 53%–69% via up-regulation of APEX1 DNA repair enzyme for 3.0–4.7 fold and down-regulating of nuclear factor-kB, tumor necrosis factora and caspase-8 expression, allowing them to prevent 4.5–6.9 fold of alanine aminotransferase (ALT) leakage in HepG2 cells. Our finding revealed that Proliverenol repressed expression of ALT, which is significantly important as possible alternative mechanism for increased blood transaminase activities. In addition, the result also showed that caspase-8 pathway seemed to be involved in the molecular pathway rather than directly inducing mitochondrial damage. Conclusions: The data support our hypothesis that Proliverenol has a potential ability in protecting cells from ethanol-induced hepatotoxicity. We propose that Proliverenol provides hepatoprotective activity through up-regulating expression of APEX1 that repress DNA fragmentation, and down-regulating expression of nuclear factor-kB, tumor necrosis factora and caspase-8, which therefore repress ALT leakage and its expression.

698–701

Authors: Aylin Ugurlu, Aysegul Karahasan Yagci, Seyhan Ulusoy, Burak Aksu, Gulgun Bosgelmez-Tinaz

Objective: To investigate the effects of plant-derived phenolic compounds (i.e. caffeic acid, cinnamic acid, ferulic acid and vanillic acid) on the production of quorum sensing regulated virulence factors such as pyocyanin, biofilm formation and swarming motility of Pseudomonas aeruginosa (P. aeruginosa) isolates. Methods: Fourteen clinical P. aeruginosa isolates obtained from urine samples and P. aeruginosa PA01 strain were included in the study. The antibacterial effects of phenolic compounds were screened by well diffusion assay. Pyocyanin and biofilm activity were measured from culture supernatants and the absorbance values were measured using a spectrophotometer. Swarming plates supplemented with phenolic acids were point inoculated with P. aeruginosa strains and the ability to swarm was determined by measuring the distance of swarming from the central inoculation site. Results: Tested phenolic compounds reduced the production of pyocyanin and biofilm formation without affecting growth compared to untreated cultures. Moreover, these compounds blocked about 50% of biofilm production and swarming motility in P. aeruginosa isolates. Conclusions: We may suggest that if swarming and consecutive biofilm formation could be inhibited by the natural products as shown in our study, the bacteria could not attach to the surfaces and produce chronic infections. Antimicrobials and natural products could be combined and the dosage of antimicrobials could be reduced to overcome antimicrobial resistance and drug side effects.

706–708

Authors: Mosaab Adl Eldin Omar, Layla Omran Elmajdoub, Mohammad Saleh Al-Aboody, Ahmed Mahmoud Elsify, Ahmed Osman Elkhtam, Abdelnasser A. Hussien

Objective: To present the molecular characterization of Cysticercus tenuicollis (C. tenuicollis) of Taenia hydatigena (T. hydatigena) from livestock isolates in Egypt, and to introduce a detailed image ofC. tenuicollis infection in ruminant animals in Upper Egypt. Methods: The prevalence rates of C. tenuicollis infections among the slaughtered animals from different organs were determined using the amplification of sequencing of the MT-CO1 gene. Results: In the present study the infection rates of C. tenuicollis were found to be 16% and 19% in sheep and goat samples respectively. Firstly we report one larval stage of T. hydatigena detected in the camel liver in Egypt. C. tenuicollis infection manifested a higher prevalence in females than in males. Those above two years of age manifested a higher infection rate than younger animals. The preferred site for the infection was the omentum: a 70% preference in sheep and a 68% preference in goats. The molecular characterization using the MT-CO1 gene of isolates from sheep, goats and camels corresponded to T. hydatigena. For this study, molecular characterizations of T. hydatigena were done for the first time in Egypt. Molecular tools are of great assistance in characterizing the C. tenuicollis parasite especially when the morphological character cannot be detected, because the metacestodes are frequently confused with infection by the hydatid cyst, especially when these occur in the visceral organs. In the present study, C. tenuicollis manifested high identity in the goat and sheep samples, while differences were found more frequently in the camel samples (10 base pair). Conclusions: Clearly molecular diagnosis for C. tenuicollis infection significantly helps to differentiate it from such other metacestodes as hydatidosis, which manifests a completely different pathogenicity and requires different control programs.

