Asian Pacific Journal of Tropical Biomedicine

Issue 8,2014 Table of Contents

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  • 1  An overview of travel-associated central nervous system infectious diseases: risk assessment, general considerations and future directions
    Morteza Izadi Arman Is’haqi Mohammad Ali Is’haqi Nematollah Jonaidi Jafari Fatemeh Rahamaty Abdolali Banki
    2014(8):589-596. DOI: 10.12980/APJTB.4.2014APJTB-2014-0065
    [Abstract](29) [HTML](0) [PDF 532.65 K](74)
    Nervous system infections are among the most important diseases in travellers. Healthy travellers might be exposed to infectious agents of central nervous system, which may require in-patient care. Progressive course is not uncommon in this family of disorders and requires swift diagnosis. An overview of the available evidence in the field is, therefore, urgent to pave the way to increase the awareness of travel-medicine practitioners and highlights dark areas for future research. In November 2013, data were collected from PubMed, Scopus, and Web of Knowledge (1980 to 2013) including books, reviews, and peer-reviewed literature. Works pertained to pre-travel care, interventions, vaccinations related neurological infections were retrieved. Here we provide information on pre-travel care, vaccination, chronic nervous system disorders, and post-travel complications. Recommendations with regard to knowledge gaps, and state-of-the-art research are made. Given an increasing number of international travellers, novel dynamic ways are available for physicians to monitor spread of central nervous system infections. Newer research has made great progresses in developing newer medications, detecting the spread of infections and the public awareness. Despite an ongoing scientific discussion in the field of travel medicine, further research is required for vaccine development, state-of-the-art laboratory tests, and genetic engineering of vectors.
    2  No structural change due to G228S substitution of haemagglutinin in emerging H6N1 influenza virus
    Beuy Joob Viroj Wiwanitkit
    2014(8):597-597. DOI: 10.12980/APJTB.4.2014C1319
    [Abstract](8) [HTML](0) [PDF 182.95 K](74)
    3  Genetic diversity of the msp-1, msp-2, and glurp genes of Plasmodium falciparum isolates along the Thai-Myanmar borders
    Kanungnit Congpuong Rungniran Sukaram Yuparat Prompan Aibteesam Dornae
    2014(8):598-602. DOI: 10.12980/APJTB.4.2014APJTB-2014-0156
    [Abstract](23) [HTML](0) [PDF 467.78 K](79)
    Objective: To study the genetic diversity at the msp-1, msp-2, and glurp genes of Plasmodium falciparum (P. falciparum) isolates from 3 endemic areas in Thailand: Tak, Kanchanaburi and Ranong provinces. Methods: A total of 144 P. falciparum isolates collected prior to treatment during January, 2012 to June, 2013 were genotyped. DNA was extracted; allele frequency and diversity of msp-1, msp-2, and glurp genes were investigated by nested polymerase chain reaction. Results: P. falciparum isolates in this study had high rate of multiple genotypes infection (96.5%) with an overall mean multiplicity of infection of 3.21. The distribution of allelic families of msp-1 was significantly different among isolates from Tak, Kanchanaburi, and Ranong but not for the msp-2. K1 and MAD20 were the predominant allelic families at the msp-1 gene, whereas alleles belonging to 3D7 were more frequent at the msp-2 gene. The glurp gene had the least diverse alleles. Population structure of P. falciparum isolates from Tak and Ranong was quite similar as revealed by the presence of similar proportions of MAD20 and K1 alleles at msp-1 loci, 3D7 and FC27 alleles at msp-2 loci as well as comparable mean MOI. Isolates from Kanchanaburi had different structures; the most prevalent alleles were K1 and RO33. Conclusions: The present study shows that P. falciparum isolates from Tak and Ranong provinces had similar allelic pattern of msp-1 and msp-2 and diversity but different from Kanchanaburi isolates. These allelic variant profiles are valuable baseline data for future epidemiological study of malaria transmission and for continued monitoring of polymorphisms associated with antimalarial drug resistance in these areas.
