Genetic diversity of noroviruses from outbreaks, sporadic cases and wastewater in Luxembourg 2008-2009.

The genetic diversity of norovirus strains obtained from gastroenteritis outbreaks, sporadic case surveillance and wastewater plants was compared in Luxembourg from October 2008 until June 2009. Except for GI.6 and GIV.1 strains detected exclusively in wastewater, all other genotypes were also found in human samples. Of the nine NoV genotypes detected, only three (GII.4, GIIb/II.3 and GIIc/II.12) were associated with institutional outbreaks. The majority of sequences from all sources belonged to genotype GII.4, including two potentially new sub-variants. Strains collected in the context of outbreaks may significantly under-represent the overall genetic diversity of NoVs circulating in a country.

Effects of pH on plaque forming unit counts and aggregation of MS2 bacteriophage.

AIM: The aim of this study was to determine whether aggregation processes in aqueous phase may explain the decrease in plaque forming unit (PFU) counts for pH close to the isoelectric point (pI) of viral particles (MS2 phages). METHODS AND RESULTS: Loss in PFU was observed for pH < or = pI (pI(MS2) = 3.9): for example, at pH 2.5, loss was approx. 3 log(10) PFU. Particle size analysis combining results of dynamic light scattering and flow particle image analysis was then applied to determine the aggregate state of viral suspensions by recording size distributions. The size of major population significantly changed to 30 nm at neutral pH to more than several micrometres when passing below the isoelectric point. CONCLUSIONS: Our study shows that MS2 phages exhibit significant aggregation processes for pH < or = pI leading to aggregate with sizes of few micrometres. This aggregation process can largely explain the decline in PFU counts. SIGNIFICANCE AND IMPACT OF THE STUDY: It is clear that viral aggregation can be a source of significant bias for PFU assays because in the presence of an aggregate the PFU count can be less than the sum of its constituent particles. Therefore, cautions should be taken in terms of conditions of storage (pH far from pI) to avoid such aggregation artefact.

Theoretical Investigations of the Influence of Pressure on the Selectivity of the Michael Addition of Diphenylmethaneamine to Stereogenic Crotonates.

SIBFA (sum of interactions between fragments ab initio computed) molecular mechanics systematics has been modified to take into account the effect of pressure on intra- and intermolecular energies. The van der Waals radii are related to the pressure, using Bridgman results on the variation of crystal volume, on one hand, and the relation between the volume of an atom and its van der Waals radius on the other. This procedure produces a decrease of the volume of the systems considered. The modified systematics is used for the study of the conformation at 1 atm and 15 kbar of two stereogenic crotonates and of the complexes formed by these two molecules with the diphenylmethaneamine and the three solvent molecules present in the experiment. The results obtained show that in the case of NMECC 1a the diastereoselectivity induced by high pressure and by the presence of methanol proceeds from an important stabilization of the pro-R reactive complex in which the crotonate has a stacked-transoid conformation. This stabilization is mainly due to intermolecular interactions. In the case of the second crotonate considered, NMCC 1b, our results indicate that pressure induces a stabilization of the pro-R and pro-S complexes having the axial conformation for which the reaction exhibits little diastereoselectivity in qualitative agreement with experimental data. This study tends to show that it is possible to account theoretically for the influence of pressure on molecular conformation and/or complex structure, using a molecular mechanics method that is able to take into account the variation of volumes of the different entities present in the system studied.

Theoretical study of the self-association of amphotericin B.

