Sedimentary Metal Bioavailability Determined By the Digestive Constraints of Marine Deposit Feeders: Gut Retention Time and Dissolved Amino Acids

Contaminant metals bound to sediments are subject to considerable solubilization during passage of the sediments through the digestive systems of deposit feeders. We examined the kinetics of this process, using digestive fluids extracted from deposit feeders Arenicola marina and Parastichopus californicus and then incubated with contaminated sediments. Kinetics are complex, with solubilization followed occasionally by readsorption onto the sediment. In general, solubilization kinetics are biphasic, with an initial rapid step followed by a slower reaction. For many sediment-organism combinations, the reaction will not reach a steady state or equilibrium within the gut retention time (GRT) of the organisms, suggesting that metal bioavailability in sediments is a time-dependent parameter. Experiments with commercial protein solutions mimic the kinetic patterns observed with digestive fluids, which corroborates our previous study that complexation by dissolved amino acids (AA) in digestive fluids leads to metal solubilization (Chen & Mayer 1998b; Environ Sci Technol 32:770-778). The relative importance of the fast and slow reactions appears to depend on the ratio of ligands in gut fluids to the amount of bound metal in sediments. High ligand to solid metal ratios result in more metals released in fast reactions and thus higher lability of sedimentary metals. Multiple extractions of a sediment with digestive fluid of A. marina confirm the potential importance of incomplete reactions within a single deposit-feeding event, and make clear that bioavailability to a single animal is Likely different from that to a community of organisms. The complex kinetic patterns lead to the counterintuitive prediction that toxification of digestive enzymes by solubilized metals will occur more readily in species that dissolve less metals.

Digestive morphophysiology of Gryllodes sigillatus (Orthoptera: Gryllidae)

The evolution of the digestive system in the Order Orthoptera is disclosed from the study of the morphophysiology of the digestive process in its major taxa. This paper deals with a cricket representing the less known suborder Ensifera Most amylase and trypsin activities occur in crop and caeca. respectively. Maltase and aminopeptidase are found in soluble and membrane-bound forms in caeca, with aminopeptidase also occurring in ventriculus. Amaranth was orally fed to Gryllodes sigillatus adults or injected into their haemolymph. The experiments were performed with starving and feeding insects with identical results. Following feeding of the dye the luminal side of the most anterior ventriculus (and in lesser amounts the midgut caeca) became heavily stained. In injected insects, the haemal side of the most posterior ventriculus was stained This suggested that the anterior ventriculus is the main site of water absorption (the caeca is a secondary one). whereas the posterior ventriculus secretes water into the gut. Thus, a putative counter-current flux of fluid from posterior to anterior ventriculus may propel digestive enzyme recycling. This was confirmed by the finding that digestive enzymes are excreted at a low rate. The fine structure of midgut caeca and ventriculus cells revealed that they have morphological features that may be related to their involvement in secretion (movement from cell to lumen) and absorption (movement from lumen to cell) of fluids. Furthermore, morphological data showed that both merocrine and apocrine secretory mechanisms occur in midgut cells. The results showed that cricket digestion differs from that in grasshopper in having (1) more membrane-bound digestive enzymes; (2) protein digestion slightly displaced toward the ventriculus; (3) midgut fluxes, and hence digestive enzyme recycling, in both starved and fed insects. (C) 2009 Elsevier Ltd. All rights reserved.

Peritrophic membrane role in enhancing digestive efficiency Theoretical and experimental models