    4  Impact of antibacterial drugs on human serum paraoxonase-1 (hPON1) activity: an in vitro study
    Hakan S?yüt Elif Duygu Kaya ?ükrü Beydemir
    2014(8):603-609. DOI: 10.12980/APJTB.4.2014APJTB-2014-0059
    [Abstract](20) [HTML](0) [PDF 732.89 K](82)
    Objective: To investigate the in vitro effects of the antibacterial drugs, meropenem trihydrate, piperacillin sodium, and cefoperazone sodium, on the activity of human serum paraoxonase (hPON1). Methods: hPON1 was purified from human serum using simple chromatographic methods, including DEAE-Sephadex anion exchange and Sephadex G-200 gel filtration chromatography. Results: The three antibacterial drugs decreased in vitro hPON1 activity. Inhibition mechanisms meropenem trihydrate was noncompetitive while piperacillin sodium and cefoperazone sodium were competitive. Conclusions: Our results showed that antibacterial drugs significantly inhibit hPON1 activity, both in vitro, with rank order meropenem trihydrate piperacillin sodium cefoperazone sodium in vitro.
    5  High levels of Zinc-α-2-Glycoprotein among Omani AIDS patients on combined antiretroviral therapy
    Sidgi Syed Anwer Hasson Mohammed Saeed Al-Balushi Muzna Hamed Al Yahmadi Juma Zaid Al-Busaidi Elias Antony Said Mohammed Shafeeq Othman Talal Abdullah Sallam Mohammed Ahmad Idris Ali Abdullah Al-Jabri
    2014(8):610-613. DOI: 10.12980/APJTB.4.201414B126
    [Abstract](16) [HTML](0) [PDF 547.45 K](78)
    Objective: To investigate the levels of zinc-α-2-glycoprotein (ZAG) among Omani AIDS patients receiving combined antiretroviral therapy (cART). Methods: A total of 80 Omani AIDS patients (45 males and 35 females), average age of 36 years, who were receiving cART at the Sultan Qaboos University Hospital (SQUH), Muscat, Oman, were tested for the levels of ZAG. In addition, 80 healthy blood donors (46 males and 34 females), average age of 26 years, attending the SQUH Blood Bank, were tested in parallel as a control group. Measurement of the ZAG levels was performed using a competitive enzyme-linked immunosorbent assay and in accordance with the manufacturer’s instructions. Results: The ZAG levels were found to be significantly higher among AIDS patients compared to the healthy individuals (P=0.033). A total of 56 (70%) of the AIDS patients were found to have higher levels of ZAG and 16 (20%) AIDS patients were found to have high ZAG levels, which are significantly (P>0.031) associated with weight loss. Conclusions: ZAG levels are high among Omani AIDS patients on cART and this necessitates the measurement of ZAG on routine basis, as it is associated with weight loss.
    6  Toxicity effects of water extracts of Holothuria atra Jaeger in mice
    Ridzwan Bin Hashim Nurul Alia Azizan Zaitunnatakin Zamli Farah Hanis Zulkipli Nurzafirah Mazlan Osama Yousef Althunibat
    2014(8):614-617. DOI: 10.12980/APJTB.4.201414B154
    [Abstract](25) [HTML](0) [PDF 1.03 M](77)
    Objective: To determine lethal median dose (LD50) and histopathological toxicity of water extract of Holothuria atra (H. atra) in mice. Methods: The behavioral changes, mortality and histopathology examination on liver were assessed in mice 14 d after the administration (i.p.) of H. atra water extract. Seven doses (10, 20, 30, 50, 100, 150 and 200 mg/kg) of H. atra were used. The control group was treated with normal saline. Results: In the acute study in mice, the water extracts of H. atra caused dose-dependent general behavior adverse affects and mortality. The main behavioral sign of toxicity was hypoactivity, noticed immediately after administration of the extract which was more obvious at the higher doses and persisted until death. Mortality increased with increasing doses, the calculated LD50 was 41 mg/kg in mice. The liver toxicity was confirmed by histopathological examination, which indicated the presence of abnormal hepatocytes with a distorted shape and undefined cell lining as well as enlarged nuclei in low doses groups. High doses groups indicated a more prominent distortion of the polyhedral hepatocytes with undefined cell lining, massive cytoplasm, pyknotic, karyorhexis and karyolytic nuclei (necrosis of hepatocytes). Control group showed polyhedral hepatocytes with defined cell lining arranged in cords and normal round nuclei, with granular cytoplasm. Conclusions: Because of the relatively low LD50 value in the acute study in mice, it may be concluded that the H. atra water extract is toxic.