720–726

Authors: Amina Hoceini, Nihel Klouche Khelil, Ilhem Ben-Yelles, Amine Mesli, Sara Ziouani, Lotfi Ghellai, Nadia Aissaoui, Fatima Nas, Mounia Arab

Objective: To compare oral hygiene practices, education and social background, food intake and oral malodor of Algerian adults suffering from dental caries with normal controls, and to determine and compare the bacterial composition of the supragingival plaques from the above-mentioned groups. Methods: Participants completed a questionnaire and were clinically examined for dental caries using decayed, missing and filled teeth index according to the criteria laid down by the World Health Organization. Supragingival plaque samples were collected from 50 caries-free adults (CF) and 50 caries-active adults (CA). Standard procedures of culture and identification of aerobic and anaerobic bacteria were used. Data were analyzed using Chi-square test. Results: A total of 117 bacterial strains were isolated from supragingival plaques in CF group subjects, 76 (64.96%) of them belonged to 9 aerobic genera, and 41 (35.04%) to 9 anaerobic genera (P < 0.05). While in the second group, 199 strains were isolated, 119 (59.80%) of the strains belonged to 10 aerobic genera and 80 (40.20%) to 10 anaerobic bacteria (P < 0.05). Streptococcus mutans, Enterococcus faecium, Aerococcus viridans, Actinomyces meyeri, Lactobacillus acidophilus and Eubacterium limosum showed a significantly higher prevalence in the CA group (P < 0.05). The findings revealed that CA group had a high sugar intake (80%). A significantly higher frequency of tooth brushing (P < 0.000) and a significantly less self-reported oral malodor (P < 0.000) and tooth pain (P < 0.000) were found in CF group, while there was no association of socioeconomic levels and intake of meal snacks with dental caries. Conclusions: This study confirms the association of some aciduric bacteria with caries formation, and a direct association of sugar intake and cultural level with dental caries. Furthermore, oral hygiene practices minimize the prevalence of tooth decay.

554–561

Authors: Assalif Demissew, Meshesha Balkew, Melaku Girma

Objective: To elucidate the larvicidal potency of neem, chinaberry and Bacillus thuringiensis israelensis (Bti) to larvae of Anopheles arabiensis under semi-field condition and adult susceptibility/resistance to the conventionally used insecticides in Tolay, Southwestern Ethiopia. Methods: Wild collected 3rd and 4th stage larvae were exposed to neem, and chinaberry seed powder dissolved in water and Bti in artificial containers at three treatment levels: 0.2, 0.1 and 0.05 g/m2 and controls were free of treatments. Larval and pupal mortalities were monitored daily and residual activities were determined. The experiments were replicated three times. The World Health Organization tube test for all classes of insecticides was conducted on adult Anopheles arabiensis reared from field collected larvae and pupae. Data were analyzed using STATA software version 11. Results: In the first application, neem powder caused 88.9%, 87.9% and 79.4% larval and pupal mortality at 0.2, 0.1 and 0.05 g/m2 after 4.3, 6.0 and 5.7 days, respectively. The corresponding killing effect of chinaberry was 80.3%, 62.1% and 30.3% after 7.0, 7.7 and 8.3 days respectively. Bti at all treatments killed 100% after 24 h except 2.7 days for 0.05 g/m2. Adult mosquitoes were susceptible only for fenitrothion and pirimiphos-methyl with 100% mortality while resistant to deltamethrin, alpha-cypermethrin, etofenprox and dichlorodiphenyl-tricgloroethane with only 9.0%, 3.0%, 5.1% and 2.0% mortalities respectively. Conclusions: Neem, chinaberry and Bti showed potent larvicidal and pupicidal activities. However, in the area, high level of mosquito resistance to pyrethroids and dichlorodiphenyl-tricgloroethane was seen which will pose serious challenge to vector control in the future. Therefore, using integrated approach including these botanical larvicides is warranted to manage insecticide resistance.

568–573

Authors: Md. Moklesur Rahman Sarker, Long Chiau Ming, Md. Zaidul Islam Sarker, M Shahabuddin Kabir Choudhuri

Objective: To assess the immunopotentiality of Ayurvedic polyherbal preparations, “Saribadi” and “Anantamul Salsa”. Methods: Freshly prepared BALB/c mice splenocytes were cultured with “Saribadi” or “Anantamul Salsa” treatment [doses of 0.25%, 0.50%, 0.75%, 1.00%, 1.50%, 2.00%, 3.00% and 4.00% (v/v)] at 37
C for 5 days. The immunoglobulin M (IgM) production and lymphocytes proliferation were determined by ELISA and MTT methods, respectively. Endotoxin contamination was assessed by treating the preparations with polymyxin B. Results: The doses of “Saribadi” [0.25%, 0.50%, 0.75% and 1.00% (v/v)] significantly increased IgM productions (0.966, 0.728, 0.695 and 0.615 mg/mL vs. control 0.265 mg/mL) and lymphocytes proliferation [absorbance 0.311, 0.394, 0.372 and 0.334 optical density (OD) vs. control 0.162 OD]. Similarly, the doses of “Anantamul Salsa” [0.50%, 0.75%, 1.00% and 1.50% (v/v)] promoted IgM productions (0.933, 0.919, 0.917 and 0.892 mg/mL vs. control 0.502 mg/mL) and the doses of “Anantamul Salsa” [0.50%, 0.75%, 1.00%, 1.50%, 2.00%, and 3.00% (v/v)] stimulated lymphocytes proliferation (absorbance 0.395, 0.326, 0.440, 0.398, 0.452 and 0.355 OD vs. control 0.199 OD). The activity of “Saribadi” and “Anantamul Salsa” was not retarded by the treatment of preparations with polymyxin B. Conclusions: Immunomodulatory activity of “Saribadi” and “Anantamul Salsa” was unveiled for the first time. “Saribadi” and “Anantamul Salsa” possess immunostimulating potential acting through the induction of lymphocyte proliferation and IgM production. These preparations may be useful in strengthening immune responses. However, further cellular and in vivo studies are required.

