Evidence of Dengue Virus Transmission and Factors Associated with the Presence of Anti-Dengue Virus Antibodies in Humans in Three Major Towns in Cameroon

Background

Dengue is not well documented in Africa. In Cameroon, data are scarce, but dengue infection has been confirmed in humans. We conducted a study to document risk factors associated with anti-dengue virus Immunoglobulin G seropositivity in humans in three major towns in Cameroon.

Methodology/Principal Findings

A cross sectional survey was conducted in Douala, Garoua and Yaounde, using a random cluster sampling design. Participants underwent a standardized interview and were blood sampled. Environmental and housing characteristics were recorded. Randomized houses were prospected to record all water containers, and immature stages of Aedes mosquitoes were collected. Sera were screened for anti-dengue virus IgG and IgM antibodies. Risk factors of seropositivity were tested using logistic regression methods with random effects.Anti-dengue IgG were found from 61.4% of sera in Douala (n = 699), 24.2% in Garoua (n = 728) and 9.8% in Yaounde (n = 603). IgM were found from 0.3% of Douala samples, 0.1% of Garoua samples and 0.0% of Yaounde samples. Seroneutralization on randomly selected IgG positive sera showed that 72% (n = 100) in Douala, 80% (n = 94) in Garoua and 77% (n = 66) in Yaounde had antibodies specific for dengue virus serotype 2 (DENV-2).Age, temporary house walls materials, having water-storage containers, old tires or toilets in the yard, having no TV, having no air conditioning and having travelled at least once outside the city were independently associated with anti-dengue IgG positivity in Douala. Age, having uncovered water containers, having no TV, not being born in Garoua and not breeding pigs were significant risk factors in Garoua. Recent history of malaria, having banana trees and stagnant water in the yard were independent risk factors in Yaounde.

Conclusion/Significance

In this survey, most identified risk factors of dengue were related to housing conditions. Poverty and underdevelopment are central to the dengue epidemiology in Cameroon.     

Prolonged treatment with prostaglandin E1 increases the rate of lipolysis in rat adipocytes

Prolonged treatment of adipocytes with certain inhibitors of lipolysis, including N(6)-phenylisopropyl adenosine (PIA) and prostaglandin E(1) (PGE(1)) leads to down-regulation of G(i). Prolonged treatment with PIA increases the rate of lipolysis, and we have reported that tumor necrosis factor-alpha (TNF alpha) stimulates lipolysis by down-regulating G(i). To determine the relationship between G(i) concentration and lipolysis, we have investigated the effect of two other acute inhibitors of lipolysis; PGE(1), which down-regulates G(i), and nicotinic acid (NA), which does not down-regulate G(i). Rat adipocytes were incubated with PIA (300 nM), PGE(1) (3 microM) or nicotinic acid (1 mM) for 24 h. The rate of lipolysis (glycerol release) was increased approximately 2 to 3-fold in PIA-treated cells, and in PGE(1)-treated cells. Conversely, the rate of lipolysis was not altered by the prolonged nicotinic acid treatment. These findings support the hypothesis that the rate of lipolysis in adipocytes is determined, at least partly, by the cellular concentration of G(i) proteins.     

Pharmacogenetic assessment of antipsychotic-induced movement disorders: contribution of the dopamine D3 receptor and cytochrome P450 1A2 genes

Tardive dyskinesia (TD) is characterized by involuntary movements predominantly in the orofacial region and develops in approximately 20% of patients during long-term treatment with typical antipsychotics. The high prevalence of TD and its disabling and potentially irreversible clinical course is an important shortcoming for treatment with typical antipsychotics. The studies presented in this article evaluate the role of single nucleotide polymorphisms in dopamine D3 receptor (DRD3) and CYP1A2 genes for propensity to develop TD in patients with schizophrenia. In theory, a combined pharmacogenetic analysis of pharmacokinetic and pharmacodynamic targets for antipsychotics should improve our ability to identify subpopulations that differ in drug safety profile. This information may in turn contribute to the design of more efficient clinical trials and thus expedite the development and regulatory approval of newer antipsychotic compounds.     

Mycobacterium ulcerans causes minimal pathogenesis and colonization in medaka (Oryzias latipes): an experimental fish model of disease transmission

Mycobacterium ulcerans causes Buruli ulcer in humans, a progressive ulcerative epidermal lesion due to the mycolactone toxin produced by the bacterium. Molecular analysis of M. ulcerans reveals it is closely related to Mycobacterium marinum, a pathogen of both fish and man. Molecular evidence from diagnostic PCR assays for the insertion sequence IS2404 suggests an association of M. ulcerans with fish. However, fish infections by M. ulcerans have not been well documented and IS2404 has been found in other mycobacteria. We have thus, employed two experimental approaches to test for M. ulcerans in fish. We show here for the first time that M. ulcerans with or without the toxin does not mount acute or chronic infections in Japanese Medaka "Oryzias latipes" even at high doses. Moreover, M. ulcerans-infected medaka do not exhibit any visible signs of infection nor disease and the bacteria do not appear to replicate over time. In contrast, similar high doses of the wild-type M. marinum or a mycolactone-producing M. marinum "DL" strain are able to mount an acute disease with mortality in medaka. Although these results would suggest that M. ulcerans does not mount infections in fish we have evidence that CLC macrophages from goldfish are susceptible to mycolactones.     

