Asian Pacific Journal of Tropical Medicine

339-346

Authors: Sravani Karri, Sanjay Sharma, Ginpreet Kaur

Malaria remains a major tropical health burden owing to the development of resistance and decreased sensitivity to the frequently used conventional antimalarial drugs. The drug like artemisinin possesses potent antimalarial activities, but has some limitations. Therefore, new strategies are to be implemented for optimal utilization of artemisinin to improve its therapeutic effectiveness and to overcome its limitations. The present review focuses on present scenario of malaria and pharmacological as well as analytical aspects of artemisinin. Data from 2000 to 2018 were collected from NCBI for understanding the various analytical techniques used for estimation of artemisinin. This review will reveal the facts about artemisinin which can be utilized to develop novel drug delivery system either in a combination or as alone for the wellbeing of the patients suffering from malaria.

353-364

Authors: Emrah Sefik Abamor, Adil Allahverdiyev, Ozlem Ayse Tosyali, Melahat Bagirova, Tayfun Acar, Zeynep Mustafaeva, Serap Derman

Objective: To prepare and characterize poly lactic-co-glycolic acid (PLGA) nanoparticles loaded with soluble leishmanial antigen or autoclaved leishmanial antigen and explore in vitro and in vivo immunogenicity of antigen encapsulated nanoparticles. Methods: Water/oil/water double emulsion technique was employed to synthesize PLGA nanoparticles, and scanning electron microscopy, Fourier transform infrared spectroscopy and Zeta-potential measurements were used to identify the characteristics of nanoparticles. Cytotoxicity of synthetized nanoparticles on J774 macrophage were investigated by MTT assays. To determine the in vitro immunostimulatory efficacies of nanoparticles, griess reaction and ELISA was used to measure the amounts of NO and cytokines. During the in vivo analysis, Balb/c mice were immunized with vaccine formulations, and protective properties of nanoparticles were measured by Leishman Donovan unit in the liver following the infection. Cytokine levels in spleens of mice were determined by ELISA. Results: MTT assay showed that neither soluble leishmanial antigen nor autoclaved leishmanial antigen encapsulated nanoparticles showed cytotoxicity against J774 macrophage cells. Contrary to free antigens, both autoclaved leishmanial antigen-nanoparticle and soluble leishmanial antigen-nanoparticle formulations led to a 10 and 16-fold increase in NO amounts by macrophages, respectively. Leishman Donovan unit calculations revealed that soluble leishmanial antigen-nanoparticles and autoclaved leishmanial antigen-nanoparticles yielded 52% and 64% protection against visceral leishmaniasis in mouse models. Besides, in vitro and in vivo tests demonstrated that by increasing IFN-γ and IL-12 levels and inhibiting IL-4 and IL- 10 secretions, autoclaved leishmanial antigen-nanoparticles and soluble leishmanial antigennanoparticles triggered Th1 immune response. Conclusions: Both autoclaved leishmanial antigen-nanoparticles and soluble leishmanial antigen-nanoparticles formulations provide exceptional in vitro and in vivo immunostimulatory activities. Hence, PLGA-based antigen delivery systems are recommended as potential vaccine candidates against visceral leishmaniasis.

375-379

Authors: Parut Suksai, Supakarn Kaewchot, Piya Sereerak, Sirinan Boonnan, Bongkotchamat Phimsin, Taksina Jaruwattananon, Kacharin Raschasin, Mutchamon Kaewparuehaschai, Sorraya Siriphet, Benjaporn Bhusri

Objective: To survey hemoplasmas infection in free ranging non-human primates from 8 provinces in Thailand. Methods: DNA from ethylenediaminetetraacetic acid blood of 262 free ranging non-human primates were identified as hemoplasmas using PCR and phylogenetic analysis based on 16S rRNA and rnpB genes. Results: A total of 148 non-human primates (56.49%) were determined positive for Candidatus Mycoplasma haemomacaque, including 125 Macaca fascicularis and 23 Macaca mulatta. Hemoplasmas can cause anemia in monkey but all positive samples were healthy. The positive rates in male and female non-human primates were not significantly different. Conclusions: Candidatus Mycoplasma infection is prevalent in free ranging Macaca fascicularis and Macaca mulatta in Thailand.