580–585

Authors: Joo Hyun Nam, Hyo Won Jung, Young-Won Chin, Woo Kyung Kim, Hyo Sang Bae

Objective: To examine the effects of Tribulus terrestris L. (T. terrestris) extract on the modulation of calcium channels to evaluate its use in topical agents for treatment of atopic dermatitis. Methods: The 70% methanol extract of T. terrestris was prepared. Human HEK293T cells with over-expressed calcium release-activated calcium channel protein 1 (Orai1), transient receptor potential vanilloid 1, or transient receptor potential vanilloid 3 (TRPV3) were treated with T. terrestris extract. Modulation of ion channels was measured using a conventional whole-cell patch-clamp technique. Results: T. terrestris extract (100 mg/mL) significantly inhibited Orai1 activity in Orai1- stromal interaction molecule 1 co-overexpressed HEK293T cells. In addition, T. terrestris extract significantly increased the TRPV3 activity compared with 2-Aminoethyl diphenylborinate (100 mmol/L), which induces the full activation of TRPV3. Conclusions: Our results suggest that T. terrestris extract may have a therapeutic potential for recovery of abnormal skin barrier pathologies in atopic dermatitis through modulating the activities of calcium ion channels, Orai1 and TRPV3. This is the first study to report the modulatory effect of a medicinal plant on the function of ion channels in skin barrier.

597–604

Authors: Ernieenor Faraliana Che Lah, Salmah Yaakop, Mariana Ahamad, Ernna George, Shukor Md Nor

Objective: To identify different stages of Ixodes granulatus (I. granulatus) based on morphological characters prior to molecular identification which is significant for confirming and identifying the nymphal stages of I. granulatus. Methods: A total of 14 individuals of adult, engorged and nymphal ticks collected from three different localities were examined morphologically using taxonomic keys, followed by PCR using cytochrome oxidase subunit I (COI). Clustering analysis based on COI sequences was carried out by constructing neighbor-joining and maximum parsimony tree to clarify the genetic variation and diversity of local I. granulatus. Results: Based on external morphological characterizations, nine individuals (64.3%) were successfully identified as I. granulatus, while five individuals were recognized only as Ixodes sp. due to lack of morphological characters visible and development during that stage. Molecular analysis of local I. granulatus using COI gene revealed 93%–94% sequence homology from available sequence in GenBank and was in concordance with the morphological identification. Furthermore, a low intraspecific variation was observed among the species of I. granulatus collected from different localities (0%–3.7%). Conclusions: These findings demonstrated for the first time the establishment of COI gene for identifying I. granulatus nymphal tick which is of paramount importance to the control of potential tick-borne infections in Malaysia. Moreover, this study provides evidence that a combination of morphology and molecular data was corroborated as an accurate tool for tick identification.

609–613

Authors: Ismail Jalil, Muhammad Arshad, Zara Rafaque, Fazle Raziq, Robina Wazir, Sajid Malik, Javid Iqbal Dasti

Objective: To study the seroprevalence of hepatitis B virus (HBV) and hepatitis delta virus (HDV) infections in patients visiting outpatient department of a major tertiary care hospital in Khyber Pakhtunkhwa region of Pakistan. Methods: Blood samples were collected from non-hospitalized patients. Serological analysis was done by ELISA and viral DNA was amplified by PCR. The amplified DNA was analyzed by agarose gel electrophoresis. Results: Altogether, 946 blood samples were screened, overall percentage of HBsAgpositive patients remained 22.41% (prevalence: 224.10/1 000; CI: 0.197 5 ± 0.250 7) with the highest incidence rates among relatively younger age groups (20–29 years). The prevalence of HBV–HDV co-infection was found to be 46.75/1 000; CI: 0.031 8 ± 0.061 7. In HBsAg-positive patients, anti-HBc-total was detected in 86.79% while 25.00% were positive for anti-HBc-immunoglobulin M. Similarly, among these patients, HBV DNA was detected in 64.13% and 10.85% were co-infected with HDV. Different symptoms were associated with the prevailing infection, including malaise (62%), anorexia (66%) and fatigue (73%). The most commonly associated symptom was abdominal discomfort. Among these patients, certain risk factors, including surgery, visit to dentist and intravenus infusions were frequently associated with the infection (c2 = 95.23; df= 11; P < 0.000 1). Conclusions: Overall, this study confirmed higher prevalence of active HBV/HDV infection, among young patients from Khyber Pakhtunkhwa region having no prior history of viral hepatitis.

