241 research outputs found
ANP32 proteins are essential for influenza virus replication in human cells
ANP32 proteins have been implicated in supporting influenza virus replication, but most of the work to date has focused on the ability of avian Anp32 proteins to overcome restriction of avian influenza polymerases in human cells. Using a CRISPR approach we show that human ANP32A and ANP32B are functionally redundant but essential host factors for mammalian-adapted influenza A virus (IAV) and influenza B virus (IBV) replication in human cells. When both proteins are absent from human cells, influenza polymerases are unable to replicate the viral genome, and infectious virus cannot propagate. Provision of exogenous ANP32A or –B recovers polymerase activity and virus growth. We demonstrate that this redundancy is absent in the murine Anp32 orthologues: murine Anp32A is incapable of recovering IAV polymerase activity, while murine Anp32B can. Intriguingly, IBV polymerase is able to use murine Anp32A. We show using a domain swap and point mutations that the LRR 5 region comprises an important functional domain for mammalian ANP32 proteins. Our approach has identified a pair of essential host factors for influenza virus replication and can be harnessed to inform future interventions
Evaluation of Five Methods for Total DNA Extraction from Western Corn Rootworm Beetles
Background: DNA extraction is a routine step in many insect molecular studies. A variety of methods have been used to isolate DNA molecules from insects, and many commercial kits are available. Extraction methods need to be evaluated for their efficiency, cost, and side effects such as DNA degradation during extraction.
Methodology/Principal Findings: From individual western corn rootworm beetles, Diabrotica virgifera virgifera, DNA extractions by the SDS method, CTAB method, DNAzol reagent, Puregene solutions and DNeasy column were compared in terms of DNA quantity and quality, cost of materials, and time consumed. Although all five methods resulted in acceptable DNA concentrations and absorbance ratios, the SDS and CTAB methods resulted in higher DNA yield (ng DNA vs. mg tissue) at much lower cost and less degradation as revealed on agarose gels. The DNeasy kit was most time-efficient but was the costliest among the methods tested. The effects of ethanol volume, temperature and incubation time on precipitation of DNA were also investigated. The DNA samples obtained by the five methods were tested in PCR for six microsatellites located in various positions of the beetle’s genome, and all samples showed successful amplifications.
Conclusion/Significance: These evaluations provide a guide for choosing methods of DNA extraction from western corn rootworm beetles based on expected DNA yield and quality, extraction time, cost, and waste control. The extraction conditions for this mid-size insect were optimized. The DNA extracted by the five methods was suitable for further molecular applications such as PCR and sequencing by synthesis
Chlorfenapyr: a new insecticide with novel mode of action can control pyrethroid resistant malaria vectors
<p>Abstract</p> <p>Background</p> <p>Malaria vectors have acquired widespread resistance to many of the currently used insecticides, including synthetic pyrethroids. Hence, there is an urgent need to develop alternative insecticides for effective management of insecticide resistance in malaria vectors. In the present study, chlorfenapyr was evaluated against <it>Anopheles culicifacies </it>and <it>Anopheles stephensi </it>for its possible use in vector control.</p> <p>Methods</p> <p>Efficacy of chlorfenapyr against <it>An. culicifacies </it>and <it>An. stephensi </it>was assessed using adult bioassay tests. In the laboratory, determination of diagnostic dose, assessment of residual activity on different substrates, cross-resistance pattern with different insecticides and potentiation studies using piperonyl butoxide were undertaken by following standard procedures. Potential cross-resistance patterns were assessed on field populations of <it>An. culicifacies</it>.</p> <p>Results</p> <p>A dose of 5.0% chlorfenapyr was determined as the diagnostic concentration for assessing susceptibility applying the WHO tube test method in anopheline mosquitoes with 2 h exposure and 48 h holding period. The DDT-resistant/malathion-deltamethrin-susceptible strain of <it>An. culicifacies </it>species C showed higher LD50 and LD99 (0.67 and 2.39% respectively) values than the DDT-malathion-deltamethrin susceptible <it>An. culicifacies </it>species A (0.41 and 2.0% respectively) and <it>An. stephensi </it>strains (0.43 and 2.13% respectively) and there was no statistically significant difference in mortalities among the three mosquito species tested (p > 0.05). Residual activity of chlorfenapyr a.i. of 400 mg/m<sup>2 </sup>on five fabricated substrates, namely wood, mud, mud+lime, cement and cement + distemper was found to be effective up to 24 weeks against <it>An. culicifacies </it>and up to 34 weeks against <it>An. stephensi</it>. No cross-resistance to DDT, malathion, bendiocarb and deltamethrin was observed with chlorfenapyr in laboratory-reared strains of <it>An. stephensi </it>and field-caught <it>An. culicifacies. </it>Potentiation studies demonstrated the antagonistic effect of PBO.</p> <p>Conclusion</p> <p>Laboratory studies with susceptible and resistant strains of <it>An. culicifacies </it>and <it>An. stephensi</it>, coupled with limited field studies with multiple insecticide-resistant <it>An. culicifacies </it>have shown that chlorfenapyr can be a suitable insecticide for malaria vector control, in multiple-insecticide-resistant mosquitoes especially in areas with pyrethroid resistant mosquitoes.</p
High sample throughput genotyping for estimating C-lineage introgression in the dark honeybee: an accurate and cost-effective SNP-based tool
The natural distribution of the honeybee (Apis mellifera L.) has been changed by humans in recent
decades to such an extent that the formerly widest-spread European subspecies, Apis mellifera
mellifera, is threatened by extinction through introgression from highly divergent commercial strains
in large tracts of its range. Conservation efforts for A. m. mellifera are underway in multiple European
countries requiring reliable and cost-efficient molecular tools to identify purebred colonies. Here, we
developed four ancestry-informative SNP assays for high sample throughput genotyping using the
iPLEX Mass Array system. Our customized assays were tested on DNA from individual and pooled,
haploid and diploid honeybee samples extracted from different tissues using a diverse range of
protocols. The assays had a high genotyping success rate and yielded accurate genotypes. Performance
assessed against whole-genome data showed that individual assays behaved well, although the
most accurate introgression estimates were obtained for the four assays combined (117 SNPs).