291-299

Authors: Kiran D. Patil, Shashikant B. Bagade, Sanjay R. Sharma, Ketan V. Hatwar

The WHO reports that billions of people and animals in tropical and subtropical regions are affected by helminthiasis as neglected tropical disease. It is predominant in underdeveloped areas; nevertheless, the increase in the number of travelers and migrants has made this infection more common. The current mass drug treatment produces severe side effects and many strains of helminths are resistant to them. None of the chemotherapeutic drugs meets the ideal requirements of anthelmintics, such as broad spectrum of activity, single dose cure, free from side effect and cost-effectiveness. Today, many researchers are screening the traditional herbal system in search of the anthelmintic herbal constituents which overcome all the problems of synthetic drugs. Several researchers proclaim anthelmintic activity of herbal medicines by using different experimental models. The present review demonstrates natural product drug discovery, outlining potential of herbal constituents from natural sources as natural leads against helminthiasis.

306-314

Authors: Hamed Behniafar1, Vahideh Moin-Vaziri2, Mehdi Mohebali3,4, Seyyed Javad Seyyed Tabaei2, Zabih Zarei5, Elham Kazemirad3, Behnaz Akhoundi4, Mohammad Kazem Saharifi-Yazdi6, Alireza Zahraei-Ramazani

Objective: To clarify the epidemiological aspects of visceral leishmaniasis in Kaleybar and Khoda-Afarin districts, north-west of Iran. Methods: A total of 1 420 human (children under 12 years) samples, 101 domestic dogs samples (Canis familiaris), and 577 female sand fly samples were collected. Sera of human and dogs were tested using the direct agglutination test, and sand flies were identified at species level using the microscopic method. Furthermore, a structured questionnaire was applied to evaluate the correlation between the potential risk factors and the related clinical signs/ symptoms with the human and dogs' seropositivity. Results: Totally, 2.18% of human samples were positive at titers≥1: 800; among them, 13 cases (41.94%) were above 1:3 200, and clinical symptoms were observed in all of them except for an 11-year old girl. Anti-Leishmania infantum antibodies were found at titer≥1: 320 in 9.90% of dogs' samples, half of them had at least one sign of canine visceral leishmaniasis. Moreover, 10 Phlebotomus species were identified in the study areas, and Phlebotomus (Larroussius) major group was the predominant species. There are significant correlations between the presence of anti-Leishmania infantum antibodies and the fever (P<0.001), anemia (P=0.001) and weight loss (P=0.016) in children. On the other hand, significant correlations were revealed between the Leishmania infection and the shelter (P=0.039), cutaneous lesion (P=0.005), lymphadenopathy (P=0.001) and weight loss (P<0.001) in the infected dogs. Conclusions: Visceral Leishmania infection is prevalent in rural areas of Kaleybar and Khoda- Afar districts located in East-Azerbaijan province, therefore active detection and treatment of visceral leishmaniasis cases should not be neglected.

321-328

Authors: Raju Sangeetha, Tamilselvan Pratheeba, Chinnasamy Ragavendran, Devarajan Natarajan

Objective: To test the mosquitocidal potential of leaf extracts of Pouteria campechiana prepared with different solvents and elucidate the structure of an isolated mosquitocidal compound. Methods: The leaf extracts of Pouteria campechiana prepared with three solvents (petroleum benzene, ethyl acetate and acetone) and potential bioactive fractions were tested against various stages of Aedes aegypti and Culex quinquefasciatus by using the WHO protocols, and the chemical profile and its functional groups were identified by GC-MS and Fourier transmission- infrared spectroscopy (FT-IR). The structure of bioactive compound was characterized by nuclear magnetic resonance (NMR) spectral technique. Results: The preliminary phytochemical results revealed the presence of alkaloids, amino acids, flavonoids, quinones, saponins, steroids, tannins, and terpenoids in the acetone extract. A significant toxic potential was observed in the acetone extract against both Aedes aegypti and Culex quinquefasciatus mosquitoes. The acetone extract exhibits remarkable larvicidal (LC50: 12.232 μg/mL and LC90: 63.970 μg/mL), pupicidal (LC50: 18.949 μg/mL and LC90: 167.669 μg/mL) and adulticidal (LC50: 20.689 μg/mL and LC90: 72.881 μg/mL) effects against Aedes aegypti. Furthermore, the same extract was subjected to isolation of bioactive compound by GC- MS and FT-IR analysis. GC-MS results showed the presence of 5 major compounds, and octacosane (18.440%) was detected as the predominant compound. The FT-IR result of acetone extract demonstrated the presence of various functional groups like alkanes/alkynes, ester, aromatic and amides. The NMR spectrum results of isolated compound were well matched to glycoside linked flavonoids. Based on the chromatography and spectral techniques the isolate molecule was identified as myricitrin by FT-IR and nuclear magnetic resonance spectral data. Conclusion: The isolated compound myricitrin possesses a significant toxic effect in all stages of Aedes aegypti and Culex quinquefasciatus mosquito’s with lowest LC50 and LC90 values.

