Enzymatic activity of Lecithin:retinol acyltransferase: A thermostable and highly active enzyme with a likely mode of interfacial activation

Lecithin:retinol acyltransferase (LRAT) plays a major role in the vertebrate visual cycle. Indeed, it is responsible for the esterification of all-trans retinol into all-trans retinyl esters, which can then be stored in microsomes or further metabolized to produce the chromophore of rhodopsin. In the present study, a detailed characterization of the enzymatic properties of truncated LRAT (tLRAT) has been achieved using in vitro assay conditions. A much larger tLRAT activity has been obtained compared to previous reports and to an enzyme with a similar activity. In addition, tLRAT is able to hydrolyze phospholipids bearing different chain lengths with a preference for micellar aggregated substrates. It therefore presents an interfacial activation property, which is typical of classical phospholipases. Furthermore, given that stability is a very important quality of an enzyme, the influence of different parameters on the activity and stability of tLRAT has thus been studied in detail. For example, storage buffer has a strong effect on tLRAT activity and high enzyme stability has been observed at room temperature. The thermostability of tLRAT has also been investigated using circular dichroism and infrared spectroscopy. A decrease in the activity of tLRAT was observed beyond 70 °C, accompanied by a modification of its secondary structure, i.e. a decrease of its α-helical content and the appearance of unordered structures and aggregated β-sheets. Nevertheless, residual activity could still be observed after heating tLRAT up to 100 °C. The results of this study highly improved our understanding of this enzyme.     

The complex translocation (9;14;14) involving IGH and CEBPE genes suggests a new subgroup in B-lineage acute lymphoblastic leukemia

Many subtypes of acute lymphoblastic leukemia (ALL) are associated with specific chromosomal rearrangements. The complex translocation t(9;14;14), a variant of the translocation (14;14)(q11;q32), is a rare but recurrent chromosomal abnormality involving the immunoglobulin heavy-chain (IGH) and CCAAT enhancer-binding protein (CEBPE) genes in B-lineage ALL (B-ALL) and may represent a new B-ALL subgroup. We report here the case of a 5-year-old girl with B-ALL, positive for CD19, CD38 and HLA-DR. A direct technique and G-banding were used for chromosomal analysis and fluorescentin situ hybridization (FISH) with BAC probes was used to investigate a possible rearrangement of the IGH andCEBPE genes. The karyotype exhibit the chromosomal aberration 46,XX,del(9)(p21),t(14;14)(q11;q32). FISH with dual-color break-apartIGH-specific and CEPBE-specific bacterial artificial chromosome (BAC) probes showed a complex t(9;14;14) associated with a deletion of cyclin-dependent kinase inhibitor 2A (CDKN2A) and paired box gene 5 (PAX5) at 9p21-13 and duplication of the fusion gene IGH-CEBPE.     

DNA barcoding of economically important freshwater fish species from north‐central Nigeria uncovers cryptic diversity

This study examines the utility of morphology and DNA barcoding in species identification of freshwater fishes from north‐central Nigeria. We compared molecular data (mitochondrial cytochrome c oxidase subunit I (COI) sequences) of 136 de novo samples from 53 morphologically identified species alongside others in GenBank and BOLD databases. Using DNA sequence similarity‐based (≥97% cutoff) identification technique, 50 (94.30%) and 24 (45.30%) species were identified to species level using GenBank and BOLD databases, respectively. Furthermore, we identified cases of taxonomic problems in 26 (49.00%) morphologically identified species. There were also four (7.10%) cases of mismatch in DNA barcoding in which our query sequence in GenBank and BOLD showed a sequence match with different species names. Using DNA barcode reference data, we also identified four unknown fish samples collected from fishermen to species level. Our Neighbor‐joining (NJ) tree analysis recovers several intraspecific species clusters with strong bootstrap support (≥95%). Analysis uncovers two well‐supported lineages within Schilbe intermedius. The Bayesian phylogenetic analyses of Nigerian S. intermedius with others from GenBank recover four lineages. Evidence of genetic structuring is consistent with geographic regions of sub‐Saharan Africa. Thus, cryptic lineage diversity may illustrate species’ adaptive responses to local environmental conditions. Finally, our study underscores the importance of incorporating morphology and DNA barcoding in species identification. Although developing a complete DNA barcode reference library for Nigerian ichthyofauna will facilitate species identification and diversity studies, taxonomic revisions of DNA sequences submitted in databases alongside voucher specimens are necessary for a reliable taxonomic and diversity inventory.     

