THE IMPORTANCE OF MOSQUITO BEHAVIOURAL ADAPTATIONS TO MALARIA CONTROL IN AFRICA

Over the past decade the use of long‐lasting insecticidal nets (LLINs), in combination with improved drug therapies, indoor residual spraying (IRS), and better health infrastructure, has helped reduce malaria in many African countries for the first time in a generation. However, insecticide resistance in the vector is an evolving threat to these gains. We review emerging and historical data on behavioral resistance in response to LLINs and IRS. Overall the current literature suggests behavioral and species changes may be emerging, but the data are sparse and, at times unconvincing. However, preliminary modeling has demonstrated that behavioral resistance could have significant impacts on the effectiveness of malaria control. We propose seven recommendations to improve understanding of resistance in malaria vectors. Determining the public health impact of physiological and behavioral insecticide resistance is an urgent priority if we are to maintain the significant gains made in reducing malaria morbidity and mortality.     

Annexin A2 is a C-terminal PCSK9-binding protein that regulates endogenous low density lipoprotein receptor levels

The proprotein convertase subtilisin/kexin-type 9 (PCSK9), which promotes degradation of the hepatic low density lipoprotein receptor (LDLR), is now recognized as a major player in plasma cholesterol metabolism. Several gain-of-function mutations in PCSK9 cause hypercholesterolemia and premature atherosclerosis, and thus, inhibition of PCSK9-induced degradation of the LDLR may be used to treat this deadly disease. Herein, we discovered an endogenous PCSK9 binding partner by Far Western blotting, co-immunoprecipitation, and pull-down assays. Following two-dimensional gel electrophoresis and mass spectrometry analysis, we demonstrated that PCSK9 binds to a approximately 33-kDa protein identified as annexin A2 (AnxA2) but not to the closely related annexin A1. Furthermore, our functional LDLR assays and small hairpin RNA studies show that AnxA2 and the AnxA2.p11 complex could prevent PCSK9-directed LDLR degradation in HuH7, HepG2, and Chinese hamster ovary cells. Immunocytochemistry revealed that PCSK9 and AnxA2 co-localize at the cell surface, indicating a possible competition with the LDLR. Structure-function analyses demonstrated that the C-terminal cysteine-histidine-rich domain of PCSK9 interacts specifically with the N-terminal repeat R1 of AnxA2. Mutational analysis of this 70-amino acid-long repeat indicated that the RRTKK81 sequence of AnxA2 is implicated in this binding because its mutation to AATAA81 prevents its interaction with PCSK9. To our knowledge, this work constitutes the first to show that PCSK9 activity on LDLR can be regulated by an endogenous inhibitor. The identification of the minimal inhibitory sequence of AnxA2 should pave the way toward the development of PCSK9 inhibitory lead molecules for the treatment of hypercholesterolemia.     

The potential and realized spread of wildfires across Canada

Given that they can burn for weeks or months, wildfires in temperate and boreal forests may become immense (eg., 10⁰ – 10⁴ km²). However, during the period within which a large fire is ‘active’, not all days experience weather that is conducive to fire spread; indeed most of the spread occurs on a small proportion (e.g., 1 – 15 days) of not necessarily consecutive days during the active period. This study examines and compares the Canada‐wide patterns in fire‐conducive weather (‘potential’ spread) and the spread that occurs on the ground (‘realized’ spread). Results show substantial variability in distributions of potential and realized spread days across Canada. Both potential and realized spread are higher in western than in eastern Canada; however, whereas potential spread generally decreases from south to north, there is no such pattern with realized spread. The realized‐to‐potential fire‐spread ratio is considerably higher in northern Canada than in the south, indicating that proportionally more fire‐conducive days translate into fire progression. An exploration of environmental correlates to spread show that there may be a few factors compensating for the lower potential spread in northern Canada: a greater proportion of coniferous (i.e., more flammable) vegetation, lesser human impacts (i.e., less fragmented landscapes), sufficient fire ignitions, and intense droughts. Because a linear relationship exists between the frequency distributions of potential spread days and realized spread days in a fire zone, it is possible to obtain one from the other using a simple conversion factor. Our methodology thus provides a means to estimate realized fire spread from weather‐based data in regions where fire databases are poor, which may improve our ability to predict future fire activity.     

A new eimeriid (Apicomplexa) species from endangered Attwater's prairie chickens (Tympanuchus cupido attwateri) in Texas

The Attwater's prairie chicken (APC; Tympanuchus cupido attwateri Bendire, 1894) has been a federally listed endangered species since 1967. Several captive propagation programs consisting of small populations are being used to keep this species from extinction. Fecal samples were collected from APCs in April 2007 and again in August 2008 from 2 separate captive propagation facilities in Texas after clinical signs of coccidiosis were observed. One Eimeria species was observed (Eimeria attwateri), which we describe as a putative new species. Sporulated oocysts are ellipsoidal, 30.0 × 18.4 (27.4-31.3 × 16.0-22.4) μm. Oocysts have a smooth wall 0.7 μm thick and lack both a micropyle and oocyst residuum, but 1 ellipsoidal polar granule is present, 2.3 × 1.9 (2.1-2.4 × 1.7-2.0) μm. Sporocysts have a nipple-like Stieda body with a rounded opposite end and are 14.0 × 7.1 (10.2-16.8 × 6.0-9.2) μm. The sporocysts contain a sporocyst residuum usually consisting of 2-4 dispersed globules, and each sporozoite contains 2 large posterior spheroid refractile bodies 3.4 μm wide. Nucleotide sequence amplified from the 18S rDNA does not match any DNA sequence information for publicly available Eimeria species, and phylogenetic reconstructions place this species with other eimerians from Galliformes. The discovery of a potentially pathogenic species of Eimeria in captive APCs is of great importance, and managers should be aware of the potential devastating effect(s) this parasite could have on the APC conservation programs.     