337-338

Authors: Ravindra Kumar Garg, Neeraj Kumar, Imran Rizvi, Hardeep Singh Malhotra

Rationale: Disseminated cysticercosis is characterized by presence of cysts in multiple body organs, like brain, skin, eyes, muscles and rarely heart and lungs. Patient concerns: A 22-year-old man presented with bilateral proptosis of 1-year duration. He also had two episodes of cysticercosis encephalitis. In the second episode of encephalopathy, the patient died. Diagnosis: Disseminated cysticercosis. Interventions: Corticosteroids (Initially intravenous dexamethasone 0.4 mg/kg/day for 2 weeks, followed by oral prednisolone 1.0 mg/kg/day). Outcomes: The patient died of cysticercosis encephalitis approximately 2 months later. Lessons: Disseminated cysticercosis in our case presented only with proptosis as he had very heavy infestation of the brain and eyes. Heavy larval infestation in a patient with disseminated cysticercosis can be life-threatening.

243-249

Authors: Amira Taman, Samar N. El-Beshbishi

Schistosomiasis is a neglected tropical disease that affects about 290 million patients worldwide. Children aged between 5 and 14 years represent 45.8% of the affected patients, in addition, schistosomiasis has been reported in Schistosoma-free areas, mostly because of tourism and immigration from endemic countries. Intestinal schistosomiasis caused by Schistosoma mansoni is mainly diagnosed via direct stool examination for egg detection. Immunological methods are favoured for disease monitoring and preliminary checking for communities in areas with low infection rates, and for patients with light and chronic infections where parasitological tests are negative. PCR-based diagnostic techniques are more sensitive, but expensive. Tegument proteins and miRNAs are promising markers for diagnosis of schistosomiasis. Here we review the diagnostic methods for schistosomiasis mansoni aiming to reach a standardized technique for diagnosis of early infection to help better control of the disease.

258-263

Authors: Chrishantha Abeysena, Sharika Peiris, Indrakantha Welgama, Upul Gunasekara, Kolitha Wickramage

Objective: To describe the symptoms of dengue at acute and post-infection stage and to identify preventive strategies adopted by the patients in the Western Province, Sri Lanka. Methods: This was a descriptive cross-sectional study conducted in the Western Province of Sri Lanka in November 2017. The study population consisted of 473 confirmed dengue patients aged from 18 to 64 years who were hospitalized between June and August 2017. A stratified random sampling technique was applied to select the participants. Data collection was done by trained interviewers, using a structured, pre-tested interviewer-administered questionnaire. Results: The most common symptoms of dengue fever at the acute stage was body ache (81.81%, n=387), followed by headache (71.67%, n=339), loss of appetite (66.80%, n=316), tiredness (53.06 %, n=251) and vomiting (41.86%, n=198). The most common symptom during the post-infection stage was body ache (48.62%, n=230), followed by fatigue (39.74%, n=188), and loss of appetite (28.75%, n=136). Adding sand to the water in places where water accumulates (41.64%, n=197) was the most common method used to prevent transmission of dengue followed by removing/draining stagnant water regularly (36.15%, n=177). Use of mosquito nets (83.08%, n=393) was the most common method in preventing mosquito bites followed by the use of fans among dengue patients (56.02%, n=265). Conclusions: The study shows a prolonged persistence of clinical symptoms during the postinfection period.

