Immune Responses to ESAT-6 and CFP-10 by FASCIA and Multiplex Technology for Diagnosis of M. tuberculosis Infection; IP-10 Is a Promising Marker

Background

There is a need for reliable markers to diagnose active and latent tuberculosis (TB). The interferon gamma release assays (IGRAs) are compared to the tuberculin skin test (TST) more specific, but cannot discriminate between recent or remote TB infection. Here the Flow-cytometric Assay for Specific Cell-mediated Immune-response in Activated whole blood (FASCIA), which quantifies expanded T-lymphoblasts by flow-cytometric analysis after long-term antigen stimulation of whole blood, is combined with cytokine/chemokine analysis in the supernatant by multiplex technology for diagnosis of Mycobacterium tuberculosis (Mtb) infection.

Methods and Findings

Consecutive patients with suspected TB (n = 85), with microbiologically verified active pulmonary TB (n = 33), extra pulmonary TB (n = 21), clinical TB (n = 11), presumed latent TB infection (LTBI) (n = 23), patients negative for TB (n = 8) and 21 healthy controls were studied. Blood samples were analyzed with FASCIA and multiplex technology to determine and correlate proliferative responses and the value of 14 cytokines for diagnosis of Mtb infection: IFN- γ, IL-2, TNF-α, IP-10, IL-12, IL-6, IL-4, IL-5, IL-13, IL-17, MIP-1β, GM-CSF, IFN-α2 and IL-10. Cytokine levels for IFN-γ, IP-10, MIP-1β, IL-2, TNF-α, IL-6, IL-10, IL-13 and GM-CSF were significantly higher after stimulation with the Mtb specific antigens ESAT-6 and CFP-10 in patients with active TB compared to healthy controls (p<0.05) and correlated with proliferative responses. IP-10 was positive in all patients with verified TB, if using a combination of ESAT-6 and CFP-10 and was the only marker significantly more sensitive in detecting active TB then IFN-γ (p = 0.012). Cytokine responses in patients with active TB were more frequent and detected at higher levels than in patients with LTBI.

Conclusions

IP-10 seems to be an important marker for diagnosis of active and latent TB. Patients with active TB and LTBI responded with similar cytokine profiles against TB antigens but proliferative and cytokine responses were generally higher in patients with active TB.     

Trans-Atlantic dispersal and large-scale lack of genetic structure in the circumpolar, arctic-alpine sedge Carex bigelowii s. l. (Cyperaceae)

Paradoxically, several of the ecologically most important plant groups in the Arctic are little understood in terms of taxonomy and biogeographic history. The circumpolar Carex bigelowii s. l. (Cyperaceae) is abundant in the Arctic and is one of the most complicated arctic plant groups. While its ecology and population genetics have been extensively studied, its taxonomy is largely unexplored. We analyzed the large-scale geographical structuring of amplified fragment length polymorphisms (AFLPs) covering most of the distribution range. We detected high levels of genetic variation, most (66%) within populations, and a fairly weak genetic structure. Only the Central Asian populations, referred to as C. orbicularis, were strongly divergent. For the remaining populations, Bayesian clustering separated three distinct clusters (one European, one amphi-Atlantic, and one broadly amphi-Beringian), probably reflecting different major glacial refugia and recent transoceanic dispersal. The isolated central European populations were most closely related to those from a larger distribution area in northern Europe. Differences in genetic diversity suggest that the Alpine and Tatra populations have experienced strong bottlenecks, whereas the Krkono?e population may have been part of a continuous distribution area during the cold stages of the Pleistocene. Finally, we discuss the relevance of our results for a uniform, range-wide taxonomic concept.     

