Analysis of apyrase 5' upstream region validates improved Anopheles gambiae transformation technique

<p>Abstract</p> <p>Background</p> <p>Genetic transformation of the malaria mosquito <it>Anopheles gambiae </it>has been successfully achieved in recent years, and represents a potentially powerful tool for researchers. Tissue-, stage- and sex-specific promoters are essential requirements to support the development of new applications for the transformation technique and potential malaria control strategies. During the <it>Plasmodium </it>lifecycle in the invertebrate host, four major mosquito cell types are involved in interactions with the parasite: hemocytes and fat body cells, which provide humoral and cellular components of the innate immune response, midgut and salivary glands representing the epithelial barriers traversed by the parasite during its lifecycle in the mosquito.</p> <p>Findings</p> <p>We have analyzed the upstream regulatory sequence of the <it>An. gambiae </it>salivary gland-specific <it>apyrase </it>(<it>AgApy</it>) gene in transgenic <it>An. gambiae </it>using a <it>piggyBac </it>transposable element vector marked by a <it>3xP3 </it>promoter:<it>DsRed </it>gene fusion. Efficient germ-line transformation in <it>An. gambiae </it>mosquitoes was obtained and several integration events in at least three different G₀families were detected. <it>LacZ </it>reporter gene expression was analyzed in three transgenic lines/groups, and in only one group was tissue-specific expression restricted to salivary glands.</p> <p>Conclusion</p> <p>Our data describe an efficient genetic transformation of <it>An. gambiae </it>embryos. However, expression from the selected region of the <it>AgApy </it>promoter is weak and position effects may mask tissue- and stage- specific activity in transgenic mosquitoes.</p

Novel cDNAs encoding salivary proteins from the malaria vector Anopheles gambiae

AbstractSeveral genes encoding salivary components of the mosquito Anopheles gambiae were identified using a selective trapping approach. Among these, five corresponded to genes expressed specifically in female glands and their role may possibly be linked to blood-feeding. Our collection included a fourth member of the D7 protein family and two polypeptides that showed weak similarity to anti-coagulants from distantly related species. Moreover, we identified two additional members of a novel group of proteins that we named glandins. The isolation of tissue-specific genes represents a first step toward a deeper molecular analysis of mosquito salivary secretions

Low-resolution genome map of the malaria mosquito Anopheles gambiae

We have microdissected divisions of the Anopheles gambiae polytene chromosomes, digested the DNAs with a restriction enzyme, and PCR-amplified the DNA fragments to generate a set of pooled probes, each corresponding to approximately 2% of the mosquito genome. These divisional probes were shown to have high complexity. Except for those derived from near the centromeres, they hybridize specifically with their chromosomal sites of origin. Thus, they can be used to map cloned DNAs by a dot blot procedure, which is much more convenient than in situ hybridization to polytene chromosomes. We discuss additional potential uses of these probes, such as easier isolation of molecular markers and genes, including those that cross-hybridize with clones available from other insects. It is expected that the probes will substantially accelerate molecular genetic analysis of this most important malaria vector

Low-resolution genome map of the malaria mosquito Anopheles gambiae

We have microdissected divisions of the Anopheles gambiae polytene chromosomes, digested the DNAs with a restriction enzyme, and PCR-amplified the DNA fragments to generate a set of pooled probes, each corresponding to approximately 2% of the mosquito genome. These divisional probes were shown to have high complexity. Except for those derived from near the centromeres, they hybridize specifically with their chromosomal sites of origin. Thus, they can be used to map cloned DNAs by a dot blot procedure, which is much more convenient than in situ hybridization to polytene chromosomes. We discuss additional potential uses of these probes, such as easier isolation of molecular markers and genes, including those that cross-hybridize with clones available from other insects. It is expected that the probes will substantially accelerate molecular genetic analysis of this most important malaria vector

Chromosomal plasticity and evolutionary potential in the malaria vector Anopheles gambiae sensu stricto: insights from three decades of rare paracentric inversions

Background: In the Anopheles gambiae complex, paracentric chromosomal inversions are nonrandomly distributed along the complement: 18/31 (58%) of common polymorphic inversions are on chromosome arm 2R, which represents only ~30% of the complement. Moreover, in An.gambiae sensu stricto, 6/7 common polymorphic inversions occur on 2R. Most of these inversions are considered markers of ecological adaptation that increase the fitness of the carriers of alternative karyotypes in contrasting habitats. However, little is known about the evolutionary forces responsible for their origin and subsequent establishment in field populations. Results: Here, we present data on 82 previously undescribed rare chromosomal inversions (RCIs) recorded during extensive field sampling in 16 African countries over a 30 year period, which may shed light on the dynamics of chromosomal plasticity in An. gambiae. We analyzed breakpoint distribution, length, and geographic distribution of RCIs, and compared these measures to those of the common inversions. We found that RCIs, like common inversions, are disproportionately clustered on 2R, which may indicate that this arm is especially prone to breakages. However, contrasting patterns were observed between the geographic distribution of common inversions and RCIs. RCIs were equally frequent across biomes and on both sides of the Great Rift Valley (GRV), whereas common inversions predominated in arid ecological settings and west of the GRV. Moreover, the distribution of RCI lengths followed a random pattern while common inversions were significantly less frequent at shorter lengths. Conclusion: Because 17/82 (21%) RCIs were found repeatedly at very low frequencies – at the same sampling location in different years and/or in different sampling locations – we suggest that BMC Evolutionary Biology 2008, 8:309 http://www.biomedcentral.com/1471-2148/8/309 RCIs are subject mainly to drift under unperturbed ecological conditions. Nevertheless, RCIs may represent an important reservoir of genetic variation for An. gambiae in response to environmental changes, further testifying to the considerable evolutionary potential hidden within this pan-African malaria vector