Reciprocal regulation of Notch and PI3K/Akt signalling in T-ALL cells in vitro

Notch signalling plays an important role in hematopoiesis and in the pathogenesis of T-ALL. Notch is known to interact with Ras and PTEN/PI3K (phosphoinositide-3 kinase)/Akt pathways. We investigated the interaction of Notch with these pathways and the possible reciprocal regulation of these signalling systems in T-ALL cells in vitro. Our analyses indicate that the PI3K/Akt pathway is constitutively active in the four T-ALL cell lines tested. Akt phosphorylation was not altered by the sequestration of growth factors, that is, Akt activation seems to be less dependent on but not completely independent of growth factors, possibly being not subject to negative feedback regulation. PTEN expression was not detected in 3/4 cell lines tested, suggesting the loss of PTEN-mediated Akt activation. Inhibition of the PI3K/Akt pathway arrests growth and enhances apoptosis, but with no modulation of expression of Bax-alpha and Bcl-2 proteins. We analysed the relationship between Notch-1 and the PI3K/Akt signalling and show that inhibition of the Akt pathway changes Notch expression; Notch-1 protein decreased in all the cell lines upon treatment with the inhibitor. Our studies strongly suggest that Notch signalling interacts with PI3K/Akt signalling and further that this occurs in the absence of PTEN expression. The consequences of this to the signalling outcome are yet unclear, but we have uncovered a significant inverse relationship between Notch and PI3K/Akt pathway, which leads us to postulate the operation of a reciprocal regulatory loop between Notch and Ras-PI3K/Akt in the pathogenesis of T-ALL.     

Trophallactic activities in the honeybee brood nest--heaters get supplied with high performance fuel

Honeybees actively regulate their brood temperature by heating between 33 and 36 degrees C if ambient temperatures are lower. Heat is generated by vibrating the flight muscles. Heating rapidly depletes the worker's internal energy; therefore heating performance is limited by the honey that is ingested before the heating process. Stored honey is the predefined fuel for flying and heating, but it is stored at a distance from the broodcomb, causing a potential logistic problem of efficient energy supply in the brood area. Our study focused on the behaviour and the thoracic temperature of the participants in trophallactic food exchanges on the broodcomb. We found that 85.5% of the recipients in a trophallactic food exchange have a higher thoracic temperature during feeding contacts than donors and after the feeding contact the former engage in brood heating more often. The donor bees have lower thoracic temperature and shuttle constantly between honey stores and the broodcomb where they transfer the stored honey to heating bees. Providing heat-emitting workers with small doses of high performance fuel contributes to an economic distribution of resources consistent with physiological conditions of the bees and the ecological requirements of the hive. The trophallaxis-based system is essential to provide the energy-intensive brood warming activity. The emerging independence from ambient temperatures is not only beneficial for brood rearing during times of sudden cold spells, but also enables the honeybees in temperate regions to raise brood in early spring and might be the decisive factor for the occurrence of honeybees in temperate climates in general.     

Purification and characterization of thermostable xylose(glucose) isomerase from Bacillus thermoantarcticus

Xylose isomerase produced by Bacillus thermoantarcticus was purified 73-fold to homogeneity and its biochemical properties were determined. It was a homotetramer with a native molecular mass of 200 kDa and a subunit molecular mass of 47 kDa, with an isoelectric point at 4.8. The enzyme had a K _m of 33 mM for xylose and also accepted D-glucose as substrate. Arrhenius plots of the enzyme activity of xylose isomerase were linear up to a temperature of 85°C. Its optimum pH was around 7.0, and it had 80% of its maximum activity at pH 6.0. This enzyme required divalent cations for its activity and thermal stability. Mn²⁺, Co²⁺ or Mg²⁺ were of comparable efficiency for xylose isomerase reaction, while Mg²⁺ was necessary for glucose isomerase reaction. Journal of Industrial Microbiology & Biotechnology (2001) 27, 234–240. Received 18 March 2001/ Accepted in revised form 03 July 2001     

Genetic structure of the tiger mosquito, Aedes albopictus, in Cameroon (Central Africa)

Background

Aedes albopictus (Skuse, 1884) (Diptera: Culicidae), a mosquito native to Asia, has recently invaded all five continents. In Central Africa it was first reported in the early 2000s, and has since been implicated in the emergence of arboviruses such as dengue and chikungunya in this region. Recent genetic studies of invasive species have shown that multiple introductions are a key factor for successful expansion in new areas. As a result, phenotypic characters such as vector competence and insecticide susceptibility may vary within invasive pest species, potentially affecting vector efficiency and pest management. Here we assessed the genetic variability and population genetics of Ae. albopictus isolates in Cameroon (Central Africa), thereby deducing their likely geographic origin.