620–624

Authors: Freddie Masaninga, Mary Katepa Bwalya, Sarai Malumo, Busiku Hamainza, Peter Songolo, Mulakwa Kamuliwo, Martin Meremikwu, Lawrence Kazembe, Jacob Mufunda, Olusegun Ayorinde Babaniyi

Objective: To assess potential determinants of uptake and highlight lessons learnt from the implementation of intermittent preventive treatment (IPTp), given to pregnant women as early as possible during the second trimester in Zambia. Methods: Data from four national malaria surveys (2006, 2008, 2010, 2012) were reviewed, and proportions of pregnant women attending antenatal clinics (ANCs) who received two or more doses of sulfadoxine–pyrimethamine (IPTp2) were compared by place of residence, education level, and wealth status. Malaria cases and deaths in pregnant women, from Health Information Management System 2011–2013, were analyzed to determine malaria burden in pregnancy in Zambia. A multiple logistic regression model was applied to identify potential determinants of IPTp uptake. Results: The proportion of pregnant women who took IPTp at ANCs increased from near zero at inception in 2001 to 61.9% in 2006; and to 72% by 2012 (P < 0.001), and overall the uptake was 1.41 times higher in 2012 compared to 2006. From 2006 to 2012, IPTp2 uptake among women with no formal education increased from 51% to 68% (P < 0.1). Likewise, uptake among pregnant women with the lowest wealth index increased from 58.2% to 61.2%. By 2012, IPTp uptake among pregnant women within the lowest wealth index increased to a similar level as the women with high wealth index (P = 0.05). Incidence of malaria cases, hospital admissions and mortality during pregnancy decreased between 2011 and 2013. Overall, increased IPTp uptake was associated with being in urban areas (OR = 1.56, 95% CI: 1.39–1.74), having college (OR = 1.83, 95% CI: 1.25–2.75) or secondary education (OR = 1.68, 95% CI: 1.44–1.96) or of being of higher wealth status (OR = 1.86, 95% CI: 1.60–2.17). Conclusions: Zambia has increased IPTp uptake through ANC for all women. The malaria control program has contributed to increasing access to health services and reducing demographic and socioeconomic disparities.

630–632

Authors: Blerina Resuli, Roberto Lisi, Daniela Musio, Vincenzo Tombolini

On September 2013, a 62-year-old man with metastatic adenocarcinoma of the lung complained tenderness and pain of the first terminal phalange of his right hand. The biopsy confirmed metastatic adenocarcinoma of the lung to the finger. A single 8-Gy fraction of palliative radiotherapy was delivered to the patient's right hand. The patient received magnetic resonance-guided focused ultrasound surgery treatment to the phalange because he showed few improvement of clinical symptoms and persistence of moderate pain after radiotherapy. After magnetic resonance-guided focused ultrasound surgery, the clinical symptoms improved significantly. No serious adverse effects were reported and the patient compliance was very high. Our patient showed improvement of clinical symptoms after combined treatment. The patient remains in good health conditions.

468–471

Authors: Giovanni Benelli

Mosquitoes (Diptera: Culicidae) represent a key threat for millions of humans and animals worldwide, since they act as vectors for important parasites and pathogens, including malaria, filariasis and a wide number of arboviruses. The recent outbreaks of Zika virus infections occurring in South America, Central America, and the Caribbean, represent the most recent four arrivals of important arboviruses in the western hemisphere, over the last 20 years, namely dengue, West Nile virus, and chikungunya. Since there are no specific treatments for Zika virus and the other arboviruses mentioned above, it should be highlighted that the eco-friendly and effective control of mosquito vectors is of pivotal importance. Besides radiation, transgenic and symbiont-based mosquito control approaches, an effective option may be the employ of biological control agents of mosquito young instars, in presence of ultra-low quantities of green-synthesized nanoparticles, which magnify their predation efficiency. Furthermore, behaviour-based control tools relying on the employ of swarming behaviour manipulation (i.e. the “lure and kill” approach), pheromone traps, sound traps need further research attention. In particular, detailed basic information on the physical and chemical cues routing mosquito swarming and mating dynamics is urgently required.