The pentrophic membrane (PM) is an anatomical structure surrounding the food bolus in most insects. Rejecting the idea that PM has evolved from coating mucus to play the same protective role as it, novel functions were proposed and experimentally tested. The theoretical principles underlying the digestive enzyme recycling mechanism were described and used to develop an algorithm to calculate enzyme distributions along the midgut and to infer secretory and absorptive sites. The activity of a Spodoptera frugiperda microvillar aminopeptidase decreases by 50% if placed in the presence of midgut contents. S. frugiperda trypsin preparations placed into dialysis bags in stirred and unstirred media have activities of 210 and 160%, respectively, over the activities of samples in a test tube. The ectoperitrophic fluid (EF) present in the midgut caeca of Rhynchosciara americana may be collected. If the enzymes restricted to this fluid are assayed in the presence of PM contents (PMC) their activities decrease by at least 58%. The lack of PM caused by calcofluor feeding impairs growth due to an increase in the metabolic cost associated with the conversion of food into body mass. This probably results from an increase in digestive enzyme excretion and useless homeostatic attempt to reestablish destroyed midgut gradients. The experimental models support the view that PM enhances digestive efficiency by: (a) prevention of non-specific binding of undigested material onto cell Surface; (b) prevention of excretion by allowing enzyme recycling powered by an ectoperitrophic counterflux of fluid; (c) removal from inside PM of the oligomeric molecules that may inhibit the enzymes involved in initial digestion; (d) restriction of oligomer hydrolases to ectoperitrophic space (ECS) to avoid probable partial inhibition by non-dispersed undigested food. Finally,PM functions are discussed regarding insects feeding on any diet. (C) 2008 Elsevier Ltd. All rights reserved.

Comparative study of digestive cells in different midgut regions of stingless bees (Hymenoptera: Apidae: Meliponinae)

Ultrastructure of the digestive cells was analyzed in three midgut regions (anterior, middle and posterior) of stingless bees. Variations occurs in the presence of lipid inclusions in the cells from posterior midgut and presence of double-membraned vesicles associated to microvilli in the anterior midgut. However, basal plasmic membrane infoldings and augmentation of surface area achieved by microvilli ale very similar in all midgut regions. These results not supported the existence of fluid fluxes in the ectoperitrophic space and suggest that digestive cells in stingless bees are polifunctional, that is, there is not midgut region specialized in secretion ol absorption as observed in other insects.

A comparative study of digestive cells in different midgut regions of stingless bees (Hymenoptera: Apidae: Meliponinae)

Ultrastructure of the digestive cells was analyzed in three midgut regions (anterior, middle and posterior) of stingless bees. Variations occurs in the presence of lipid inclusions in the cells from posterior midgut and presence of double-membraned vesicles associated to microvilli in the anterior midgut. However, basal plasmic membrane infoldings and augmentation of surface area achieved by microvilli are very similar in all midgut regions. These results not supported the existence of fluid fluxes in the ectoperitrophic space and suggest that digestive cells in stingless bees are polifunctional, that is, there is not midgut region specialized in secretion or absorption as observed in other insects.

Structures and Concentrations of Surfactants in Gut Fluid of the Marine Polychaete Arenicola Marina

Marine invertebrate deposit feeders secrete surfactants into their gut fluid in concentrations sufficient to induce micelle formation, enhancing solubilization of sedimentary lipids. We isolated and identified 3 related surfactant molecules from the deposit-feeding polychaete lugworm Arenicola marina. Surfactants were isolated and separated by a combination of solvent extraction and thin-layer and gas chromatography. Identification was performed using mass and infrared spectrometry, coupled to various derivatization and hydrolysis reactions. A. marina produces a mixture of related yet distinct anionic surfactants composed of branched, C9, saturated and unsaturated fatty acids that are amide linked to leucine or glycine residues, showing some similarity to crustacean surfactants. The critical micelle concentration of the mixture of these surfactants in gut fluid was about 2 mM, and total concentrations ranged from 5.5 to 19.5 mM. The hydrophilic amide linkage helps to explain previous observations that gut surfactants do not adsorb onto sediment transiting the gut.

Growth response of broilers to lysine levels and hydrolyzed porcine digestive mucosa (Palbio) inclusion in diet from 1 to 21 d of age

Palbio (PAL, Palbio 50 RD, Bioibérica, Spain) is a protein concentrate based on hydrolyzed porcine digestive mucosa dried under a fluid bed system over a soybean carrier, currently used in piglet feeds. The digestibility of PAL is very high and the product may be an excellent source of protein for young chicks. An experiment was conducted with 1,280 straight-run one-d-old Ross 308 chicks to evaluate the growth response of broilers to dietary inclusion of PAL.

