Design of a dual-promoter expression vector harboring Sag1 and Gra7 genes from Toxoplasma gondii (RH strain).

Toxoplasmosis, a parasitic disease caused by Toxoplasma gondii, has possible irreparable consequences in immunocompromised patients and fetuses. Finding an effective method of prevention, such as vaccination, is crucial because of the global distribution of the parasite and the lack of effective anti-toxoplasmosis drugs. The Sag1 and Gra7 antigens of T. gondii can induce strong humoral and cell-mediated immune responses. Therefore, to develop a novel DNA vaccine against toxoplasmosis, we prepared a eukaryotic construct expressing the Sag1 and Gra7 genes of T. gondii (RH strain). We then verified the ability of this construct to produce the corresponding Sag1 and Gra7 antigens in mammalian cells. Using specific primers, the complete coding sequences of Sag1 and Gra7 genes were amplified by polymerase chain reaction (PCR) from the genomic DNA of T. gondii. Then, both genes were subcloned into pVitro2-neo-mcs plasmid. The pVitro-Sag1-Gra7 construct was subjected to colony PCR, enzymatic digestion, and sequencing to confirm successful subcloning. Sag1 and Gra7 expression in HeLa cells was investigated. Sag1 and Gra7 were successfully subcloned in pVitro2-neo-mcs plasmid. The expression of Sag1 and Gra7 in HeLa cells was confirmed through Western blot analysis. The recombinant pVitro-Sag1-Gra7 construct that simultaneously produces Sag1 and Gra7 antigens in one mammalian cell may be used to develop a novel protective vaccine against toxoplasmosis.

Equal sensitivity of the new generation QuantiFERON-TB Gold plus in direct comparison with the previous test version QuantiFERON-TB Gold IT.

QuantiFERON-TB Gold IT analyses interferon-γ release from CD4(+) T cells after stimulation with specific tuberculosis (TB) antigens. Its sensitivity is approximately 80% for active TB. A new test generation (QFTGplus) also analyses the response of CD8(+) T cells. We investigated both test generations in a direct head-to-head comparison in a German pulmonary hospital. Sensitivity rates for active TB were identical, no matter whether diagnosis was bacteriologically confirmed or not.

Wild rodents and their ectoparasites in Baluchistan area, southeast of Iran.

During 2008-2009 a total of 67 individuals of rodents, Tetera indica, Meriones hurrianae, Meriones libycus and Gerbillus nanus were trapped in three areas, Bampor, Daman and Qasre Qand from Iranshahr and Nikshahr districts. There is a significant difference between comparative abundance of four species (P<0.05). A total of 1422 ectoparasites collected including 299 mites (21%), 127 fleas (8.9%), 972 lice (68.4%) and 24 ticks (1.7%). Significant findings amongst the ectoparasites is the lice group with three species identified, Laelaps accuninata, Andralaelaps hermophrodita and Paracheylaellaps pyriformis being the first record in the study areas. All four captured genera of rodents are known as main/ potential reservoir hosts of zoonotic cutaneous leishmaniasis. The migration habit of rodents may affect the spatial distribution of parasitic ticks and their transmitted diseases like CCHF, which has been reported in recent years from Sistan and Baluchestan province. Monitoring of rodent populations and their ectoparasites will help to predict the potential of zoonotic arthropod-borne diseases.

Molecular Monitoring of Plasmodium vivax Infection after Radical Treatment in Southeastern Iran.

BACKGROUND: The aim was to evaluate the relapse risk of vivax malaria in patients who received radical treatment in Hormozgan Province, a malarious area located on southeast of Iran. METHODS: A total of 95 symptomatic vivax malaria infected patients were enrolled in urban health centers of Bandar-Abbas, Minab, Bandar-Jask and Bashagard districts of Hormozgan Province, southeast of Iran from January 2008 to March 2009 for consideration as a case- series study. DNA was extracted from parasite infected whole blood samples. A polymorphic region of Plasmodium vivax merozoite surface protein 1 (pvMSP1) was selected and a PCR method was employed for all the samples to amplify the specific variable gene fragment. The obtained fragments in primary and secondary samples were sequenced. Both nucleotide and amino acid sequences of the samples were investigated for returned patients. RESULTS: 3.2% of the patients experienced a second attack between 83-199 days after the initial episode of infection. Alignment of nucleotide and their deduced amino acid sequences between pair sequences of primary and secondary isolates revealed 8 and 6 dissimilarities respectively for the first case, and 9 and 7 dissimilarities for the second case. Although microscopical examination of recurrent thick blood smear of the third patient confirmed new P. vivax infection, the venous blood sample was accidentally missed. Sequencing results of primary and returned isolates 1P, 1S, 2P, 2S and 3P in this study showed an identity with BP13, T117, BP13, TC28 and Chesson genotypes respectively. CONCLUSION: The returned (secondary) isolates may account to be for the sake of reinfection.

On-line identification of unstable iridoids from Jamesbrittenia fodina by HPLC-MS and HPLC-NMR.

HPLC-UV-MS analysis of the methanol extract of Jamesbrittenia fodina (Wild) O. M. Hilliard (Scrophulariaceae) revealed the presence of different iridoid cinnamic esters; however, isolation of these constituents was prevented by instability problems. HPLC-UV-MS and HPLC-NMR analysis of the mixtures obtained after a tentative isolation indicated that, in the first instance, instability was due to a light-induced cis/trans isomerisation of the cinnamic moieties. Further investigation of related compounds showed an additional instability problem linked to other chemical transformations. A detailed HPLC-NMR-MS study of these fractions demonstrated that the modifications occurred on the rhamnose moiety of these iridoids. It could be concluded that the second type of instability was attributable to transesterification of the cinnamic moiety on the rhamnose unit. The recording of stop-flow HPLC-NMR spectra for specific HPLC peaks permitted the direct monitoring of these transformations. Based on these on-line data, six new unstable aucubin derivatives were efficiently characterised.

