Parasitism byEphedrus cerasicola [Hym.: Aphidiidae] developing in different stages ofMyzus persicae [Hom.: Aphididae]

The parasitoidEphedrus cerasicola Stary oviposited in the 4 nymphal instars and in newly moulted apterous adults ofMyzus persicae (Sulzer). Development and reproduction of unparasitized and parasitized aphids at 21°C were compared. Unparasitized aphids developed to adults in 6.5 days and started to reproduce after 7 days. Longevity varied between 7 and 42 days. Net reproductive rate (R₀) was 40.7. In contrast to older nymphs, aphids parasitized in the 1 st instar almost never reached the adult stage before mummification. Aphids parasitized in 2nd, 3rd and 4th instar and as newly moulted adults produced respectively 0.07 %, 2 %, 23 % and 32 % of offspring produced by unparasitized aphids. Corresponding reproductive periods were 1, 1.4, 3 and 4 days. Host age at parasitization had a slight effect on the parasitoid's developmental rate and had no effect on egg or pupal survival, or on the sex ratio of the emerging parasitoids.     

Lipidomics of familial longevity

Middle‐aged offspring of nonagenarians, as compared to their spouses (controls), show a favorable lipid metabolism marked by larger LDL particle size in men and lower total triglyceride levels in women. To investigate which specific lipids associate with familial longevity, we explore the plasma lipidome by measuring 128 lipid species using liquid chromatography coupled to mass spectrometry in 1526 offspring of nonagenarians (59 years ± 6.6) and 675 (59 years ± 7.4) controls from the Leiden Longevity Study. In men, no significant differences were observed between offspring and controls. In women, however, 19 lipid species associated with familial longevity. Female offspring showed higher levels of ether phosphocholine (PC) and sphingomyelin (SM) species (3.5–8.7%) and lower levels of phosphoethanolamine PE (38:6) and long‐chain triglycerides (TG) (9.4–12.4%). The association with familial longevity of two ether PC and four SM species was independent of total triglyceride levels. In addition, the longevity‐associated lipid profile was characterized by a higher ratio of monounsaturated (MUFA) over polyunsaturated (PUFA) lipid species, suggesting that female offspring have a plasma lipidome less prone to oxidative stress. Ether PC and SM species were identified as novel longevity markers in females, independent of total triglycerides levels. Several longevity‐associated lipids correlated with a lower risk of hypertension and diabetes in the Leiden Longevity Study cohort. This sex‐specific lipid signature marks familial longevity and may suggest a plasma lipidome with a better antioxidant capacity, lower lipid peroxidation and inflammatory precursors, and an efficient beta‐oxidation function.     

High Content Analysis of Primary Macrophages Hosting Proliferating Leishmania Amastigotes: Application to Anti-leishmanial Drug Discovery

Background/Objectives

Human leishmaniases are parasitic diseases causing severe morbidity and mortality. No vaccine is available and numerous factors limit the use of current therapies. There is thus an urgent need for innovative initiatives to identify new chemotypes displaying selective activity against intracellular Leishmania amastigotes that develop and proliferate inside macrophages, thereby causing the pathology of leishmaniasis.

Methodology/Principal Findings

We have developed a biologically sound High Content Analysis assay, based on the use of homogeneous populations of primary mouse macrophages hosting Leishmania amazonensis amastigotes. In contrast to classical promastigote-based screens, our assay more closely mimics the environment where intracellular amastigotes are growing within acidic parasitophorous vacuoles of their host cells. This multi-parametric assay provides quantitative data that accurately monitors the parasitic load of amastigotes-hosting macrophage cultures for the discovery of leishmanicidal compounds, but also their potential toxic effect on host macrophages. We validated our approach by using a small set of compounds of leishmanicidal drugs and recently published chemical entities. Based on their intramacrophagic leishmanicidal activity and their toxicity against host cells, compounds were classified as irrelevant or relevant for entering the next step in the drug discovery pipeline.