Diet shift of a facultative scavenger, the wolverine, following recolonization of wolves

1. Wolves Canis lupus L. recolonized the boreal forests in the southern part of the Scandinavian peninsula during the late 1990s, but so far there has been little attention to its effect on ecosystem functioning. Wolf predation increases the availability of carcasses of large prey, especially moose Alces alces L., which may lead in turn to a diet switch in facultative scavengers such as the wolverine Gulo gulo L. 2. Using 459 wolverine scats collected during winter-spring 2001-04 for DNA identity and dietary contents, we compared diet inside and outside wolf territories while controlling for potential confounding factors, such as prey density. We tested the hypothesis that wolverine diet shifted towards moose in the presence of wolves, while taking into account possible sexual segregation between the sexes. Occurrence of reindeer, moose and small prey was modelled against explanatory covariates using logistic mixed-effects models. Furthermore, we compared diet composition and breadth among habitats and sexes. 3. Occurrence of reindeer, moose and small prey in the diet varied with prey availability and habitat. As expected, diet contained more moose and less reindeer and small prey in the presence of wolves. Their diet in tundra consisted of 40% reindeer Rangifer tarandus L., 39% moose and 9% rodents. In forest with wolf, their diet shifted to 76% moose, 18% reindeer and 5% rodents; compared to 42% moose, 32% reindeer and 15% rodents in forest without wolf. This diet switch could not be explained by higher moose density in wolf territories. Female diet consisted of more small prey than for males, but there was a tendency for females to use the highly available moose carrion opportunistically and to hunt less on small prey within wolf territories. 4. Our study highlights how wolves increase scavenging opportunities for wolverines, and how sexual differences in diet may also apply to large scavengers. Due to their more restricted home range, female wolverines are forced to rely more on hunting small prey. The relatively high occurrence of wolf kills, however, forms an important food source to wolverines in this area. The recolonization of wolves may therefore have contributed to the consequent recolonization of wolverines into the same area.     

Cardiac-restricted expression of the carboxyl-terminal fragment of GRK3 Uncovers Distinct Functions of GRK3 in regulation of cardiac contractility and growth: GRK3 controls cardiac alpha1-adrenergic receptor responsiveness

G protein-coupled receptor kinase-2 and -3 (GRK2 and GRK3) in cardiac myocytes catalyze phosphorylation and desensitization of different G protein-coupled receptors through specificity controlled by their carboxyl-terminal pleckstrin homology domain. Although GRK2 has been extensively investigated, the function of cardiac GRK3 remains unknown. Thus, in this study cardiac function of GRK3 was investigated in transgenic (Tg) mice with cardiac-restricted expression of a competitive inhibitor of GRK3, i.e. the carboxyl-terminal plasma membrane targeting domain of GRK3 (GRK3ct). Cardiac myocytes from Tg-GRK3ct mice displayed significantly enhanced agonist-stimulated alpha(1)-adrenergic receptor-mediated activation of ERK1/2 versus cardiac myocytes from nontransgenic littermate control (NLC) mice consistent with inhibition of GRK3. Tg-GRK3ct mice did not display alterations of cardiac mass or left ventricular dimensions compared with NLC mice. Tail-cuff plethysmography of 3- and 9-month-old mice revealed elevated systolic blood pressure in Tg-GRK3ct mice versus control mice (3-month-old mice, 136.8 +/- 3.6 versus 118.3 +/- 4.7 mm Hg, p < 0.001), an observation confirmed by radiotelemetric recording of blood pressure of conscious, unrestrained mice. Simultaneous recording of left ventricular pressure and volume in vivo by miniaturized conductance micromanometry revealed increased systolic performance with significantly higher stroke volume and stroke work in Tg-GRK3ct mice than in NLC mice. This phenotype was corroborated in electrically paced ex vivo perfused working hearts. However, analysis of left ventricular function ex vivo as a function of increasing filling pressure disclosed significantly reduced (dP/dt)(min) and prolonged time constant of relaxation (tau) in Tg-GRK3ct hearts at elevated supraphysiological filling pressure compared with control hearts. Thus, inhibition of GRK3 apparently reduces tolerance to elevation of preload. In conclusion, inhibition of cardiac GRK3 causes hypertension because of hyperkinetic myocardium and increased cardiac output relying at least partially on cardiac myocyte alpha(1)-adrenergic receptor hyper-responsiveness. The reduced tolerance to elevation of preload may cause impaired ability to withstand pathophysiological mechanisms of heart failure.     