On-line identification of unstable catalpol derivatives from Jamesbrittenia fodina by LC-mS and LC-NMR.

LC-UV-MS analysis of the methanol extract of Jamesbrittenia fodina O. M. Hilliard (Scrophulariaceae) revealed the presence of cinnamic ester derivatives. Two isomeric pairs of these constituents were detected, but could not be isolated. In order to identify these unstable compounds, LC-1H-NMR spectra were obtained for each individual isomer and standard NMR measurements were performed in-mixture. The spectra clearly demonstrated that the fractions consisted of mixtures of cis and trans cinnamoyl catalpol glycoside esters.

Study of two plants used in traditional medicine in Zimbabwe for skin problems and rheumatism: Dioscorea sylvatica and Urginea altissima.

Two plants used in Zimbabwean traditional medicine, Dioscorea sylvatica (Dioscoreaceae) and Urginea altissima (Liliaceae), produce mild inflammation and itching when rubbed on the skin. Investigations of the causes of these cutaneous reactions showed that raphides of calcium oxalate are, at least in part, responsible for the effects. Histamine could not be detected.

Xanthones from Hypericum roeperanum.

Four new xanthones have been isolated from the roots of Hypericum roeperanum. Their structures have been established by a combination of spectroscopic and chemical methods as 1,6-dihydroxy-5-methoxy-4',5'-dihydro-4',4',5'-trimethylfurano- (2',3':3,4)-xanthone (5-O-methyl-2-deprenylrheediaxanthone B), 1,6-dihydroxy-5-methoxy-6',6'-dimethylpyrano-(2',3':3,4)-xanthone (5-O-methylisojacareubin), 1,3,5,6-tetrahydroxy-2-(2',2'-dimethyl-4'-isopropenyl)-cyclopen tanylxanthone (5-O-demethylpaxanthonin) and 1,3,5,6-tetrahydroxy-4-trans-sesquilavandulylxanthone (roeperanone). In addition, 2-hydroxyxanthone, 5-hydroxy-2-methoxyxanthone, 1,5-dihydroxy-2-methoxyxanthone, 2-deprenyl rheediaxanthone B, isojacareubin and calycinoxanthone D have been isolated and characterized. Some of the isolated xanthones exhibited antifungal activity against Candida albicans.

Antifungal and antibacterial chalcones from Myrica serrata.

The dichloromethane extract of the leaves of Myrica serrata inhibits growth of Cladosporium cucumerinum, Bacillus subtilis, and Escherichia coli on TLC plates. Activity-guided fractionation led the isolation of 2',4'-dihydroxy-6'-methoxy-3',5'-dimethylchalcone (1), 2',4'-dihydroxy-6'-methoxy-5'-methylchalcone (aurentiacin A) (2), 2',6'-dihydroxy-4'-methoxy-3',5'-dimethyldihydrochalcone (3), 2'-hydroxy-4',6'-dimethoxy-3'-methyldihydrochalcone (4), and 2', 6'-dihydroxy-4'-methoxy-3'-methyldihydrochalcone (5). In addition, the flavanones demethoxymatteucinol (6) and cryptostrobin (7) were also identified.

Larvicidal constituents of Melantheria albinervia.

Bioactivity-guided fractionation has led to the isolation of two larvicidal diterpenes, active against the yellow fever-transmitting mosquito Aedes aegypti, from a dichloromethane root extract of Melantheria albinervia (Asteraceae), a plant from Zimbabwe. These diterpenes were identified as ent-kaur-16-en-19-oic acid and 9(11),16-kauradien-19-oic acid. The diterpenes were also weakly active against Bacillus subtilis.

Antimicrobial activity of essential oil from Hoslundia opposita.

The essential oil from the leaves of Hoslundia opposite was extracted by hydrodistillation. GC/MS analysis of the volatile oil showed that it contains largely the sesquiterpenes and sesquiterpene alcohols. The essential oil was tested against eight different bacterial species and one fungal species. The antibacterial and antifungal properties of the essential oil from Hoslundia opposite were determined by using seeded agar plates with wells into which was placed the oil, and flasks of yeast extract and sucrose broth for mycelial growth of Aspergillus niger. After incubation for 24 hours, the diameter of the inhibition zone was measured for the antibacterial tests and after seven days, the dry weight of the mycelia was measured and a percentage of inhibition calculated using controls where no samples were added. The results obtained showed that the essential oil from this plant has significant activity against Aspergillus niger, Acinetobacter calcoacetica, Brochothrix thermosphacta and Flavobacterium suaveolens. These were most affected by the volatile oil from Hoslundia opposita.

Indigenous plant remedies in Zimbabwe.

Two household surveys undertaken in Zimbabwe between 1981 and 1983 revealed extensive use of indigenous plant remedies in the home-management of childhood diarrhoea and many adult illnesses. Names of the local plants, trees and shrubs are listed, together with the part of the plant used and the type of condition treated. The usage of medicinal plants underscores the need for further study of indigenous pharmacopoeias and the therapeutic properties of plants. The role of indigenous plant remedies within local health care systems is also worthy of closer investigation.