    7  Prevention of renal dysfunction by nutraceuticals prepared from oil rich plant foods
    Sahar Y. Al-Okbi Doha A. Mohamed Thanaa E. Hamed Reham SH. Esmail Souria M. Donya
    2014(8):618-626. DOI: 10.12980/APJTB.4.201414B66
    [Abstract](35) [HTML](0) [PDF 943.02 K](80)
    Objective: To investigate the protective effect of extracts prepared from avocado, walnut, flaxseed and Eruca sativa seeds in a rat model of kidney dysfunction induced by intraperitoneal cisplatin. Methods: Ethanol and petroleum ether extracts mixture was prepared from each plant. Six groups of rats were conducted; control healthy, cisplatin group and four test groups where rats were given daily oral dose of each extract mixture before cisplatin injection. Different biochemical and cytogenetic parameters and kidney histopathology were determined. Acute toxicity was tested for the nutraceuticals. Total phenolic contents, fatty acids (FA) and unsaponifiable matter were assessed in the extracts. Results: Walnut ethanol extract showed the highest content of total phenolic. FA analysis revealed that all the studied plants were rich in unsaturated FA. Gas-liquid chromatographic investigation of the unsaponifiable matter showed the presence of campesterol, stigmasterol and β-sitosterol in all the studied plants. Cisplatin treatment induced significant increase in plasma urea, creatinine and malondialdehyde along with significant reduction of plasma albumin, total protein, catalase and total antioxidant as well as reduction in creatinine clearance. Histopathological examination proved the induction of kidney dysfunction. Some sorts of chromosomal aberration and sperm-shape abnormalities were noticed after cisplatin treatment. Administration of extracts mixtures produced improvements in biochemical, histopathological and cytogenetic parameters. Conclusions: Administration of the studied nutraceuticals proved to possess protective role against cisplatin-induced nephrotoxicity, chromosomal aberration and abnormal sperms. All studied nutraceuticals showed complete safety.
    8  Polyketide and benzopyran compounds of an endophytic fungus isolated from Cinnamomum mollissimum: biological activity and structure
    Carolina Santiago Lin Sun Murray Herbert Gibson Munro Jacinta Santhanam
    2014(8):627-632. DOI: 10.12980/APJTB.4.2014APJTB-2014-0030
    [Abstract](23) [HTML](0) [PDF 691.05 K](85)
    Objective: To study bioactivity and compounds produced by an endophytic Phoma sp. fungus isolated from the medicinal plant Cinnamomum mollissimum. Methods: Compounds produced by the fungus were extracted from fungal broth culture with ethyl acetate. This was followed by bioactivity profiling of the crude extract fractions obtained via high performance liquid chromatography. The fractions were tested for cytotoxicity to P388 murine leukemic cells and antimicrobial activity against bacteria and pathogenic fungi. Compounds purified from active fractions which showed antibacterial, antifungal and cytotoxic activities were identified using capillary nuclear magnetic resonance analysis, mass spectrometry and admission to AntiMarin database. Results: Three known compounds, namely 4-hydroxymellein, 4,8-dihydroxy-6-methoxy-3- methyl-3,4-dihydro-1H-isochromen-1-one and 1-(2,6-dihydroxyphenyl) ethanone, were isolated from the fungus. The polyketide compound 4-hydroxymellein showed high inhibitory activity against P388 murine leukemic cells (94.6%) and the bacteria Bacillus subtilis (97.3%). Meanwhile, 4,8-dihydroxy-6-methoxy-3-methyl-3,4-dihydro-1H-isochromen-1-one, a benzopyran compound, demonstrated moderate inhibitory activity against P388 murine leukemic cells (48.8%) and the fungus Aspergillus niger (56.1%). The second polyketide compound, 1 (2,6-dihydroxyphenyl) ethanone was inactive against the tested targets. Conclusions: These findings demonstrate the potential of endophytes as producers of pharmacologically important compounds, including polyketides which are major secondary metabolites in fungi.