477–484

Authors: Robert Soumay Houmsou, Sam Mao Panda, Sambo Obadiah Elkanah, Liatu Columbus Garba, Binga Emmanuel Wama, Elizabeth Une Amuta, Santaya Larit Kela

Objective: To determine schistosomiasis level and risk factors that exposed school-aged children to infection as well as to model schistosomiasis map in relation to altitude and rainfall in Gashaka Local Government Area, Taraba State, Nigeria. Methods: The study was conducted between January 2014 and June 2014. Urine and faecal samples were collected from 1 080 school-aged children and processed using the filtration and formol–ether concentration techniques, respectively. Results: Irrespective of the schistosomes species, a point prevalence of 10.18% was reported out of the 1 080 children examined. Males were significantly infected than their female counterparts (11.75% vs. 8.43%) (c2 = 4.86; P = 0.027), as well as children aged 6–10 (11.65%, 72/618) and 11–15 years (10.29%, 35/340) than the other age groups (c2 = 9.274; P = 0.026). No significant difference was observed in schistosomiasis between children whose parents were educated (11.11%, 57/513) and not educated (9.88%, 53/536) (c2 = 1.342; P = 0.247) and those whose parents are farmers (9.74%, 53/544) and non-farmers (10.63%, 57/536) (c2 = 0.787; P = 0.375). Proximity to water bodies (distance < 500 m) (odds ratio = 1.809, confidence interval = 1.057–3.094; P = 0.003 1) and fishing (odds ratio = 2.632, confidence interval = 1.397–4.958; P = 0.003) were the risk factors exposing children to infection. The spatial distribution pattern of schistosomiasis showed that the infection was significantly higher in Serti A (22.2%, 26/180) and Mayo-Selbe (21.1%, 38/180) than the other localities (c2 = 92.99; P = 0.000). Conclusions: This study reported a moderate level of infection among school-aged children with proximity to water bodies and fishing as the main risk factors. The spatial distribution of schistosomiasis in the area will guide in efficient and effective control programmes at local level. It is recommended that continued efforts be made to scale-up distribution of praziquantel to high risk areas so as to curb the progression of the disease.

492–500

Authors: Jugjeet Singh Ramkissoon, Mohamad Fawzi Mahomoodally, Anwar Hussein Subratty, Nessar Ahmed

Objective: To investigate the inhibitory activity of ten culinary herbs and spices namely on glucose-mediated glycation (GMG) and fructose-mediated glycation (FMG) of bovine serum albumin. Methods: Fluorescence was used as an index of albumin glycation using glucose and fructose as substrates in the presence of infusions and ethanolic extracts of ten culinary herbs and spices. Antioxidant activity of the extracts was evaluated using reducing power, metal ion chelating and superoxide radical scavenging assays. Phytochemicals profile was analysed using 13 standard methods. Results: FMG was found to be significantly higher than GMG (95 and 84 AU, respectively; P < 0.05). Infusions and ethanolic extracts showed significant (P < 0.05) inhibitory activity on both GMG and FMG when compared to appropriate controls. No significant difference (P > 0.05) was found in the percentage glycation inhibitory activity of infusions compared to ethanolic extracts. The mean percentage inhibitory activity of the extracts for GMG (45.9%) and for FMG (45.1%) was not significantly different (P > 0.05). Qualitative phytochemical analysis showed the presence of alkaloids, flavonoids, tannins, terpenoids, anthraquinones, steroids, reducing sugars, proteins, phenols, saponins, phlobatannins, and cardiac glycosides. Conclusions: The higher rate of fluorescence generation by fructation suggests that glycation by fructose deserves much attention as a glycating agent. Data herein showed that the extracts inhibited GMG and FMG. Thus, these edible plants could be a natural source of antioxidants and anti-glycation agent for preventing advanced glycation endproducts-mediated complications.

506–510

Authors: Noreen Orianna Koumba Madingou, Aristide Traore, Alain Souza, Marlaine Michelle Boukandou Mounang1, Raissa Reine Aworet Samseny, Sylvin Ouedraogo, Alfred Sababenedyo Traore

Objective: To investigate the toxicity of aqueous extract of Guibourtia tessmannii (Harms) J. Leonard (G. tessmannii) and evaluate its safety. Methods: NMRI mice were used to determine the acute toxicity of G. tessmannii. Increasing concentrations of the plant extracts were administered intraperitoneally or by force-feeding. General behavior and death were monitored and recorded daily for 7 days. In order to determine the sub-acute toxicity of the extract, several doses were administered by oral gavage daily for 28 days in adult Wistar rats. Different parameters were assessed including body weight, food and water intake, biochemical parameters and several vital organ weights. Results: LD50 of 328.78 mg/kg was obtained by i.p. route and more than 5 000 mg/kg was obtained in acute toxicity by oral route. In sub-acute toxicity, no significant alteration was observed in body weight or vital organs, food and water intake, and biochemical parameters. Conclusions: The results showed that the aqueous extract of G. tessmannii has low toxicity intraperitoneally and no sub-acute toxicity via oral intake.

516–519

Authors: Dolores Limongi, Cartesio D'Agostini, Marco Ciotti

Sepsis remains a leading cause of death in the intensive care units and in all age groups worldwide. Early recognition and diagnosis are key to achieving improved outcomes. Therefore, novel biomarkers that might better inform clinicians treating such patients are surely needed. The main attributes of successful biomarkers would be high sensitivity, specificity, possibility of bedside monitoring and financial accessibility. A panel of sepsis biomarkers along with currently used laboratory tests will facilitate earlier diagnosis, timely treatment and improved outcome may be more effective than single biomarkers. In this review, we summarize the most recent advances on sepsis biomarkers evaluated in clinical and experimental studies.

