Trypanosoma cruzi infection follow-up in a sylvatic vector of Chagas disease: Comparing early and late stage nymphs

Chagas disease is caused by Trypanosoma cruzi and transmitted by the triatomine Mepraia spinolai in the southwest of South America. Here, we examined the T. cruzi-infection dynamics of field-caught M. spinolai after laboratory feeding, with a follow-up procedure on bug populations collected in winter and spring of 2017 and 2018. Bugs were analyzed twice to evaluate T. cruzi-infection by PCR assays of urine/fecal samples, the first evaluation right after collection and the second 40 days after the first feeding. We detected bugs with: the first sample positive and second negative (+/-), the first sample negative and second positive (-/+), and with both samples positive or negative (+/+; -/-). Bugs that resulted positive on both occasions were the most frequent, with the exception of those collected in winter 2018. Infection rate in spring was higher than winter only in 2018. Early and late stage nymphs presented similar T. cruzi-infection rates except for winter 2017; therefore, all nymphs may contribute to T. cruzi-transmission to humans. Assessment of infection using two samples represents a realistic way to determine the infection a triatomine can harbor. The underlying mechanism may be that some bugs do not excrete parasites unless they are fed and maintained for some time under environmentally controlled conditions before releasing T. cruzi, which persists in the vector hindgut. We suggest that T. cruzi-infection dynamics regarding the three types of positive-PCR results detected by follow-up represent: residual T. cruzi in the rectal lumen (+/-), colonization of parasites attached to the rectal wall (-/+), and presence of both kinds of flagellates in the hindgut of triatomines (+/+). We suggest residual T. cruzi-infections are released after feeding, and result 60–90 days after infection persisting in the rectal lumen after a fasting event, a phenomenon that might vary between contrasting seasons and years.     

The SR Ca2+ ATPase of the Antarctic scallop Adamussium colbecki: cold adaptation and heavy metal effects

Cell calcium is accumulated in intracellular stores by sarco-endoplasmic reticulum Ca²⁺ ATPases functionally interacting with the membrane lipid environment. Cold adaptations of membrane lipids in Antarctic Sea organisms suggest possible adaptive effects also on sarco-endoplasmic reticulum Ca²⁺ ATPases. We investigated the SR Ca²⁺ ATPase of an Antarctic scallop, Adamussium colbecki, by characterising the enzyme activity and studying temperature effects. Ca²⁺ ATPase, assayed by following ATP hydrolysis, was thapsigargin- and vanadate-sensitive, showed maximum activity under 2 μM Ca²⁺, 200 mM KCl and pH 7.2, and had a K _M for ATP of 22 ± 7 μM. Temperature effects showed an Arrhenius inversion between −1.8 and 0°C, indicating cold adaptation, an Arrhenius break at 10°C, and a collapse above 20°C. A. colbecki accumulates high amounts of cadmium in the digestive gland; heavy metal effects on sarco-endoplasmic reticulum Ca²⁺ ATPases were therefore tested, finding an IC₅₀ = 0.9 μM for Hg²⁺ and 3 μM for Cd²⁺. Finally, SDS-PAGE analysis showed a main band at about 100 kDa, which was identified as sarco-endoplasmic reticulum Ca²⁺ ATPase after trypsin digestion, and accounted for 60% total protein. Accepted: 10 December 1998     

Purification and Characterization of Novel Antifungal Compounds from the Sourdough Lactobacillus plantarum Strain 21B

Sourdough lactic acid bacteria were selected for antifungal activity by a conidial germination assay. The 10-fold-concentrated culture filtrate of Lactobacillus plantarum 21B grown in wheat flour hydrolysate almost completely inhibited Eurotium repens IBT18000, Eurotium rubrum FTDC3228, Penicillium corylophilum IBT6978, Penicillium roqueforti IBT18687, Penicillium expansum IDM/FS2, Endomyces fibuliger IBT605 and IDM3812, Aspergillus niger FTDC3227 and IDM1, Aspergillus flavus FTDC3226, Monilia sitophila IDM/FS5, and Fusarium graminearum IDM623. The nonconcentrated culture filtrate of L. plantarum 21B grown in whole wheat flour hydrolysate had similar inhibitory activity. The activity was fungicidal. Calcium propionate at 3 mg ml⁻¹ was not effective under the same assay conditions, while sodium benzoate caused inhibition similar to L. plantarum 21B. After extraction with ethyl acetate, preparative silica gel thin-layer chromatography, and chromatographic and spectroscopic analyses, novel antifungal compounds such as phenyllactic and 4-hydroxy-phenyllactic acids were identified in the culture filtrate of L. plantarum 21B. Phenyllactic acid was contained at the highest concentration in the bacterial culture filtrate and had the highest activity. It inhibited all the fungi tested at a concentration of 50 mg ml⁻¹ except for P. roqueforti IBT18687 and P. corylophilum IBT6978 (inhibitory concentration, 166 mg ml⁻¹). L. plantarum 20B, which showed high antimold activity, was also selected. Preliminary studies showed that phenyllactic and 4-hydroxy-phenyllactic acids were also contained in the bacterial culture filtrate of strain 20B. Growth of A. niger FTDC3227 occurred after 2 days in breads started with Saccharomyces cerevisiae 141 alone or with S. cerevisiae and Lactobacillus brevis 1D, an unselected but acidifying lactic acid bacterium, while the onset of fungal growth was delayed for 7 days in bread started with S. cerevisiae and selected L. plantarum 21B.     