Digestive Peptidase Evolution In Holometabolous Insects Led To A Divergent Group Of Enzymes In Lepidoptera.

Trypsins and chymotrypsins are well-studied serine peptidases that cleave peptide bonds at the carboxyl side of basic and hydrophobic l-amino acids, respectively. These enzymes are largely responsible for the digestion of proteins. Three primary processes regulate the activity of these peptidases: secretion, precursor (zymogen) activation and substrate-binding site recognition. Here, we present a detailed phylogenetic analysis of trypsins and chymotrypsins in three orders of holometabolous insects and reveal divergent characteristics of Lepidoptera enzymes in comparison with those of Coleoptera and Diptera. In particular, trypsin subsite S1 was more hydrophilic in Lepidoptera than in Coleoptera and Diptera, whereas subsites S2-S4 were more hydrophobic, suggesting different substrate preferences. Furthermore, Lepidoptera displayed a lineage-specific trypsin group belonging only to the Noctuidae family. Evidence for facilitated trypsin auto-activation events were also observed in all the insect orders studied, with the characteristic zymogen activation motif complementary to the trypsin active site. In contrast, insect chymotrypsins did not seem to have a peculiar evolutionary history with respect to their mammal counterparts. Overall, our findings suggest that the need for fast digestion allowed holometabolous insects to evolve divergent groups of peptidases with high auto-activation rates, and highlight that the evolution of trypsins led to a most diverse group of enzymes in Lepidoptera.

Human Herpesvirus Infections Of The Central Nervous System: Laboratory Diagnosis Based On Dna Detection By Nested Pcr In Plasma And Cerebrospinal Fluid Samples.

Infections of the central nervous systems (CNS) present a diagnostic problem for which an accurate laboratory diagnosis is essential. Invasive practices, such as cerebral biopsy, have been replaced by obtaining a polymerase chain reaction (PCR) diagnosis using cerebral spinal fluid (CSF) as a reference method. Tests on DNA extracted from plasma are noninvasive, thus avoiding all of the collateral effects and patient risks associated with CSF collection. This study aimed to determine whether plasma can replace CSF in nested PCR analysis for the detection of CNS human herpesvirus (HHV) diseases by analysing the proportion of patients whose CSF nested PCR results were positive for CNS HHV who also had the same organism identified by plasma nested PCR. In this study, CSF DNA was used as the gold standard, and nested PCR was performed on both types of samples. Fifty-two patients with symptoms of nervous system infection were submitted to CSF and blood collection. For the eight HHV, one positive DNA result-in plasma and/or CSF nested PCR-was considered an active HHV infection, whereas the occurrence of two or more HHVs in the same sample was considered a coinfection. HHV infections were positively detected in 27/52 (51.9%) of the CSF and in 32/52 (61.5%) of the plasma, difference not significant, thus nested PCR can be performed on plasma instead of CSF. In conclusion, this findings suggest that plasma as a useful material for the diagnosis of cases where there is any difficulty to perform a CSF puncture.

Cloning, purification and comparative characterization of two digestive lysozymes from Musca domestica larvae