528–533

Authors: Ebrahim Babapour, Azam Haddadi, Reza Mirnejad, Seyed-Abdolhamid Angaji, Nour Amirmozafari

Objective: To check biofilm formation by Acinetobacter baumannii (A. baumannii) clinical isolates and show their susceptibility to different antibiotics and investigate a possible link between establishment of biofilm and multidrug resistance. Methods: This study was performed on clinical samples collected from patients with nosocomial infections in three hospitals of Tehran. Samples were initially screened by culture and biochemical tests for the presence of different species of Acinetobacter. Identifications were further confirmed by PCR assays. Their susceptibilities to 11 antibiotics of different classes were determined by disc diffusion method according to Clinical and Laboratory Standards Institute guidelines. The ability to produce biofilm was investigated using methods: culture on Congo red agar, microtiter plate, and test tube method. Results: From the overall clinical samples, 156 specimens were confirmed to contain A. baumannii. The bacteria were highly resistant to most antibiotics except polymyxin B. Of these isolates, 10.26% were able to produce biofilms as shown on Congo red agar. However, the percentage of bacteria with positive biofilm in test tube, standard microtiter plate, and modified microtiter plate assays were 48.72%, 66.66%, and 73.72%, respectively. At least 92% of the biofilm forming isolates were multidrug resistant. Conclusions: Since most of the multidrug resistant strains produce biofilm, it seems necessary to provide continuous monitoring and determination of antibiotic susceptibility of clinical A. baumannii. This would help to select the most appropriate antibiotic for treatment.

539–545

Authors: Sothorn Anucherngchai, Thanawan Tejangkura, Thapana Chontananarth

Objective: To investigate the prevalence of cercarial trematode infection in snails and to examine the reconstruction of the phylogenetic relationship to explain the molecular system of cercarial stage trematodes to estimate the infection rate of in the definite host from the Chao-Phraya Basin. Methods: The snails were collected from 10 provinces of the Chao-Phraya Basin, Thailand by stratified sampling method. The snails were examined for cercarial infection by the crushing method. All DNA specimens were amplified with internal transcribed spacer 3 (ITS3) and ITS4 primer based on PCR technique. The sequence data were aligned and used to reconstruct the phylogenetic tree by unweighted pair-group method with arithmetic means with 10 000 bootstraps. Results: The overall rate of cercarial infection was found to be 5.90% (122/2 067). Snails in the family Thiaridae were found to be in the highest prevalence followed by Lymnaeidae, Bithyniidae, Planorbidae, Viviparidae, and Ampullariidae, respectively, while the Buccinidae family (Clea helena) did not reveal any infections. The frequently found species of cercariae were parapleurolophocercous cercariae, cercariae and megarulous cercariae. The monophyletic tree separated the snails into five groups comprised of Heterophyidae, Strigeidae, Lecithodendriidae, Philophthalmidae and Echinostomatidae using the sequence of Angiostrongylus cantonensis as an out-group. Conclusions: This study was the first to report on cercarial infection in the Chao-Phraya Basin, Thailand. This revealed that a high variety of freshwater snails were infected by cercariae stage trematodes with a high prevalence. The sequence data of ITS2 can be used to investigate the phylogenetic relationships of trematodes at the family level and in each clade of different families separated by the definitive hosts.

376-381

Authors: Lawrence N. Kazembe, Don P. Mathanga

Objective: To estimate risk factors of urban malaria in Blantyre, Malawi, with the goal of understanding the epidemiology and ecology of the disease, and informing malaria elimination policies for African urban cities that have markedly low prevalence of malaria. Methods: We used a case-control study design, with cases being children under the age of five years diagnosed with malaria, and matched controls obtained at hospital and communities. The data were obtained from Ndirande health facility catchment area. We then fitted a multivariate spatial logistic model of malaria risk. Covariate and risk factors in the model included child-specific, household and environmental risk factor (nearness to garden, standing water, river and swamps). The spatial component was assumed to follow a Gaussian process and model fitted using Bayesian inference. Results: Our findings showed that children who visited rural areas were 6 times more likely to have malaria than those who did not [odds ratio (OR) = 6.66, 95% confidence interval (CI): 4.79–9.61]. The risk of malaria increased with age of the child (OR = 1.01, 95% CI: 1.003–1.020), but reduced with high socio-economic status compared to lower status (OR = 0.39, 95% CI: 0.25–0.54 for the highest level and OR = 0.67, 95% CI: 0.47– 0.94 for the medium level). Although nearness to a garden, river and standing water showed increased risk, these effects were not significant. Furthermore, significant spatial clusters of risk emerged, which does suggest other factors do explain malaria risk variability apart from those established above. Conclusions: As malaria in urban areas is highly fuelled by rural-urban migration, emphasis should be to optimize information, education and communication prevention strategies, particularly targeting children from lower socio-economic position.