In Vivo Addition of Poly(A) Tail and AU-Rich Sequences to the 3′ Terminus of the Sindbis Virus RNA Genome: a Novel 3′-End Repair Pathway

Alphaviruses are mosquito-transmitted RNA viruses that cause important diseases in both humans and livestock. Sindbis virus (SIN), the type species of the alphavirus genus, carries a 11.7-kb positive-sense RNA genome which is capped at its 5′ end and polyadenylated at its 3′ end. The 3′ nontranslated region (3′NTR) of the SIN genome carries many AU-rich motifs, including a 19-nucleotide (nt) conserved element (3′CSE) and a poly(A) tail. This 3′CSE and the adjoining poly(A) tail are believed to regulate the synthesis of negative-sense RNA and genome replication in vivo. We have recently demonstrated that the SIN genome lacking the poly(A) tail was infectious and that de novo polyadenylation could occur in vivo (K. R. Hill, M. Hajjou, J. Hu, and R. Raju, J. Virol. 71:2693–2704, 1997). Here, we demonstrate that the 3′-terminal 29-nt region of the SIN genome carries a signal for possible cytoplasmic polyadenylation. To further investigate the polyadenylation signals within the 3′NTR, we generated a battery of mutant genomes with mutations in the 3′NTR and tested their ability to generate infectious virus and undergo 3′ polyadenylation in vivo. Engineered SIN genomes with terminal deletions within the 19-nt 3′CSE were infectious and regained their poly(A) tail. Also, a SIN genome carrying the poly(A) tail but lacking a part or the entire 19-nt 3′CSE was also infectious. Sequence analysis of viruses generated from these engineered SIN genomes demonstrated the addition of a variety of AU-rich sequence motifs just adjacent to the poly(A) tail. The addition of AU-rich motifs to the mutant SIN genomes appears to require the presence of a significant portion of the 3′NTR. These results indicate the ability of alphavirus RNAs to undergo 3′ repair and the existence of a pathway for the addition of AU-rich sequences and a poly(A) tail to their 3′ end in the infected host cell. Most importantly, these results indicate the ability of alphavirus replication machinery to use a multitude of AU-rich RNA sequences abutted by a poly(A) motif as promoters for negative-sense RNA synthesis and genome replication in vivo. The possible roles of cytoplasmic polyadenylation machinery, terminal transferase-like enzymes, and the viral polymerase in the terminal repair processes are discussed.     

Impact of Three Rounds of Mass Drug Administration on Lymphatic Filariasis in Areas Previously Treated for Onchocerciasis in Sierra Leone

Background

1974–2005 studies across Sierra Leone showed onchocerciasis endemicity in 12 of 14 health districts (HDs) and baseline studies 2005–2008 showed lymphatic filariasis (LF) endemicity in all 14 HDs. Three integrated annual mass drug administration (MDA) were conducted in the 12 co-endemic districts 2008–2010 with good geographic, programme and drug coverage. Midterm assessment was conducted 2011 to determine impact of these MDAs on LF in these districts.