Environmental phenology and geographical gradients in moose body mass

Intraspecific body mass in ungulates has often been shown to increase with latitude. The biological basis for such latitudinal gradients is, however, poorly known. Here we examined whether satellite-derived indices of environmental phenology, based on the normalised difference vegetation index (NDVI), as well as variables derived from meteorological stations, altitude, and population density, can explain latitudinal gradients and regional variation in body mass of Norwegian moose. The best model gave a considerably better fit than latitude alone, and included all explanatory environmental variables. Accordingly, heavy moose were found in areas with short and intense summers that were followed by long, cold winters, at low altitude relative to the tree-limit, and with low population density relative to the available plant biomass. This relationship was stronger for yearlings than for calves, except for the effect of population density. This indicates that differences in the characteristics of the vegetation quality and environmental phenology, as well as winter harshness and population density, are important factors that shape both the latitudinal and other geographical gradients in moose body mass.     

Plerocercoids of the cestode Diphyllobothrium ditremum in brown trout Salmo trutta: substantial increase in infection after establishment of European minnow Phoxinus phoxinus

This study focuses on plerocercoids of the cestode Diphyllobothrium ditremum in brown trout Salmo trutta from the subalpine lake Øvre Heimdalsvatn in south–central Norway. Salmo trutta was the only fish species in this lake until European minnow Phoxinus phoxinus was registered in 1969. The P. phoxinus population increased substantially in the following years. In contrast with the 1969–1972 period, when plerocercoids of D. ditremum were practically absent in S. trutta, there was a high prevalence and intensity of infection in the 2013 S. trutta samples. Because the life cycle of D. ditremum involves two larval stages, in copepods and salmonids and mature worms in piscivorous birds, such as mergansers and loons, a change in feeding ecology of S. trutta or changes in population densities of copepods, fish or birds might have influenced the infection pattern. No relationships between D. ditremum infection and muscle‐tissue δ¹⁵N signature or Hg concentration were found, indicating that infection is not a result of piscivory or cannibalism. Furthermore, consumption of copepods by S. trutta during summer and autumn was low. On the other hand, the number of piscivorous birds has increased, probably due to the presence of P. phoxinus as a new and numerous prey. An increased number of final D. ditremum hosts may have produced a higher output of cestode eggs, resulting in more infected copepods that in turn are consumed by S. trutta. Indirectly, P. phoxinus may therefore have caused the observed increased infection in S. trutta and thereby imposed further negative effects on S. trutta in high mountain areas.     

Granule containing cells and fibres in the sinus venosus of elasmobranchs

The concentrations of catecholamines in the heart chambers of elasmobranchs were measured by the fluorimetric method of Bertler et al. (1958). Noradrenaline (NA) can be detected in all the chambers, but the sinus venosus is by far the richest in NA. This can either be due to the presence of storage sites for this amine in the sinus wall, or to a transport of amine to the sinus venosus from the anterior chromaffin bodies. The sinus wall contains large numbers of "granule containing cells" and axon-like processes, both with numerous dense-core vesicles of about 1800 A diameter. The dense-core vesicles contain a uranophilic matrix indicating the presence of protein, phospholipids and/or nucleic acid. The reactions failed to demonstrate amine, which may be due to a loss of amine by diffusion, to a relatively low intravesicular amine concentration, or, to the absence of amines in these granule-containing cells and processes. Heavy accumulations of granule-containing processes occur in the subendothelial area. The endothelium contains fenestrae and pores through which granule-containing fibres protrude into the venous cavity. Granule-containing cells are innervated by presumed cholinergic nerve endings. It is suggested that the granule-containing cells and fibres belong to the neurosecretory system with a cholinergic input, releasing the contents of the dense-core vesicles into the blood stream at the level of the venous cavity.