cDNA coding for two digestive lysozymes (MdL1 and MdL2) of the Musca domestica housefly was cloned and sequenced. MdL2 is a novel minor lysozyme, whereas MdL1 is the major lysozyme thus far purified from M. domestica midgut. MdL1 and MdL2 were expressed as recombinant proteins in Pichia pastoris, purified and characterized. The lytic activities of MdL1 and MdL2 upon Micrococcus lysodeikticus have an acidic pH optimum (4.8) at low ionic strength (μ = 0.02), which shifts towards an even more acidic value, pH 3.8, at a high ionic strength (μ = 0.2). However, the pH optimum of their activities upon 4-methylumbelliferyl N-acetylchitotrioside (4.9) is not affected by ionic strength. These results suggest that the acidic pH optimum is an intrinsic property of MdL1 and MdL2, whereas pH optimum shifts are an effect of the ionic strength on the negatively charged bacterial wall. MdL2 affinity for bacterial cell wall is lower than that of MdL1. Differences in isoelectric point (pI) indicate that MdL2 (pI = 6.7) is less positively charged than MdL1 (pI = 7.7) at their pH optima, which suggests that electrostatic interactions might be involved in substrate binding. In agreement with that finding, MdL1 and MdL2 affinities for bacterial cell wall decrease as ionic strength increases.

Central nitrergic system regulation of neuroendocrine secretion, fluid intake and blood pressure induced by angiotensin-II

Background: Nitric oxide (NO) synthesis has been described in several circumventricular and hypothalamic structures in the central nervous system that are implicated in mediating central angiotensin-II (ANG-II) actions during water deprivation and hypovolemia. Neuroendocrine and cardiovascular responses, drinking behavior, and urinary excretions were examined following central angiotensinergic stimulation in awake freely-moving rats pretreated with intracerebroventricular injections of N omega-nitro-L-arginine methyl ester (L-NAME, 40 mu g), an inhibitor of NO synthase, and L-arginine (20 ug), a precursor of NO. Results: Injections of L-NAME or ANG-II produced an increase in plasma vasopressin (VP), oxytocin (OT) and atrial natriuretic peptide (ANP) levels, an increase in water and sodium intake, mean arterial blood pressure and sodium excretion, and a reduction of urinary volume. L-NAME pretreatment enhanced the ANG-II response, while L-arginine attenuated VP and OT release, thirst, appetite for sodium, antidiuresis, and natriuresis, as well as pressor responses induced by ANG-II. Discussion and conclusion: Thus, the central nitrergic system participates in the angiotensinergic responses evoked by water deprivation and hypovolemia to refrain neurohypophysial secretion, hydromineral balance, and blood pressure homeostasis.

Effects of Laser on the Synovial Fluid in the Inflammatory Process of the Knee Joint of the Rabbit

Objective: The purpose of this study was to evaluate the effects of low-level laser (LLL) energy on the clinical signs of inflammation and the cellular composition of synovial fluid (SF) in the inflamed knee of the rabbit. Background Data: There are few findings related to the effects of LLL on SF in inflammatory processes and there is little knowledge about the optimal parameters for reducing joint inflammation. Materials and Methods: Inflammation in the right knee of 36 rabbits was induced by intracapsular injection (0.2 mL) of Terebinthina commun (Tc). The animals were randomly assigned to three groups: acute experimental group (AEG), chronic experimental group (CEG), and control group (CG), which only received Tc. Each group was divided in two subgroups of six animals each. The AEG and CEG groups began to receive laser treatment 2 and 5 d after the induction of inflammation, respectively. Laser irradiation at a wavelength of 830 nm, power output of 77 mW, and power density of 27.5 W/cm(2) was applied daily for 7 d for either 0.12 sec or 0.32 sec, resulting in doses of 3.4 J/cm(2) and 8 J/cm(2), respectively. Body mass, joint perimeter, joint temperature, and the morphology of the SF were analyzed. Results: There was no statistically significant differences between groups in the body mass, joint perimeter, and SF morphology. Conclusion: Laser irradiation with the selected parameters produced only a few subtle differences in the inflammatory signs and the SF. The lack of effects may have been due to the short irradiation time.