Catecholamines are key modulators of ventricular repolarization patterns in the ball python (Python regius)

Ectothermic vertebrates experience daily changes in body temperature, and anecdotal observations suggest these changes affect ventricular repolarization such that the T-wave in the ECG changes polarity. Mammals, in contrast, can maintain stable body temperatures, and their ventricular repolarization is strongly modulated by changes in heart rate and by sympathetic nervous system activity. The aim of this study was to assess the role of body temperature, heart rate, and circulating catecholamines on local repolarization gradients in the ectothermic ball python (Python regius). We recorded body-surface electrocardiograms and performed open-chest high-resolution epicardial mapping while increasing body temperature in five pythons, in all of which there was a change in T-wave polarity. However, the vector of repolarization differed between individuals, and only a subset of leads revealed T-wave polarity change. RNA sequencing revealed regional differences related to adrenergic signaling. In one denervated and Ringer’s solution–perfused heart, heating and elevated heart rates did not induce change in T-wave polarity, whereas noradrenaline did. Accordingly, electrocardiograms in eight awake pythons receiving intra-arterial infusion of the β-adrenergic receptor agonists adrenaline and isoproterenol revealed T-wave inversion in most individuals. Conversely, blocking the β-adrenergic receptors using propranolol prevented T-wave change during heating. Our findings indicate that changes in ventricular repolarization in ball pythons are caused by increased tone of the sympathetic nervous system, not by changes in temperature. Therefore, ventricular repolarization in both pythons and mammals is modulated by evolutionary conserved mechanisms involving catecholaminergic stimulation.     

Biological Control Potential of Wolbachia -infected Versus Uninfected Wasps: Laboratory and Greenhouse Evaluation of Trichogramma cordubensis and T. deion Strains

The effects of thelytoky-inducing Wolbachia ( &#102 -proteobacteria) on Trichogramma cordubensis and T. deion (Hymenoptera, Trichogrammatidae) were studied in laboratory and greenhouse conditions. One infected (thelytokous, all female) line of each wasp species was compared with its conspecific uninfected (arrhenotokous, sexual) counterpart for several fecundity and dispersal traits. Arrhenotokous lines had a higher fecundity than their thelytokous counterparts, which suggests that Wolbachia negatively affect the fecundity of Trichogramma females. The arrhenotokous females dispersed more in the laboratory than their thelytokous counterparts. In the greenhouse, the opposite effect or no difference between lines was found, indicating that the laboratory set-up used to measure dispersal is not useful to predict relative dispersal of the females in the greenhouse. Calculations show that by releasing 100 adult wasps of both lines, thus including arrhenotokous males in the sexual line, more eggs are parasitized by the thelytokous wasps. Therefore, in spite of their lower individual female fecundity, thelytokous lines have a better potential for biological control than their arrhenotokous counterparts.     

Spectrophotometer-aided evaluation of cyanogenic potential in white clover (Trifolium repens L.)