Conclusions/Significance

Our assay represents a new screening platform that overcomes several limitations in anti-leishmanial drug discovery. First, the ability to detect toxicity on primary macrophages allows for discovery of compounds able to cross the membranes of macrophage, vacuole and amastigote, thereby accelerating the hit to lead development process for compounds selectively targeting intracellular parasites. Second, our assay allows discovery of anti-leishmanials that interfere with biological functions of the macrophage required for parasite development and growth, such as organelle trafficking/acidification or production of microbicidal effectors. These data thus validate a novel phenotypic screening assay using virulent Leishmania amastigotes growing inside primary macrophage to identify new chemical entities with bona fide drug potential.     

Mapping the Binding Interface between an HIV-1 Inhibiting Intrabody and the Viral Protein Rev

HIV-1 Rev is the key protein in the nucleocytoplasmic export and expression of the late viral mRNAs. An important aspect for its function is its ability to multimerize on these mRNAs. We have recently identified a llama single-domain antibody (Nb₁₉₀) as the first inhibitor targeting the Rev multimerization function in cells. This nanobody is a potent intracellular antibody that efficiently inhibits HIV-1 viral production. In order to gain insight into the Nb₁₉₀-Rev interaction interface, we performed mutational and docking studies to map the interface between the nanobody paratope and the Rev epitope. Alanine mutants of the hyper-variable domains of Nb₁₉₀ and the Rev multimerization domains were evaluated in different assays measuring Nb₁₉₀-Rev interaction or viral production. Seven residues within Nb₁₉₀ and five Rev residues are demonstrated to be crucial for epitope recognition. These experimental data were used to perform docking experiments and map the Nb₁₉₀-Rev structural interface. This Nb₁₉₀-Rev interaction model can guide further studies of the Nb₁₉₀ effect on HIV-1 Rev function and could serve as starting point for the rational development of smaller entities binding to the Nb₁₉₀ epitope, aimed at interfering with protein-protein interactions of the Rev N-terminal domain.     

PCSK9 SNP rs11591147 is associated with low cholesterol levels but not with cognitive performance or noncardiovascular clinical events in an elderly population

Proprotein convertase subtilisin-like/kexin type 9 (PCSK9) is a protein involved in LDL-cholesterol metabolism. The single-nucleotide polymorphism (SNP) rs11591147 has been associated with lower LDL-cholesterol and a lower risk of coronary heart disease. Because PCSK9 has high affinity to the LDL receptor, inhibiting PCSK9 is a testable therapeutic target for lipid-lowering therapy. Currently, several approaches to inhibit PCSK9 are under development, but it is unknown what the effects of those inhibitors will be on cognition or noncardiovascular clinical events. In this study, we assessed the association between rs11591147 and cognitive performance, activities of daily living (ADL), and noncardiovascular clinical events within 5,777 participants of the PROspective Study of Pravastatin in the Elderly at Risk (PROSPER). Rs11591147 was associated with 10% to 16% lower LDL cholesterol levels (P = 3.62 × 10⁻¹²), but was not associated with cognitive performance, ADL, or noncardiovascular clinical events in the PROSPER study. Our findings suggest that lower cholesterol levels due to genetic variation in the PCSK9 gene are not associated with cognitive performance, functional status, or noncardiovascular clinical events.     

Intramyocellular lipid content in human skeletal muscle

Fat can be stored not only in adipose tissue but also in other tissues such as skeletal muscle. Fat droplets accumulated in skeletal muscle [intramyocellular lipids (IMCLs)] can be quantified by different methods, all with advantages and drawbacks. Here, we briefly review IMCL quantification methods that use biopsy specimens (biochemical quantification, electron microscopy, and histochemistry) and non-invasive alternatives (magnetic resonance spectroscopy, magnetic resonance imaging, and computed tomography). Regarding the physiological role, it has been suggested that IMCL serves as an intracellular source of energy during exercise. Indeed, IMCL content decreases during prolonged submaximal exercise, and analogously to glycogen, IMCL content is increased in the trained state. In addition, IMCL content is highest in oxidative, type 1 muscle fibers. Together, this, indeed, suggests that the IMCL content is increased in the trained state to optimally match fat oxidative capacity and that it serves as readily available fuel. However, elevation of plasma fatty acid levels or dietary fat content also increases IMCL content, suggesting that skeletal muscle also stores fat simply if the availability of fatty acids is high. Under these conditions, the uptake into skeletal muscle may have negative consequences on insulin sensitivity. Besides the evaluation of the various methods to quantify IMCLs, this perspective describes IMCLs as valuable energy stores during prolonged exercise, which, however, in the absence of regular physical activity and with overconsumption of fat, can have detrimental effects on muscular insulin sensitivity.     