Association of Trypanosoma vivax in extracellular sites with central nervous system lesions and changes in cerebrospinal fluid in experimentally infected goats

Changes in cerebrospinal fluid (CSF) and anatomical and histopathological central nervous system (CNS) lesions were evaluated, and the presence of Trypanosoma vivax in CNS tissues was investigated through PCR. Twelve adult male goats were divided into three groups (G): G1, infected with T. vivax and evaluated during the acute phase; G2, infected goats evaluated during the chronic phase; and G3, consisting of non-infected goats. Each goat from G1 and G2 was infected with 1.25 x 10(5) trypomastigotes. Cerebrospinal fluid (CSF) analysis and investigation of T. vivax was performed at the 15(th) day post-infection (dpi) in G1 goats and on the fifth day after the manifestation of nervous system infection signs in G2 goats. All goats were necropsied, and CNS fragments from G1 and G2 goats were evaluated by PCR for the determination of T. vivax. Hyperthermia, anemia and parasitemia were observed from the fifth dpi for G1 and G2, with the highest parasitemia peak between the seventh and 21(st) dpi. Nervous system infection signs were observed in three G2 goats between the 30(th) and 35(th) dpi. CSF analysis revealed the presence of T. vivax for G2. Meningitis and meningoencephalitis were diagnosed in G2. PCR were positive for T. vivax in all the samples tested. In conclusion, T. vivax may reach the nervous tissue resulting in immune response from the host, which is the cause of progressive clinical and pathological manifestations of the CNS in experimentally infected goats.

On the accuracy of two-fluid model predictions for a particular gas-solid riser flow

Literature presents a huge number of different simulations of gas-solid flows in risers applying two-fluid modeling. In spite of that, the related quantitative accuracy issue remains mostly untouched. This state of affairs seems to be mainly a consequence of modeling shortcomings, notably regarding the lack of realistic closures. In this article predictions from a two-fluid model are compared to other published two-fluid model predictions applying the same Closures, and to experimental data. A particular matter of concern is whether the predictions are generated or not inside the statistical steady state regime that characterizes the riser flows. The present simulation was performed inside the statistical steady state regime. Time-averaged results are presented for different time-averaging intervals of 5, 10, 15 and 20 s inside the statistical steady state regime. The independence of the averaged results regarding the time-averaging interval is addressed and the results averaged over the intervals of 10 and 20 s are compared to both experiment and other two-fluid predictions. It is concluded that the two-fluid model used is still very crude, and cannot provide quantitative accurate results, at least for the particular case that was considered. (C) 2009 Elsevier Inc. All rights reserved.

AnSBBR Applied to the Treatment of Metalworking Fluid Wastewater: Effect of Organic and Shock Load

An investigation was performed regarding the application of a mechanically stirred anaerobic sequencing batch biofilm reactor containing immobilized biomass on inert polyurethane foam (AnSBBR) to the treatment of soluble metalworking fluids to remove organic matter and produce methane. The effect of increasing organic matter and reactor fill time, as well as shock load, on reactor stability and efficiency have been analyzed. The 5-L AnSBBR was operated at 30 A degrees C in 8-h cycles, agitation of 400 rpm, and treated 2.0 L effluent per cycle. Organic matter was increased by increasing the influent concentration (500, 1,000, 2,000, and 3,000 mg chemical oxygen demand (COD)/L). Fill times investigated were in the batch mode (fill time 10 min) and fed-batch followed by batch (fill time 4 h). In the batch mode, organic matter removal efficiencies were 87%, 86%, and 80% for influent concentrations of 500, 1,000, and 2,000 mgCOD/L (1.50, 3.12, and 6.08 gCOD/L.d), respectively. At 3,000 mgCOD/L (9.38 gCOD/L.d), operational stability could not be achieved. The reactor managed to maintain stability when a shock load twice as high the feed concentration was applied, evidencing the robustness of the reactor to potential concentration variations in the wastewater being treated. Increasing the fill time to 4 h did not improve removal efficiency, which was 72% for 2,000 mgCOD/L. Thus, gradual feeding did not improve organic matter removal. The concentration of methane formed at 6.08 gCOD/L was 5.20 mmolCH(4), which corresponded to 78% of the biogas composition. The behavior of the reactor during batch and fed-batch feeding could be explained by a kinetic model that considers organic matter consumption, production, and consumption of total volatile acids and methane production.