Methodology/Principal Findings

The mf prevalence and intensity in the 12 districts were determined using the thick blood film method and results compared with baseline data from 2007–2008. Overall mf prevalence fell from 2.6% (95% CI: 2.3%–3.0%) to 0.3% (95% CI: 0.19%–0.47%), a decrease of 88.5% (p = 0.000); prevalence was 0.0% (100.0% decrease) in four districts: Bo, Moyamba, Kenema and Kono (p = 0.001, 0.025, 0.085 and 0.000 respectively); and seven districts had reductions in mf prevalence of between 70.0% and 95.0% (p = 0.000, 0.060, 0.001, 0.014, 0.000, 0.000 and 0.002 for Bombali, Bonthe, Kailahun, Kambia, Koinadugu, Port Loko and Tonkolili districts respectively). Pujehun had baseline mf prevalence of 0.0%, which was maintained. Only Bombali still had an mf prevalence ≥1.0% (1.58%, 95% CI: 0.80%–3.09%)), and this is the district that had the highest baseline mf prevalence: 6.9% (95% CI: 5.3%–8.8%). Overall arithmetic mean mf density after three MDAs was 17.59 mf/ml (95% CI: 15.64 mf/ml–19.55 mf/ml) among mf positive individuals (65.4% decrease from baseline of 50.9 mf/ml (95% CI: 40.25 mf/ml–61.62 mf/ml; p = 0.001) and 0.05 mf/ml (95% CI: 0.03 mf/ml–0.08 mf/ml) for the entire population examined (96.2% decrease from baseline of 1.32 mf/ml (95% CI: 1.00 mf/ml–1.65 mf/ml; p = 0.000)).

Conclusions/Significance

The results show that mf prevalence decreased to <1.0% in all but one of the 12 districts after three MDAs. Overall mf density reduced by 65.0% among mf-positive individuals, and 95.8% for the entire population.     

Changes in leaf gas exchange, water relations, biomass production and solute accumulation in Phragmites australis under hypoxic conditions

The physiological responses to hypoxic stress were studied in the common reed, Phragmites australis (Cav.) Trin. ex Steudel. Growth, leaf gas exchange, water (and ion) relations and osmotic adjustment were determined in hydroponically grown plants exposed to 10, 20 and 30 days of oxygen deficiency. The highest growth of reed seedlings was found in normoxic (aerobic) conditions. Treatment effects on biomass production were relatively consistent within each harvest. Leaf water potential and osmotic potential declined significantly as hypoxia periods increased. However, leaf turgor pressure showed a consistent pattern of increase, suggesting that reed plants adjusted their water status by osmotic adjustment in response to root hypoxia. After 20 and 30 days in the low oxygen treatment, net CO₂ assimilation and stomatal conductance were positively associated and the former variable also had a strong positive relationship with transpiration. Short-term hypoxic stress had a slight effect on the ionic status (K⁺, Ca²⁺ and Mg²⁺) of reed plants. In contrast, soluble sugar concentrations increased more under hypoxic conditions as compared to normoxia. These findings indicate that hypoxia slightly affected the physiological behavior of reed plants.     

Upper Cenomanian bivalves from the Guir Basin (southwestern Algeria): Order Veneroida

The Upper Cretaceous rocks are widely distributed and well exposed in south Algeria and consist in beds rich in macrofauna. For the first time, twenty veneroid species (Bivalvia) are systematically described from the upper Cenomanian deposits of the Guir Basin. While three species were reported since long before [Granocardium desvauxi (Coquand), G. productum (J. de C. Sowerby) and Glossus aquilinus (Coquand)]; Lucina fallax Forbes, Crassatella (Rochella) tenuicostata (Seguenza), Protocardia hillana (J. Sowerby), G. productum (J. de C. Sowerby) var. byzacenica (Pervinquière), Arctica cordata (Sharpe), A. humei (Cox), A. inornata (d’Orbigny), A. picteti (Coquand), Tenea delettrei (Coquand), Paraesa faba (J. de C. Sowerby), Meretrix desvauxi (Coquand) were previously unknown from the Cretaceous of Algerian Sahara. Because of reduced degree of preservation, Sphaera cf. corrugata J. Sowerby, Maghrebella cf. forgemoli (Coquand), Maghrebella sp., Granocardium cf. carolinum (d’Orbigny), Protocardia sp. and Meretrix sp. are tentatively determined. The studied material, found in the lower part of the “Calcaires de Sidi Mohamed Ben Bouziane” Formation, evidences palaeobiogeographic affinities occurring over a wide geographical area: from North Africa, southern Europe to Middle East and India. The present study provides new information to the knowledge of the upper Cenomanian palaeobiology of the studied region.