    9  Physico-chemical analysis and antimicrobial potential of Apis dorsata, Apis mellifera and Ziziphus jujube honey samples from Pakistan
    Hira Fahim Javid Iqbal Dasti Ihsan Ali Safia Ahmed Muhammad Nadeem
    2014(8):633-641. DOI: 10.12980/APJTB.4.2014APJTB-2014-0095
    [Abstract](20) [HTML](0) [PDF 908.33 K](72)
    Objective: To evaluate physico-chemical properties and antimicrobial potential of indigenous honey samples against different reference strains including Escherichia coli ATCC 8739, Enterobacter aerogenes ATCC 13048, Pseudomonas aeroginosa ATCC 9027, Bacillus subtilis ATCC 6633, Staphylococcus aureus ATCC 25923, Salmonella typhi ATCC 14028, Klebsiella pneumonia ATCC 13883, Aspergillus niger ATCC 16404, Rhizopus oligosporus PCSIR1, Candida albicans ATCC 14053 and Candida utilis ATCC 9950. Methods: By using standard methods samples were evaluated for their antimicrobial properties including additive effect of starch and non-peroxidase activity, antioxidative properties (phenol contents, flavonoid contents, 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity). Prior to this evaluation, complete physico-chemical properties including pH, color, ash contents, protein contents, moisture contents, hydroxymethyl furfural contents, total sugar contents, reducing sugar and non-reducing sugar contents were analyzed. Results: Relatively higher ash contents were found in the Siddar honey i.e. (0.590 0±0.033 6)% and small honey showed relatively higher protein contents i.e. (777.598±9.880) mg/kg. The moisture contents of tested honey samples ranged between 13.8%-16.6%, total sugar contents from 61.672%- 72.420% and non-reducing sugar contents from 1.95%-3.93%. Presences of phenolic contents indicate higher antioxidant potential of these honey samples. All bacteria showed clear inhibition zones in response to tested honey samples whereas fungi and yeast showed inhibition at higher concentrations of these honey samples. For Escherichia coli, Bacillus subtilis, Salmonella typhi, Pseudomonas aeroginosa and Aspergillus niger, overall the small honey showed the higher activity than other honey samples. Conclusion: Physico-chemical analysis of honey samples confirmed good quality of honey according to the standards set by European Union Commission and Codex Alimentarius Commission. Evaluation of these honey samples confirms antimicrobial potential of particular types of honeys indigenous to Pakistan.
    10  An efficient method in breaking of dormancy from Bunium persicum (Boiss) Fedtsch seeds: a valuable herb of Middle East and Central Asia
    Yousef Emamipoor Mahmood Maziah
    2014(8):642-649. DOI: 10.12980/APJTB.4.2014APJTB-2014-0042
    [Abstract](25) [HTML](0) [PDF 2.26 M](71)
    Objective: To develop a protocol for breaking of seed dormancy and increasing the seed germination rate of Bunium persicum. Methods: The seeds were treated with 3.1, 6.3, 12.5, 25, 50 and 100 µmol/L of benzyl aminopurine, gibberellic acid (GA3), thidiazuron (TDZ) and forchlorfenuron. Then, seeds were transferred to two different temperature conditions including room temperature (25 °C) and chilling temperature (2-5 °C). Results: The treatment of moist seeds with chilling temperature (2-5 °C) broke seed dormancy and showed maximum germination, which was 54.7% after 60 d treatment. Also, the treatment of dry seeds with chilling temperature broke seed dormancy with 9.3% germination rate after 120 d. Treatment of seeds with different level of plant growth regulators showed that under moist-room condition, there was evidence of higher and lower seed germination rate: GA3 (100 µmol/L) with 46.7% and TDZ (50 µmol/L) with 6.67% respectively. In addition, the results showed that under moist-chilling condition, TDZ (6.3 µmol/L) with 53.3% seed germination rate had higher influence on breaking seed dormancy. Treatment of seeds with combination of TDZ and GA3 under moist-chilling condition revealed higher rate of breaking of seed dormancy when 6.3 µmol/L TDZ was combined with 100 µmol/L GA3, showing 93.7% germination rate. Conclusions: The effect of plant growth regulators coupled with chilling temperature on breaking of seed dormancy could provide a large number of seedlings while the long juvenile time which is the next restricting factor of plantation still remained. Thus, the subsequent growth of seedlings to provide a large number of corms is necessary for successful plantation.