Hyaluronic acid alone versus hyaluronic acid associated with adelmidrol for intra-articular treatment of knee osteoarthritis: a long-term follow-up

Background: Hyaluronic acid (HA) has been used for many years for intra-articular treatment of knee osteoarthritis with satisfactory results. HA associated with Adelmidrol – an anti-neuroinflammatory compound – have been only recently introduced in orthopedic clinical practice with good preliminary results. Objective: To investigate whether HA associated with Adelmidrol provides better results than HA alone. Methods: Two cohorts of patients with moderate knee osteoarthritis were treated. Cohort 1 received 5 weekly intra-articular injections of HA during 2017 while Cohort 2, 4 weekly intra-articular injections of HA associated with Adelmidrol during 2018. The patients of the two Cohorts were assessed by WOMAC scale, SF-12 questionnaire and PGIC scale at 1 week (T0), 6 months (T1), 1 year (T2), and 2 years (T3) after the end of treatment. All the data were statistically analyzed. A p-value of &lt;0.05 was considered statistically significant. Results: According to the WOMAC Scale Cohort 1 had higher mean scores than Cohort 2 at each follow-up time, with a statistically significant difference between the two cohorts at T3 (p&lt;0.03) for all the WOMAC components, except for Stiffness. WOMAC Total mean score worsened statistically significantly only in Cohort 1, from T1 to T3 (T2 vs T1: p=0.0033; T3 vs T2: p=0.0007). The same happened for WOMAC Physical Function (T2 vs T1: p=0.0146; T3 vs T2: p=0.0046) and WOMAC Pain (T2 vs T1: p=0.0004; T3 vs T2: p=0.0002). WOMAC Stiffness worsened statistically significantly in Cohort 1 from T2 to T3 (T3 vs T2: p=0.0041), while in Cohort 2 no change on WOMAC scale was statistically significant at any time-point, for any components. The mean scores of the SF-12 questionnaire were better in Cohort 2 than in Cohort 1 at each follow-up time for both the Physical and the Mental components, with a statistically significant difference between the two groups for the latter, at T0 (p=0.0001). In both cohorts the mean score of the Physical component decreased from T0 to T3, but the difference was not statistically significant between the two groups (p=0.25). The mean score of the Mental component slightly increased in Cohort 1 and decreased in Cohort 2, without statistically significant differences between the two groups at any time-point. PGIC showed that Cohort 2 scored significantly better than Cohort 1 at T3 (p=0.0336). Conclusions: Overall, HA associated with Adelmidrol gave better long-term results than HA alone

Genetic complexity of Plasmodium falciparum in two ethnic groups of Burkina Faso with marked differences in susceptibility to malaria.

We have characterized Plasmodium falciparum genotypes among the Mossi and Fulani sympatric ethnic groups in villages in Burkina Faso during the rainy season. Differences in clinical malaria presentation and in immune responses to malaria occur between the two groups. Asexual parasite rate, density, and gametocyte rate were higher among the Mossi than the Fulani. There was no difference in frequencies of alleles of the P. falciparum merozoite surface protein 1 (msp-1), msp-2, and glutamate-rich protein (glurp) genes among the parasites in each group. However, there were significant differences in the mean number of P. falciparum clones in the two populations, with there being more in the Mossi than in the Fulani. This effect was especially marked in older children. These differences can most probably be attributed to genetic differences in immune responsiveness to malaria between the two ethnic groups

Thermodynamics and statistical mechanics of frozen systems in inherent states

We discuss a Statistical Mechanics approach in the manner of Edwards to the ``inherent states'' (defined as the stable configurations in the potential energy landscape) of glassy systems and granular materials. We show that at stationarity the inherent states are distributed according a generalized Gibbs measure obtained assuming the validity of the principle of maximum entropy, under suitable constraints. In particular we consider three lattice models (a diluted Spin Glass, a monodisperse hard-sphere system under gravity and a hard-sphere binary mixture under gravity) undergoing a schematic ``tap dynamics'', showing via Monte Carlo calculations that the time average of macroscopic quantities over the tap dynamics and over such a generalized distribution coincide. We also discuss about the general validity of this approach to non thermal systems.Comment: 10 pages, 16 figure