384-389

Authors: Luisa De Martino, Francesca Paola Nocera, Karina Mallardo, Sandra Nizza, Eleonora Masturzo, Filomena Fiorito, Giuseppe Iovane, Piergiorgio Catalanotti

Objective: To update the recent knowledge of the microbiological causes of canine otitis externa in Campania Region (Italy) and the antibiotic susceptibility patterns of the isolated strains. Methods: A total of 122 dogs were examined by otoscopy, and auricular swab samples were collected from both ears in 74 dogs presenting clinical bilateral otitis and from single ears in 48 dogs displaying clinical unilateral otitis. Cytological examination, bacteriological analysis and antimicrobial susceptibility tests were performed. Results: Thirty-one out of 122 dogs were positive for yeast species (25.4%, 95% confidence interval (CI): 18.2%–34.2%) with a higher prevalence of Malassezia pachydermatis (21/31 isolates, 67.7%, CI: 48.5%–82.7%), and a total of 91 out of 122 dogs were positive for bacterial species (74.6%; CI: 65.8%–81.8%) with a higher prevalence of Staphylococcus pseudintermedius (45/143 isolates, 31.5%, CI: 24.1%–39.8%). These results are the first description of Streptococcus agalactiae-associated otitis. The yeasts isolated showed high levels of susceptibility to all antifungal agents tested; on the contrary all the isolated bacterial strains were highly resistant to at least four out of ten antimicrobial classes. Both Gram-positive and Gram-negative bacteria showed high resistance to amoxicillin/clavulanate and kanamycin hence they are not recommended as initial empirical therapy for the otitis treatment. Conclusions: This update illustrates an increase in antibiotic resistances providing an insight into the current knowledge of the therapeutic procedures followed on canine otitis externa in Italy. It also emphasizes the importance of considering the results of the microbiological and sensitivity tests to decide on an appropriate antibiotic therapy.

396-399

Authors: Louise Helena Guimarães de Oliveira, Patricia Alexandria Paiva Silva de Sousa, Fabrine Felipe Hilario, Gabriel Joventino Nascimento, João Paulo Saraiva Morais, Everaldo Paulo de Medeiros, Manoel Francisco de Sousa, Fabiola da Cruz Nunes

Objective: To investigate the effects of Agave sisalana (A. sisalana) extract on Aedes aegypti (Ae. aegypti) primary cell culture. Methods: Cells of Ae. aegypti were exposed to different concentrations of A. sisalana crude extract (0.18–6.00 mg/mL) for 24 h. Then, the cells were labeled with propidium iodide and subjected to fluorescence microscopy to verify cell viability. In addition, nitric oxide production was measured. Results: Results showed that cells exposed to 6 mg/mL of the crude extract presented a greater percentage of death when compared to control (73.8% ± 9.6% vs. 34.6% ± 9.6%). Furthermore, there was an increase in the nitric oxide production in cells exposed to 6 mg/ mL of A. sisalana crude extract [(0.81 ± 0.08) mmol/L] compared to control group [(0.41 ± 0.18) mmol/L]. Conclusions: The results show that A. sisalana is cytotoxic to Ae. aegypti and may be used as raw material for new eco-friendly and inexpensive insecticides, since sisal industry discards the liquid waste for the extraction of plant fiber.

404-409

Authors: Latifah Saiful Yazan, Napsiah Abd Rahman, Kim Wei Chan, Wan Nor Hafiza Wan Abd Ghani, Yin Sim Tor, Jhi Biau Foo

Objectives: To determine the cytotoxicity of crude ethanolic extract, n-butanol fraction and aqueous fraction on selected cancer cell lines, and to observe the morphological changes of the cancer cells treated with n-butanol fraction. Methods: The cytotoxic effect of n-butanol fraction, crude ethanolic extract and aqueous fraction on breast cancer (MCF-7 and MDA-MB-231), colon cancer (HT29), lung cancer (A549), cervical cancer (HeLa) and normal mouse fibroblast (3T3) cell lines was determined using MTT assay. The morphological changes of the treated cells were observed under an inverted light microscope. Results: n-Butanol fraction was the most cytotoxic towards HT29 and MCF-7 cells in a dose-dependent manner compared to crude ethanolic extract and aqueous fraction (P < 0.05). The IC50 of n-butanol fraction for HT29 and MCF-7 was (780.00 ± 28.28) and (895.00 ± 7.07) mg/mL, respectively. Cell shrinkage, membrane blebbing and formation of apoptotic bodies were noted following treatment of HT29 cells with n-butanol fraction. Conclusions: In conclusion, n-butanol fraction was more cytotoxic than crude ethanolic extract and aqueous fraction towards the selected cancerous cell lines and induced apoptosis in HT29 cells. 1. Introduction Kenaf (Hibiscus cannabinus L.) belongs to the Malvaceae (Mallow)