    11  GC/GCMS analysis of the petroleum ether and dichloromethane extracts of Moringa oleifera roots
    Shaheen Faizi Saima Sumbul Muhammed Ali Versiani Rubeena Saleem Aisha Sana Hira Siddiqui
    2014(8):650-654. DOI: 10.12980/APJTB.4.201414B141
    [Abstract](18) [HTML](0) [PDF 282.84 K](82)
    Objective: To explore the phytochemical constituents from petroleum ether and dichloromethane extracts of Moringa oleifera (M. oleifera) roots using GC/GC-MS. Methods: A total of 5.11 kg fresh and undried crushed root of M. oleifera were cut into small pieces and extracted with petroleum ether and dichloromethane (20 L each) at room temperature for 2 d. The concentrated extracts were subjected to their GC-MS analysis. Results: The GC-MS analysis of the petroleum ether and dichloromethane extracts of M. oleifera roots, which showed promising biological activities, has resulted in the identification 102 compounds. These constituents belong to 15 classes of compounds including hydrocarbons, fatty steroids, halocompounds, urea and acids, esters, alcohols, isothiocyanate, thiocyanate, pyrazine, aromatics, alkamides, cyanides, N-hydroxyimine derivatives, unsaturated alkenamides, alkyne and indole. GC/GC-MS studies on petroleum ether extract of the roots revealed that it contained 39 compounds, belonging to nine classes. Cyclooctasulfur S8 has been isolated as a pure compound from the extract. The major compounds identified from petroleum ether extract were trans- 13-docosene (37.9%), nonacosane (32.6%), cycloartenol (28.6%) nonadecanoic acid (13.9%) and cyclooctasulfur S8 (13.9%). Dichloromethane extract of the roots was composed of 63 compounds of which nasimizinol (58.8%) along with oleic acid (46.5%), N-benzyl-N-(7-cyanato heptanamide (38.3%), N-benzyl-N-(1-chlorononyl) amide (30.3%), bis [3-benzyl prop-2-ene]-1-one (19.5%) and N, N-dibenzyl-2-ene pent 1, 5-diamide (11.6%) were the main constituents. Conclusions: This study helps to predict the formula and structure of active molecules which can be used as drugs. This result also enhances the traditional usage of M. oleifera which possesses a number of bioactive compounds.
    12  Glucose-6-phosphate dehydrogenase (G6PD) deficiency is associated with asymptomatic malaria in a rural community in Burkina Faso
    Abdoul Karim Ouattara Cyrille Bisseye Bapio Valery Jean Télesphore Elvira Bazie Birama Diarra Tegwindé Rebeca Compaore Florencia Djigma Virginio Pietra Remy Moret Jacques Simpore
    2014(8):655-658. DOI: 10.12980/APJTB.4.2014APJTB-2014-0100
    [Abstract](31) [HTML](0) [PDF 231.14 K](78)
    Objective: To investigate 4 combinations of mutations responsible for glucose-6-phosphate dehydrogenase (G6PD) deficiency in a rural community of Burkina Faso, a malaria endemic country. Methods: Two hundred individuals in a rural community were genotyped for the mutations A376G, G202A, A542T, G680T and T968C using TaqMan single nucleotide polymorphism assays and polymerase chain reaction followed by restriction fragment length polymorphism. Results: The prevalence of the G6PD deficiency was 9.5% in the study population. It was significantly higher in men compared to women (14.3% vs 6.0%, P=0.049). The 202A/376G G6PD A-was the only deficient variant detected. Plasmodium falciparum asymptomatic parasitaemia was significantly higher among the G6PD-non-deficient persons compared to the G6PD-deficient (P<0.001). The asymptomatic parasitaemia was also significantly higher among G6PD non-deficient compared to G6PD-heterozygous females (P<0.001). Conclusions: This study showed that the G6PD A- variant associated with protection against asymptomatic malaria in Burkina Faso is probably the most common deficient variant.