272-282

Authors: Subin Varghese Thomas, John Paul, Akhila Hosur Shrungeswara, Subhankar Biswas, Aarti Abhishek Shah, Chandrashekar Kodangala Subraya, Yogendra Nayak

Objective: To evaluate the anticancer potentials of Annona muricata fruit by in vitro and in vivo methods. Methods: The ethanolic extract of Annona muricata fruit was prepared by Soxhlet extraction method and further fractionated with petroleum ether, ethyl acetate and chloroform. The fractions were tested for cytotoxicity, apoptosis, scratch wound assay, and cell cycle analysis. IC50, apoptotic index and percentage cell migration were determined using HepG2 cells. For the in vivo studies, hepatocellular carcinoma was induced by administering 0.01% diethylnitrosamine (DEN) in drinking water in Wistar rats. In pre-treatment, rats were co-administered 200 mg/kg of fruit extract with DEN for 14 weeks. In post-treatment, the extract was co-administered after 8-weeks of DEN-induction for 14 weeks. Liver function test, haematological test, oxidative stress markers, relative liver weight, number of cancer nodules and histopathological parameters were determined. Results: Annona muricata fruit extract =significantly lowered cell proliferation counts. The chloroform-fraction possessed higher activity [IC50=(53.7±4.3) μg/mL]. The chloroform fraction inhibited cell migration, which was significant compared to curcumin. Further investigations regarding the mode of anticancer activity revealed that the chloroform fraction induced apoptosis. The cell cycle analysis indicated that cells were being arrested at G0/G1. In the in vivo studies, the DEN-control group showed a significant decrease in body weights with increased mortality rate, hepatic nodules, and impairment of liver function compared to normal rats. The rats pre-treated and post-treated with the extract showed positive results with significant improvement in the parameters that were adversely affected by DEN. In addition, other adverse effects of DEN, such as blood dyscrasias and hepatic endogenous antioxidant, were significantly attenuated by Annona muricata fruit extract. Conclusions: The Annona muricata fruit extract has anticancer activity when tested by in vitro and in vivo hepatocellular cancer models.

288-290

Authors: Yi Hern Tan, Lishan Jessica Quah, Humaira Shafi

Rationale: Acinetobacter radioresistens is a non-fermentative Gram-negative coccobacillus that is environmentally ubiquitous and is an uncommon cause of pneumonia in an immunocompetent patient with no known chronic medical illness. Patient concerns: A middle-aged Asian male with a smoking history presented with fever and cough. Physical examination was unremarkable. Chest imaging was consistent with pulmonary parenchymal infection and blood culture grew Acinetobacter radioresistens. Diagnosis: Community acquired pneumonia with Acinetobacter radioresistens bacteremia. Interventions: The patient received a combination of intravenous and oral ampicillinsulbactam over 2 weeks. Outcomes: Repeat blood cultures showed resolution of bacteremia. Completion of antimicrobial treatment saw resolution of respiratory symptoms and radiological pneumonic changes. Lessons: Acinetobacter radioresistens causing community-acquired pneumonia in an immunocompetent host has never been described before. It may be a novel emerging infectious agent in pulmonary infections. Its clinical course in this immunocompetent patient appears to be relatively benign.

195-203

Authors: Kabita Gogoi, Gokul Baishya, Biswajit Saikia, Nabin Chandra Barua, Chandrajit Dohutia, Akalesh Kumar Verma, Anil Prakash

Objective: To obtain suitable artimisinin-based drug candidates with high antimalarial activity. Methods: Three different reaction schemes were used to synthesize a total of 15 artemisininbased compounds. The first synthetic scheme involved the synthesis of diazido aliphatic and aromatic compounds from commercially available dihalides and azido derivatives of artemisinin. The second scheme consisted of the reaction of dibromoaliphatic compounds with sodium azide in dimethylformamide which yielded the desired compounds. Artemisinin-based compounds on treatment with sodium azide and bromotrimethylsilane in dichloromethane produced the most potent compound GB-2. Another potent compound GB-1 was synthesized from artemisinin by treatment with alcohols in the presence of Aberlyst-15 in anhydrous dichloromethane. The third scheme involved the Huisgen 1,3-dipolar cycloaddition between the synthesized aliphatic and aromatic diazides and two alkyne derivatives of artemisinin to obtain the desired artemisinin dimers with average yields. Results: The best in vitro antiplasmodial activity was shown by the compound GB-2 registering IC50 value 0.066 μg/mL against chloroquine-sensitive and 0.865 μg/mL against chloroquineresistant strains of Plasmodium falciparum. It suppressed 59.0% parasitaemia in vivo of rodent malaria parasite Plasmodium berghei in Swiss albino model at 50 μg/kg body weight dosage. Molecular docking interactions of Plasmodium falciparum ATP6 (PfATP6) protein revealed strong bonding of GB-2 with Thr255 residue which is likely to be the reason for excellent antimalarial activity of this compound. Conclusion: Two compounds GB-1 and GB-2 exhibited excellent in vitro antiplasmodial activity and fair in vivo antimalarial activity. Of the two, GB-2 showed better activity which could be attributed to its strong bonding interactions with Thr255 as evidenced from the molecular docking study. Study helped in identifying artemisinin analogues possessing good antimalarial properties and further research in structural alterations of the selected molecules should be carried out which may result in obtaining potent drug candidates against the malarial parasite.