Familial longevity is marked by enhanced insulin sensitivity

Insulin resistance is a risk factor for various age-related diseases. In the Leiden Longevity study, we recruited long-lived siblings and their offspring. Previously, we showed that, compared to controls, the offspring of long-lived siblings had a better glucose tolerance. Here, we compared groups of offspring from long-lived siblings and controls for the relation between insulin and glucose in nonfasted serum (n = 1848 subjects) and for quantitation of insulin action using a two-step hyperinsulinemic-euglycemic clamp (n = 24 subjects). Groups of offspring and controls were similar with regard to sex distribution, age, and body mass index. We observed a positive bi-phasic linear relationship between ln (insulin) levels and nonfasted glucose with a steeper slope from 10.7mU L(-1) insulin onwards in controls compared to offspring (P = 0.02). During the clamp study, higher glucose infusion rate was required to maintain euglycemia during high-dose insulin infusion (P = 0.036) in offspring, reflecting higher whole-body insulin sensitivity. After adjustment for sex, age, and fat mass, the insulin-mediated glucose disposal rate (GDR) was higher in offspring than controls (42.5 ± 2.7 vs. 33.2 ± 2.7 micromol kg(-1) min(-1) , mean ± SE, P = 0.025). The insulin-mediated suppression of endogenous glucose production and lipolysis did not differ between groups (all P > 0.05). Furthermore, GDR was significantly correlated with the mean age of death of the parents. In conclusion, offspring from long-lived siblings are marked by enhanced peripheral glucose disposal. Future research will focus on identifying the underlying biomolecular mechanisms, with the aim to promote health in old age.     

Effects of aerobic exercise therapy and cognitive behavioural therapy on functioning and quality of life in amyotrophic lateral sclerosis: protocol of the FACTS-2-ALS trial

Background

Neuropathic pain must be correctly diagnosed for optimal treatment. The questionnaire named Neuropathic Pain Symptom Inventory (NPSI) was developed in its original French version to evaluate the different symptoms of neuropathic pain. We hypothesized that the NPSI might also be used to differentiate neuropathic from non-neuropathic pain.

Methods

We translated the NPSI into German using a standard forward-backward translation and administered it in a case-control design to patients with neuropathic (n = 68) and non-neuropathic pain (headache and osteoarthritis, n = 169) to validate it and to analyze its discriminant properties, its sensitivity to change, and to detect neuropathic pain subgroups with distinct profiles.

Results

Using a sum score (the NPSI-G score), we found sensitivity to change (r between 0.37 and 0.5 for pain items of the graded chronic pain scale) and could distinguish between neuropathic and other pain on a group basis, but not for individual patients. Post hoc development of a discriminant score with optimized diagnostic properties to distinguish neuropathic pain from non-neuropathic pain resulted in an instrument with high sensitivity (91%) and acceptable specificity (70%). We detected six different pain profiles in the patient group with neuropathic pain; three profiles were found to be distinct.

Conclusions

The NPSI-G potentially combines the properties of a diagnostic tool and an instrument to identify subtypes of neuropathic pain.     

Functional responses to prey density ofEphedrus cerasicola [Hym.: Aphidiidae], an aphidiid parasitoid ofMyzus persicae [Hom.: Aphididae]

The functional responses of the parasitoidEphedrus cerasicola Starý parasitizingMyzus persicae (Sulzer) on a paprika plant, were studied for single females at different host densities (1 to 120 aphids per 1.8 dm³) during 3 different time periods (1 hr, 6 hrs, 24 hrs) at 21 °C. The response during 1 hr was curvilinear and is described by the random parasite equation according toRogers (1972). A maximum of 5–6 aphids could be parasitized during 1 hr. After 6 hrs, the response was linear, although it showed a slight sigmoid trend. In the 24-hr experiment, the response was curvilinear, but due to high rate of parasitization together with superparasitism no curve was estimated. The area of discovery was calculated for all densities and exposure times and ranged from 0.15 to 0.79 (1 hr), 0.69 to 1.96 (6 hrs), and 0.81 to 3.21 (24 hrs).