    13  Digestive fungal flora in asymptomatic subjects in Bobo-Dioulasso, Burkina Faso
    Bamba Sanata Ouédraogo Abdoul Salam Sangaré Ibrahim Zida Adama Cissé Mamoudou Karou D. Simplice Simpore Jacques Guiguemdé T. Robert Hennequin Christophe
    2014(8):659-662. DOI: 10.12980/APJTB.4.201414B27
    [Abstract](16) [HTML](0) [PDF 355.41 K](79)
    Objective: To identify Candida species in asymptomatic subjects in Bobo-Dioulasso (Burkina Faso) by the matrix-assisted laser desorption ionization-time of flight mass spectrometry. Methods: A cross-sectional study was conducted from January to February 2013 in Bobo-Dioulasso to collect fecal and urine specimens from voluntary donors. Fungal strains were isolated on Sabouraud dextrose agar and analyzed using matrix-assisted laser desorption ionisation-time-of-flight mass spectrometry. Results: A total of 135 samples including stools (78.5%, 106/135) and urine (21.5%, 29/135) were analyzed. The results revealed that fecal specimens contained mainly Candida krusei (C. krusei) (42.5%) followed by Candida albicans (29.3%), Candida glabrata (18.0%) and Candida tropicalis (C. tropicalis) (4.7%). C. krusei (34.6%) was also found to be the most frequently identified in urine samples followed by Candida albicans (27.0%), C. tropicalis (15.4%) and Candida parapsilosis. However, uncommon species such as Candida nivariensis, Candida kefyr, Candida norvegensis, Candida parapsilosis, Candida lusitaniae and Candida robusta were also identified from fecal and urines samples. Conclusions: This study noted the emergence of species such as C. krusei, Candida glabrata, Candida parapsiolosis, C. tropicalis, Candida nivariensis, Candida norvegensis, and others. It is an imperative to take into account the existence of these species in the therapeutic management of patients in Bobo-Dioulasso.
    14  Iron-chelating and anti-lipid peroxidation properties of 1-(N-acetyl-6-aminohexyl)-3-hydroxy-2-methylpyridin-4-one (CM1) in long-term iron loading β-thalassemic mice
    Kanokwan Kulprachakarn Nittaya Chansiw Kanjana Pangjit Chada Phisalaphong Suthat Fucharoen Robert C. Hider Sineenart Santitherakul Somdet Srichairatanakool
    2014(8):663-668. DOI: 10.12980/APJTB.4.2014APJTB-2014-0155
    [Abstract](21) [HTML](0) [PDF 353.57 K](79)
    Objective: To evaluate the iron-chelating properties and free-radical scavenging activities of 1-(N-acetyl-6-aminohexyl)-3-hydroxy-2-methylpyridin-4-one (CM1) treatment in chronic iron-loaded β-thalassemic (BKO) mice. Methods: The BKO mice were fed with a ferrocene-rich diet and were orally administered with CM1 [50 mg/(] for 6 months. Blood levels of non-transferrin bound iron, labile plasma iron, ferritin (Ft) and malondialdehyde were determined. Results: The BKO mice were fed with an iron diet for 8 months which resulted in iron overload. Interestingly, the mice showed a decrease in the non-transferrin bound iron, labile plasma iron and malondialdehyde levels, but not the Ft levels after continuous CM1 treatment. Conclusions: CM1 could be an effective oral iron chelator that can reduce iron overload and lipid peroxidation in chronic iron overload β-thalassemic mice.
    15  Adult Klebsiella pneumoniae meningitis in Qatar: clinical pattern of ten cases
    Fahmi Yousef Khan Mohammed Abukhattab Mohammed AbuKamar Deshmukh Anand
    2014(8):669-672. DOI: 10.12980/APJTB.4.201414B100
    [Abstract](8) [HTML](0) [PDF 231.05 K](74)
    Objective: To describe the clinical presentation, underlying diseases, antimicrobial susceptibility, treatment and outcome of Klebsiella pneumoniae meningitis patients. Methods: This retrospective study involved all patients with 15 years of age or older who admitted to Hamad General Hospital with culture proven Klebsiella pneumoniae meningitis from January 1, 2007 to December 31, 2012. Results: A total of ten cases were identified (nine males and one female). Their mean age was (43.3±12.8) years. Eight patients (80%) had nosocomial meningitis with neurosurgery being the most frequent associated condition. Fever and altered consciousness were the most frequent symptom. Cerebrospinal fluid showed elevated protein and glucose levels. Gram stain showed Gram-negative rods in 50% of cases, while positive cerebrospinal fluid culture results were found in all patients. Multidrug resistance was observed in two cases, and all patients had received appropriate empirical and definitive antibiotic treatments. The mean duration of intravenous antimicrobial treatment was (19.3±7.0) d and all patients with external ventricular drains underwent removal of the device, while in-hospital mortality was 50%. Conclusions: The number of cases was too small to come up with therapeutic and prognostic conclusions. Further large-scale prospective study is needed.

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