The best compromise between accuracy and genotyping costs was achieved when combining two
assays (62 SNPs). We provide a ready-to-use cost-effective tool for accurate molecular identification
and estimation oinfo:eu-repo/semantics/publishedVersio
Insect Pollinated Crops, Insect Pollinators and US Agriculture: Trend Analysis of Aggregate Data for the Period 1992–2009
In the US, the cultivated area (hectares) and production (tonnes) of crops that require or benefit from insect pollination (directly dependent crops: apples, almonds, blueberries, cucurbits, etc.) increased from 1992, the first year in this study, through 1999 and continued near those levels through 2009; aggregate yield (tonnes/hectare) remained unchanged. The value of directly dependent crops attributed to all insect pollination (2009 USD) decreased from 10.69 billion in 2001, but increased thereafter, reaching 11.68 billion and 15.45 billion in 1996 to 5.39 billion and 4.99 and $7.04 billion. Trend analysis demonstrates that US producers have a continued and significant need for insect pollinators and that a diminution in managed or wild pollinator populations could seriously threaten the continued production of insect pollinated crops and crops grown from seeds resulting from insect pollination
Pneumococcal serotype trends, surveillance and risk factors in UK adult pneumonia, 2013-18.
BACKGROUND: Changes over the last 5 years (2013-18) in the serotypes implicated in adult pneumococcal pneumonia and the patient groups associated with vaccine-type disease are largely unknown. METHODS: We conducted a population-based prospective cohort study of adults admitted to two large university hospitals with community-acquired pneumonia (CAP) between September 2013 and August 2018. Pneumococcal serotypes were identified using a novel 24-valent urinary monoclonal antibody assay and from blood cultures. Trends in incidence rates were compared against national invasive pneumococcal disease (IPD) data. Persons at risk of vaccine-type pneumonia (pneumococcal conjugate vaccine (PCV)13 and pneumococcal polysaccharide vaccine (PPV)23) were determined from multivariate analyses. FINDINGS: Of 2934 adults hospitalised with CAP, 1075 (36.6%) had pneumococcal pneumonia. The annual incidence of pneumococcal pneumonia increased from 32.2 to 48.2 per 100 000 population (2013-18), predominantly due to increases in PCV13non7-serotype and non-vaccine type (NVT)-serotype pneumonia (annual incidence rate ratio 1.12, 95% CI 1.04 to 1.21 and 1.19, 95% CI 1.10 to 1.28, respectively). Incidence trends were broadly similar to IPD data. PCV13non7 (56.9% serotype 3) and PPV23non13 (44.1% serotype 8) serotypes were identified in 349 (32.5%) and 431 (40.1%) patients with pneumococcal pneumonia, respectively. PCV13-serotype pneumonia (dominated by serotype 3) was more likely in patients in the UK pneumococcal vaccination clinical risk group (adjusted OR (aOR) 1.73, 95% CI 1.31 to 2.28) while PPV23-serotype pneumonia was more likely in patients outside the clinical risk group (aOR 1.54, 95% CI 1.13 to 2.10). INTERPRETATION: The incidence of pneumococcal CAP is increasing, predominantly due to NVT serotypes and serotype 3. PPV23-serotype pneumonia is more likely in adults outside currently identified clinical risk groups
Genetic structure of drone congregation areas of Africanized honeybees in southern Brazil
As yet, certain aspects of the Africanization process are not well understood, for example, the reproductive behavior of African and European honeybees and how the first Africanized swarms were formed and spread. Drone congregation areas (DCAs) are the ideal place to study honeybee reproduction under natural conditions since hundreds of drones from various colonies gather together in the same geographical area for mating. In the present study, we assessed the genetic structure of seven drone congregations and four commercial European-derived and Africanized apiaries in southern Brazil, employing seven microsatellite loci for this purpose. We also estimated the number of mother-colonies that drones of a specific DCA originated from. Pairwise comparison failed to reveal any population sub-structuring among the DCAs, thus indicating low mutual genetic differentiation. We also observed high genetic similarity between colonies of commercial apiaries and DCAs, besides a slight contribution from a European-derived apiary to a DCA formed nearby. Africanized DCAs seem to have a somewhat different genetic structure when compared to the European
Ocean acidification reduces demersal zooplankton that reside in tropical coral reefs
The in situ effects of ocean acidification on zooplankton communities remain largely unexplored. Using natural volcanic CO2
seep sites around tropical coral communities, we show a threefold reduction in the biomass of demersal zooplankton in
high-CO2 sites compared with sites with ambient CO2. Differences were consistent across two reefs and three expeditions.
Abundances were reduced in most taxonomic groups. There were no regime shifts in zooplankton community composition and
no differences in fatty acid composition between CO2 levels, suggesting that ocean acidification affects the food quantity but
not the quality for nocturnal plankton feeders. Emergence trap data show that the observed reduction in demersal plankton
may be partly attributable to altered habitat. Ocean acidification changes coral community composition from branching to
massive bouldering coral species, and our data suggest that bouldering corals represent inferior daytime shelter for demersal
zooplankton. Since zooplankton represent a major source of nutrients for corals, fish and other planktivores, this ecological
feedback may represent an additional mechanism of how coral reefs will be affected by ocean acidification
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