In comparison with ordinary methods of colorimetric evaluation of cyanogenic potential based on visual evaluation of the alkaline picrate reaction, a spectrophotometer-aided method could be more accurate (since it determines the exact amount of hydrogen cyanide released by the plant material), and less time-consuming as it can be performed on bulk material rather than on a number of individual plants. Ten white clover populations were evaluated by a spectrophotometer-aided method and by two visual evaluation criteria. All methods indicated the presence of large variation between populations. Visual methods gave almost identical results and allowed only for the distinction between cyanogenic and substantially acyanogenic populations. The results were only moderately consistent with those obtained by the spectrophotometer-aided method, which could detect the presence of variation also between cyanogenic populations. The effects of various incubation times (from 4 to 48 h) and of the addition of β-glucosidase on hydrogen cyanide release were also investigated. Comparable results for ranking of populations could be obtained over a range of incubation times, but at least 24 h were needed for a reliable estimation of the hydrogen cyanide produced by plants. The addition of enzyme did not increase the released cyanide. The effect of season and/or conditions of evaluation was marked on mean cyanogenic potential but limited on ranking of populations. Copyright © 2000 John Wiley & Sons, Ltd.     

An enhanced chimeric firefly luciferase-inspired enzyme for ATP detection and bioluminescence reporter and imaging applications

Firefly luciferases, which emit visible light in a highly specific ATP-dependent process, have been adapted for a variety of applications, including gene reporter assays, whole-cell biosensor measurements, and in vivo imaging. We previously reported the approximately 2-fold enhanced activity and 1.4-fold greater bioluminescence quantum yield properties of a chimeric enzyme that contains the N-domain of Photinus pyralis luciferase joined to the C-domain of Luciola italica luciferase. Subsequently, we identified 5 amino acid changes based on L. italica that are the main determinants of the improved bioluminescence properties. Further engineering to enhance thermal and pH stability produced a novel luciferase called PLG2. We present here a systematic comparison of the spectral and physical properties of the new protein with P. pyralis luciferase and demonstrate the potential of PLG2 for use in assays based on the detection of femtomole levels of ATP. In addition, we compared the performance of a mammalian codon-optimized version of the cDNA for PLG2 with the luc2 gene in HEK293T cells. Using an optimized low-cost assay system, PLG2 activity can be monitored in mammalian cell lysates and living cells with 4.4-fold and approximately 3.0-fold greater sensitivity, respectively. PLG2 could be an improved alternative to Promega’s luc2 for reporter and imaging applications.     

Patterns of sexual dimorphism in flight agility in territorial and non-territorial Odonata

Journal of Ethology - Adult dragonflies (Anisoptera) and damselflies (Zygoptera) are amongst the most accomplished flying insects on the planet. The main functions of spatial displacement by flight...     

Blood antioxidant status and urinary levels of catecholamine metabolites in β-thalassemia

It has been reported that iron overload in β-thalassemia leads to an enhanced generation of reactive oxygen species and to oxidative stress. We have studied the oxidant/antioxidant imbalance in the blood of 48 transfusion-dependent β-thalassemic patients (TLP) (17 males, 31 females, 11–22 year), under chelation therapy, and in 40 sex and age matched healthy controls (CTR). Plasma and lymphocyte levels of vitamin E (Vit E), ubiquinol (CoQ₁₀H₂), ubiquinone (CoQ₁₀), plasma concentrations of vitamin A (Vit A), β-carotene, lycopene, vitamin C (Vit C), total thiols, fatty acid patterns of phospholipids (PL-FA), and plasma and urinary markers of lipoperoxidation (TBA-RM, conjugated dienes, and azelaic acid (AZA), as well as the urinary levels of catecholamine and serotonin metabolites, were evaluated by gas chromatography-mass spectrometry (GC-MS), HPLC and spectrophotometry. Routine laboratory blood analyses were performed on the same samples; 39/48 TLP were HCV positive. Blood samples were collected just before transfusion, the 24 h urine samples the day before. Our results clearly showed that a severe oxidative stress occurs in the plasma of TLP in comparison with CTR. In fact, the levels of lipophilic antioxidants and ascorbate were severely depleted: CoQ₁₀H₂ (-62.5%), total CoQ₁₀ (-35.1%), Vit E (-43.8%, β-carotene (-31.1%), lycopene (-63.7%), Vit A (-35.9%), Vit C (-23.1%). The impairment of the antioxidant status was associated with elevated plasma levels of by-products of lipoperoxidation and urinary concentrations of catecholamine metabolites and of AZA, indicating a high degree of both neurological stress and lipoperoxidation. A significant positive correlation was found between vitamin E and non-transferrin-bound iron (NTBI) (r = -0.81; p < 0.001), while no correlation was found between antioxidant depletion and ferritin serum levels, average blood consumption, or the presence of clinical complications. The administration of selective antioxidants along with an appropriate diet might represent a promising way of counteracting oxidative damage and its deleterious effects on the progression of the disease.