418-421

Authors: Henry Borb´on, Sandra V´aldes, Javier Alvarado-Mes´en, Roy Soto, Ilena Vega

Objective: To evaluate the antimicrobial and antifungal activities of the aqueous and partitioned extract of sea anemone Anthopleura nigrescens (A. nigrescens). Methods: The sea anemone A. nigrescens was collected, minced, homogenized, lyophilized and then further partitioned with diethyl ether, acetone, ethanol and water. These fractions were evaluated for antimicrobial activity against bacterial and fungal pathogens. Results: Acetone extract was found to produce a pronounced inhibition of 7.0 mm against Proteus vulgaris and diethyl ether extract inhibited Pseudomonas aeruginosa with an inhibition zone of 6.5 mm. In antifungal activity, ethanol extract showed good activity against Botrytis cinerea, Trichoderma harzianum and Rhizopus oryzae compared with other strains. Acetone and ethanol extract of A. nigrescens showed activity against all of pathogens tested. Slight activity was observed in the water extract with inhibition zone of 1.5 mm. Conclusions: The present study revealed that sea anemone A. nigrescens may also contain some biologically active agents which have potential activity against pathogenic microorganisms.

429-440

Authors: Khaled Mohamed Mohamed Koriem

Multiple sclerosis (MS) is the most famous autoimmune disease attacking the central nervous system. It attacks people from age 20–50 years old and the females' attacks double than males' attacks. MS is an autoimmune disease affecting principally the central nervous system that cause nerve sheath demyelination followed by axon damage and paralysis. MS symptoms include muscle weakness, weak reflexes, muscle spasm, difficult in move, miss-coordination and unbalance with others. There are many factors may be responsible for MS: microbial, viral, smoking, stress, environmental toxins, contaminated diet, and gout. MS is wide spread in the populations in North Europe and this related to lack of vitamin D due to decrease of sunlight exposure. MS biomarkers include nitric oxide, interleukin-6, nitric oxide synthase, fetuin-A and osteopontin. MS is not a genetic disease where MS occurs when human leukocyte antigen system related genes are changed in chromosome 6. The physiology of MS is monitored by activation of immuneinflammatory, oxidative, and nitrosative stress pathways. MS is including two main steps: (1) myelin sheath destruction and formation of lesions and, (2) inflammation. Four types of MS can be distinguished: relapsing-remitting, primary progressive, secondary progressive and progressive relapsing. Nine treatments have been accepted for relapsingremitting MS type: interferon b-1a, interferon b-1b, mitoxantrone, natalizumab, glatiramer acetate, fingolimod, dimethyl fumarate, teriflunomide, and alemtuzumab, however, the only treatment used is mitoxantrone for progressive MS but many of MS treatments side effects are recorded. Complementary treatments also used in MS treatments such as: vitamin D, Yoga, medicinal plants, oxygen therapy, acupuncture and reflexology.

443-445

Authors: Mahdieh Rezaei, Shahrzad Azizi, Shima Shahheidaripour, Sara Rostami

Transmissible venereal tumor (TVT) is a coitally transmitted tumor of dogs with widespread distribution. The present study describes the occurrence of the primary oral and nasal TVT in a 10-year-old, female, mix-breed dog. The case was presented with a history of anorexia, inability to swallow and dyspnea. Clinical examinations revealed the emaciation, muzzle deformity due to the presence of a friable, fleshy, cauliflower-like mass in the oral cavity and submandibular lymphadenopathy. TVT was diagnosed based on histopathological findings. The dog was discharged with therapeutic intervention with vincristine. Unfortunately, the case died before readmission because of the progressive worsening of the general condition. Our findings highlight the need for considering TVT for the differential diagnosis of the extragenital masses in dogs.

451-454

Authors: Kazem Baesi, Majedeh Moradbeigi, Mehrdad Ravanshad, Ashrafolnesa Baghban

Objective: To survey the level and patterns of reverse transcriptase-based drug resistance and subtype distribution among antiretroviral-treated HIV-infected patients receiving only reverse transcriptase inhibitors in Iran. Methods: A total of 25 samples of antiretroviral therapy experienced patients with no history of using protease inhibitors were collected. After RNA extraction, reverse transcriptase-nested PCR was performed. The final products were sequenced and then analysed for drug-resistant mutations and subtypes. Results: No drug resistant mutations were observed among the 25 subjects. The results showed the following subtypes among patients: CRF 35_AD (88%), CRF 28_BF (8%), and CRF 29_BF (4%). Conclusions: A significant increase in drug resistance has been noted in recentlyinfected patients worldwide. Subtype distributions are needed to perform properlydesigned surveillance studies to continuously monitor rates and patterns of transmitted drug resistance and subtypes to help guide therapeutic approaches and limit transmission of these variants.