Variation analysis and gene annotation of eight MHC haplotypes: The MHC Haplotype Project

The human major histocompatibility complex (MHC) is contained within about 4 Mb on the short arm of chromosome 6 and is recognised as the most variable region in the human genome. The primary aim of the MHC Haplotype Project was to provide a comprehensively annotated reference sequence of a single, human leukocyte antigen-homozygous MHC haplotype and to use it as a basis against which variations could be assessed from seven other similarly homozygous cell lines, representative of the most common MHC haplotypes in the European population. Comparison of the haplotype sequences, including four haplotypes not previously analysed, resulted in the identification of >44,000 variations, both substitutions and indels (insertions and deletions), which have been submitted to the dbSNP database. The gene annotation uncovered haplotype-specific differences and confirmed the presence of more than 300 loci, including over 160 protein-coding genes. Combined analysis of the variation and annotation datasets revealed 122 gene loci with coding substitutions of which 97 were non-synonymous. The haplotype (A3-B7-DR15; PGF cell line) designated as the new MHC reference sequence, has been incorporated into the human genome assembly (NCBI35 and subsequent builds), and constitutes the largest single-haplotype sequence of the human genome to date. The extensive variation and annotation data derived from the analysis of seven further haplotypes have been made publicly available and provide a framework and resource for future association studies of all MHC-associated diseases and transplant medicine. Horton and Gibson contributed equally to this work.     

The helicase domain of the TMV replicase proteins induces the N-mediated defence response in tobacco

Tobacco mosaic virus (TMV) induces the hypersensitive response (HR) in tobacco plants containing the N gene. This defence response is characterized by cell death at the site of virus infection and inhibition of viral replication and movement. A previous study indicated that a portion of the TMV replicase containing a putative helicase domain is involved in HR induction. Here, this observation is confirmed and extended by showing that non-viral expression of a 50 kDa TMV helicase fragment (p50) is sufficient to induce the N-mediated HR in tobacco. Like the HR elicited by TMV infection, transgenic expression of p50 induces a temperature-sensitive defence response. We demonstrate that recombinant p50 protein has ATPase activity, as suggested by the presence of conserved sequence motifs found in ATPase/helicase enzymes. A point mutation that alters one of these motifs abolishes ATPase activity in vitro but does not affect HR induction. These results suggest that features of the TMV helicase domain, independent of its enzymatic activity, are recognized by N-containing tobacco to induce TMV resistance.     

Thermal biology of African lovebirds and Australian grass parakeets

We compared aspects of the thermal biology of two groups of small parrots, of similar body mass, each derived from a range of habitat types, varying in aridity, but indigenous to either southern Africa or Australia. By accounting for phylogenetic differences, we were able to question whether arid zone species have lower metabolic rates and greater thermal tolerances than mesic species in relation to the “pre-adapted” and “post-arrival adaptation” hypotheses. Four species of African lovebird (Agapornis) and four species of Australian grass parakeet (one Neopsephotus and three Neophema species) were investigated. The Rosy-faced Lovebird (Agapornis roseicollis), Bourke's Parakeet (Neopsephotus bourkii) and the Scarlet-chested Parakeet (Neophema splendida) were categorised as arid zone species, Fischer's Lovebird (Agapornis fischeri), the Black-masked Lovebird (Agapornis personatus) and the Elegant Parakeet (Neophema elegans) as semi-arid zone species, and the Black-cheeked Lovebird (Agapornis nigrigenis) and the Turquoise Parakeet (Neophema pulchella) as mesic zone species. Conventional and phylogenetically independent statistical methods yielded no significant differences in the basal metabolic rates of birds from different habitats or between the species assemblages from Africa and Australia.     

Truncated amplification: a method for high-fidelity template-driven nucleic acid amplification

The error rate of conventional PCR is problematic when amplifying from single cells or amplifying segments for protein functional analysis by in vitro translation. We describe truncated amplification, a method for high-fidelity amplification in which DNA polymerase errors are not propagated efficiently and original DNA templates exert greater influence on the amplification process. Truncated amplification utilizes pairs of oligonucleotides and thermal cycling, but it differs from PCR. Truncated amplification amplifies non-exponentially with one or two chimeric oligonucleotides and produces truncated terminal products that are no more than three rounds of replication from the original template. Exon 6 of the p53 gene was utilized as a model system to demonstrate proof of principle. Chimeric oligonucleotides containing three 3'-->5' reversed-deoxynucleotides or 2'-OMe-ribonucleotides at 6-8 nucleotides from the 3 'terminus retained sequence specificity and primer extension activity. With PfuTurbo but not with Taq or Vent (exo-) DNA polymerases, the modified nucleotides completely truncated the DNA polymerase elongation. The resulting truncated terminal products are not templates for further amplification because of the short length of the 3' complementary region. Truncated amplific ation can amplify quadratically or geometrically depending on whether two or one chimeric oligonucleotides are used. Truncated amplification is a promising approach when template-driven amplification is desired to increase thefrequency of error-free products.