The aim of this present work is the study of self-association of amphotericin B (AmB) at a molecular levels, because of its importance in the toxicity of this antibiotic. Molecular mechanics calculations have been performed considering different conformations of the polar head of AmB, the two most stable ones we have determined (B and C) and the one issued from the X-ray data. Our calculations have shown that both head-to-head and head-to-tail stable dimers were found within an energy range between -30 and -40 kcal/mol, the very stable head-to-head dimer with the polar head within C conformation having an energy of -46.8 kcal/mol. We have shown that both electrostatic and Van der Waals terms contribute to the total interaction energy but their relative weight depends on the conformation of the polar head and on the head-to-head and head-to-tail structures involved in the dimer. Thus the electrostatic contribution does no particularly stabilize the head-to-tail dimer. Furthermore an explicit calculation of the dipole moment in the ground state of AmB has disproved the current assertion upon the greatest stabilization of head-to-tail dimers by electrostatic dipole-dipole interaction. Among all the dimers we have calculated, we have found a group denoted G1 with a geometrical structure consistent with absorption data, namely a blue-shift of the dimer main absorption band with regard to the monomer one. In this group G1 we have found two isoenergetic (-38.8 kcal/mol) very stable head-to-head and head-to-tail dimers. We have found that, as a rule, the self-association of AmB in dimers is more favourable than the complexation with the cholesterol and, in a less extent, with the ergosterol. It seems that these features could be also observed for some trimers, that we have roughly calculated.

Theoretical study of the complexation of amphotericin B with sterols.

The aim of this present work was the study of the intermolecular complexes between amphotericin B (AmB) and either cholesterol or ergosterol. In such complexes the intermolecular interaction energy mainly proceeds from both Van der Waals and H-bonding (via water molecules) forces. Our calculations have shown that the Van der Waals forces slightly favor the AmB-ergosterol complex. Several relative positions of the sterol with regard to AmB lead to energy minima: sterol may be either in contact with the AmB polar head or repelled towards the end of the macrolide ring. It appeared that the role played by some water molecules was to maintain the sterol close to the AmB polar head.

Comparative theoretical study of the conformations of amphotericin methyl ester and amphotericin B polar heads in the presence of water.

Amphotericin methyl ester (AmE) is an interesting derivative of amphotericin B (AmB) because of its enhancement of selectivity against the fungicells. Both AmB and AmE molecules differ by the structure of their polar heads. This work deals with a theoretical study of conformations of the polar head of AmE in the presence of hydration water molecules. The results will be compared with our previous work concerning AmB.

Theoretical study of the geometrical arrangement of GT and GA wobble pairs in two short duplexes, Proton NMR chemical shifts and interaction energy calculations.

NMR shielding constants are calculated for the base protons of duplexes formed by the dodecamer d(CGTGAATTCGCG) and the decamer d(CCAAGATTGG). A good agreement with experimental data is obtained for B-DNA helices in which the wobble GT and GA pairs are in the plane of the corresponding GC pairs of the parent duplexes formed by d(CGCGAATTCGCG) and d(CCAAGCTTGG), if the glycosyl bonds of T and G or A and G are symmetrical with respect to the dyad axis of the Watson-Crick GC pair. Interaction energy calculations show that this type of geometrical arrangement, which implies a distortion of the ribonphosphate backbone of both strands of the duplexes are more stable than those in which only one strand has its conformation modified by the presence of the wobble pair. For the duplex containing the GA pair, NMR chemical shifts as well as interaction energy computations favour the Watson-Crick hydrogen bonding scheme. The variation of the different contributions (intrastrand, interstrand, pair-pair) to the interaction energy between the bases of the duplexes, with the geometrical arrangement of the wobble pairs, is reported.

[Value of immunology in the diagnosis of cutaneous onchocerciasis]. / Intérêt de l'immunologie dans le diagnostic de l'onchocercose cutanée.

In a study of 135 cases, the authors present the dermatologic lesions occasioned by onchocercosis, study and discuss the value of the following three diagnostic procedures: the skin Snip technic, the Mazzotti test, and indirect fluorescent antibody test as used in a region of Africa where onchocercosis is, with dipetalonemosis, the principal filariasis. The skin Snip showed positive in only 11,8% of cases while each of the other technics permitted diagnosis of more than 80% of cases studied. Use of the Mazzotti test in conjunction with indirect fluorescent antibody test proved the origin of dermatologic lesions to be onchocercotic in 100% of the study group while both tests remained negative for subjects in the control group. The authors conclude that, in individual practice the 3 tests should be used conjointly, while, for purposes of mass screening, the determination of fluorescent antibodies must be performed as an adjunct to clinical examination and skin Snip.