Methods and Results

Mosquitoes were sampled in 2007 in 12 localities in southern Cameroon and analyzed for polymorphism at six microsatellite loci and in two mitochondrial DNA regions (ND5 and COI). All the microsatellite markers were successfully amplified and were polymorphic, showing moderate genetic structureamong geographic populations (F_ST = 0.068, P<0.0001). Analysis of mtDNA sequences revealed four haplotypes each for the COI and ND5 genes, with a dominant haplotype shared by all Cameroonian samples. The weak genetic variation estimated from the mtDNA genes is consistent with the recent arrival of Ae. albopictus in Cameroon. Phylogeographic analysis based on COI polymorphism indicated that Ae. albopictus populations from Cameroon are related to tropical rather than temperate or subtropical outgroups.

Conclusion

The moderate genetic diversity observed among Cameroonian Ae. albopictus isolates is in keeping with recent introduction and spread in this country. The genetic structure of natural populations points to multiple introductions from tropical regions.     

Bgp2, a new member of the carcinoembryonic antigen-related gene family, encodes an alternative receptor for mouse hepatitis viruses.

Murine coronaviruses such as mouse hepatitis virus (MHV) infect mouse cells via cellular receptors that are isoforms of biliary glycoprotein (Bgp) of the carcinoembryonic antigen gene family (G. S. Dveksler, C. W. Dieffenbach, C. B. Cardellichio, K. McCuaig, M. N. Pensiero, G.-S. Jiang, N. Beauchemin, and K. V. Holmes, J. Virol. 67:1-8, 1993). The Bgp isoforms are generated through alternative splicing of the mouse Bgp1 gene that has two allelic forms called MHVR (or mmCGM1), expressed in MHV-susceptible mouse strains, and mmCGM2, expressed in SJL/J mice, which are resistant to MHV. We here report the cloning and characterization of a new Bgp-related gene designated Bgp2. The Bgp2 cDNA allowed the prediction of a 271-amino-acid glycoprotein with two immunoglobulin domains, a transmembrane, and a putative cytoplasmic tail. There is considerable divergence in the amino acid sequences of the N-terminal domains of the proteins coded by the Bgp1 gene from that of the Bgp2-encoded protein. RNase protection assays and RNA PCR showed that Bgp2 was expressed in BALB/c kidney, colon, and brain tissue, in SJL/J colon and liver tissue, in BALB/c and CD1 spleen tissue, in C3H macrophages, and in mouse rectal carcinoma CMT-93 cells. When Bgp2-transfected hamster cells were challenged with MHV-A59, MHV-JHM, or MHV-3, the Bgp2-encoded protein served as a functional MHV receptor, although with a lower efficiency than that of the MHVR glycoprotein. The Bgp2-mediated virus infection could not be inhibited by monoclonal antibody CC1 that is specific for the N-terminal domain of MHVR. Although CMT-93 cells express both MHVR and Bgp2, infection with the three strains of MHV was blocked by pretreatment with monoclonal antibody CC1, suggesting that MHVR was the only functional receptor in these cells. Thus, a novel murine Bgp gene has been identified that can be coexpressed in inbred mice with the Bgp1 glycoproteins and that can serve as a receptor for MHV strains when expressed in transfected hamster cells.     

Electron transfer dissociation of peptides generated by microwave D-cleavage digestion of proteins

The nonenzymatic digestion of proteins by microwave D-cleavage is an effective technique for site-specific cleavage at aspartic acid (D). This specific cleavage C-terminal to D residues leads to inherently large peptides (15-25 amino acids) that are usually relatively highly charged (above +3) when ionized by electrospray ionization (ESI) due to the presence of several basic amino acids within their sequences. It is well-documented that highly charged peptide ions generated by ESI are well-suited for electron transfer dissociation (ETD), which produces c- and z-type fragment ions via gas-phase ion/ion reactions. In this paper, we describe the sequence analysis by ETD tandem mass spectrometry (MS/MS) of multiply charged peptides generated by microwave D-cleavage of several standard proteins. Results from ETD measurements are directly compared to CID MS/MS of the same multiply charged precursor ions. Our results demonstrate that the nonenzymatic microwave D-cleavage technique is a rapid (<6 min) and specific alternative to enzymatic cleavage with Lys-C or Asp-N to produce highly charged peptides that are amenable to informative ETD.