216-223

Authors: Eleonora Cella, Domenico Benvenuto, Daniele Donati, Francesco Garilli, Silvia Angeletti, Stefano Pascarella, Massimo Ciccozzi

Objective: To study the genetic diversity of Culex theileri flavivirus and the spread of this virus among Spain, Portugal and Turkey. Methods: A database consisting of 55 sequences of the NS5/3’UTR region of Culex theileri flavivirus group downloaded from GenBank were aligned and manual edited with Bioedit. ModelTest v. 3.7 was used to select the simplest evolutionary model that adequately fitted the sequence data. Maximum likelihood analysis was performed using MEGA7. The phylogenetic signal of the dataset was investigated by the likelihood mapping analysis. Results: The phylogenetic tree showed three clusters. Myanmar sequences clusterd together with Turkish sequences, Spain and Portugal strains grouped together and two Turkish sequences grouped separately. Selective pressure analysis showed a moderate percentage of sites (22.5%) under pervasive negative selection and only 1% under pervasive positive selection. The sites subject to selective pressure in CTFV RdRp NS5 fragments have been located onto the predicted three-dimensional structure. Conclusions: Phylogenetic and evolutionary analysis can be an important tool for understanding the evolutionary impact of the probable contemporary existence between nonpathogenic and pathogenic flaviviruses among these vectors.

232-238

Authors: Ismail Elshahaw, Fatma AbouElenien

Objective: To identify the serological epidemiology of Cryptosporidium infections and to follow up on the changes in the infection profile in Southern Egypt in order to establish a suitable scheme for control and prevention of cryptosporidiosis. Methods: A total of 1 912 (960 from human and 952 from animals) stool specimens and sera were screened for Cryptosporidium species using modified Ziehl Neelsen technique and a newly-developed enzyme-linked immunosorbent assay (ELISA). Environmental risk factors and socioeconomic data were surveyed by questionnaire between September 2016 and December 2017. Results: Totally, 20.83% of the human subjects were positive for Cryptosporidium infection tested by ELISA. The seropositivity was positively correlated with age. The prevalence of Cryptosporidium infections in females was significantly higher than in males (P<0.05). The sensitivity and specificity of ELISA for Cryptosporidium were 99.06% and 88.88%, respectively. Furthermore, a high prevalence of Cryptosporidium in domestic animals (42.20%). Conclusions: The study observed that Cryptosporidium infections are common in the study area, with water sanitation, socioeconomic level; eating habits and hygienic status are considered the main risk factors for cryptosporidiosis. Therefore, environmental sanitation and health education will be useful in reducing the prevalence of infection.

147-152

Authors: Rinku Rozera, Surajpal Verma, Ravi Kumar, Anzarul Haque, Anshul Attri

Dengue fever is a disease that is caused by five dengue virus (DENV) serotypes. It is endemic in more than 128 countries. Millions of people are affected by this disease. But still, there is no specific treatment available till now to combat it. Some of the preventive measures using plant extracts showed certain promising ovicidal activity against Aedes aegypti. There are some vaccines as preventive measure and antiviral agents to cure the dengue fever, which are under clinical trials but their results have not been reported or approved yet. Dengvaxia is the only licensed vaccine to prevent dengue fever in some South American countries, but it is not approved by other regulatory authorities worldwide. Scientific research has been going on to find a permanent cure for dengue fever, but till now, no successful invention has been done. There are some general treatments for dengue patient by employing analgesics and fluid replacement; however, specific treatment is needed to overcome the lethal effect of dengue fever. Since this disease has affected millions of people and not much invention has been made in this field; therefore, advancement in dengue therapy is required for the safety and well-being of dengue patients. Herein we collectively describe the different chemotherapeutics agents, alternative methods like vaccines and antiviral agents to prevent and cure this lethal disease. Furthermore, the future perspective for the treatment of dengue is discussed in this review.

163-169

Authors: Izumi Mashima, Citra F. Theodorea, Boonyanit Thaweboon, Sroisiri Thaweboon, Tippanart Vichayanrat, Frank A. Scannapieco, Futoshi Nakazawa

Objective: To investigate the composition of the salivary microbiome of 50 healthy Thai children. Methods: A total 76 provinces in Thailand are grouped into 5 geographical clusters based on unique economics, foods and lifestyles. Geographical locations and the results of an oral assessment were also considered. Genomic DNA was extracted from stimulated saliva samples. Subsequently, amplicon libraries were prepared by 16S Metagenomic Sequencing Library Preparation. The amplicons were sequenced using an Illumina Miseq platform followed by bioinformatics and statistical analyses. Results: The correlation between oral hygiene status and caries history varied from r 2 =0.887 to r 2 =0.999 in the geographical groups, suggesting oral hygiene status a strong association between caries history. Twenty taxonomic groups were found in all subjects and constituted 93.6%-96.5% of the microbiome. Of these, genus Veillonella and Prevotella showed significant differences in their proportions between the geographical groups (P<0.05). Furthermore, the proportion of Veillonella parvula, as well as Rothia aeria and Rothia dentocariosa tended to increase with worse oral hygiene status, which was also related to higher dental caries history. Conclusions: The differences in the salivary microbiome as related to geographic regions suggest that environmental factors, which may include dietary habits, could influence the predominant bacteria found in the mouth of Thai children, especially the genus Veillonella and Prevotella. The ratio of Veillonella parvula, Rothia aeria and Rothia dentocariosa may be indicators of worse oral hygiene status and future caries in this population.

175-180

Authors: Ahmed Dahiru Balami, Hadiza Umar Meleh

Objective: To determine the spread of misinformation on salt water among Nigerians, salt water use for Ebola prophylaxis, and the role played by the social media during the 2014 Ebola outbreak. Methods: Information was collected from the general Nigerian population through online Google forms which were majorly disseminated via Facebook and WhatsApp platforms. The data retrieved was analysed using descriptive statistics, Chi-square test and multivariate logistic regression. Results: A total of 703 questionnaires were included in the final analysis. The respondents’ mean age was (30.2±6.7) years, predominantly male (73.2%). Almost all of them (95.0%) had received some message to use salt water, 37.4% of whom received such message on social media. Around a half of them ever made an attempt to verify the health information they received on social media (50.9%), and about a quarter (24.0%) of them had used salt water during the outbreak. Many of them had used salt water because they believed it would not harm them (52.0%). Salt water was less likely to be used by those who had received such message on social media, as well as by those who had received some contrary information to salt use. Conclusions: Social media could serve as an effective tool in propagating health information and should be actively engaged by health agencies, to spread accurate information.

185-194

Authors: Shu-Wei Zheng

Rationale: Disseminated nocardiosis due to Nocardia otitidiscaviarum is rarely reported in immunocompetent hosts. Patient concerns: A 59 year old male patient complained of painful soft tissue swellings and fever for two days. Diagnosis: Disseminated nocardiosis due to Nocardia otitidiscaviarum. Interventions: Initial antimicrobial therapy with imipenem and trimethoprim/sulfamethoxazole was switched to 6 weeks of trimethoprim/sulfamethoxazole, linezolid and tigecycline after sensitivity test results were available. Thereafter, the patient was switched to maintenance trimethoprim/sulfamethoxazole and moxifloxacin. Prednisolone was gradually tapered. Outcomes: Soft tissue swelling and pain disappeared and the patient was discharged uneventfully. Lessons: Disseminated nocardiosis due to Nocardia otitidiscaviarum should be suspected in immunocompetent hosts with risk factors such as medication with prednisolone. Early identification of the causative species and susceptibility results is crucial given the diverse resistance patterns amongst various Nocardia species.