Digestive enzyme activities and gastrointestinal fermentation in wood-eating catfishes

To determine what capabilities wood-eating and detritivorous catfishes have for the digestion of refractory polysaccharides with the aid of an endosymbiotic microbial community, the pH, redox potentials, concentrations of short-chain fatty acids (SCFAs), and the activity levels of 14 digestive enzymes were measured along the gastrointestinal (GI) tracts of three wood-eating taxa (Panaque cf. nigrolineatus “Marañon”, Panaque nocturnus, and Hypostomus pyrineusi) and one detritivorous species (Pterygoplichthys disjunctivus) from the family Loricariidae. Negative redox potentials (−600 mV) were observed in the intestinal fluids of the fish, suggesting that fermentative digestion was possible. However, SCFA concentrations were low (<3 mM in any intestinal region), indicating that little GI fermentation occurs in the fishes’ GI tracts. Cellulase and xylanase activities were low (<0.03 U g−1), and generally decreased distally in the intestine, whereas amylolytic and laminarinase activities were five and two orders of magnitude greater, respectively, than cellulase and xylanase activities, suggesting that the fish more readily digest soluble polysaccharides. Furthermore, the Michaelis–Menten constants (Km) of the fishes’ β-glucosidase and N-acetyl-β-d-glucosaminidase enzymes were significantly lower than the Km values of microbial enzymes ingested with their food, further suggesting that the fish efficiently digest soluble components of their detrital diet rather than refractory polysaccharides. Coupled with rapid gut transit and poor cellulose digestibility, the wood-eating catfishes appear to be detritivores reliant on endogenous digestive mechanisms, as are other loricariid catfishes. This stands in contrast to truly “xylivorous” taxa (e.g., beavers, termites), which are reliant on an endosymbiotic community of microorganisms to digest refractory polysaccharides

Substrate concentration and enzyme allocation can affect rates of microbial decomposition

Abstract. A large proportion of the world&apos;s carbon is stored as soil organic matter (SOM). However, the mechanisms regulating the stability of this SOM remain unclear. Recent work suggests that SOM may be stabilized by mechanisms other than chemical recalcitrance. Here, we show that the mineralization rate of starch, a plant polymer commonly found in litter and soil, is concentration dependent, such that its decomposition rate can be reduced by as much as 50% when composing less than ;10% of SOM. This pattern is largely driven by low activities of starch-degrading enzymes and low inducibility of enzyme production by microbial decomposers. The same pattern was not observed for cellulose and hemicellulose degradation, possibly because the enzymes targeting these substrates are expressed at constitutively high levels. Nevertheless, given the heterogeneous distribution of SOM constituents, our results suggest a novel low-concentration constraint on SOM decomposition that is independent of chemical recalcitrance. These results may help explain the stability of at least some SOM constituents, especially those that naturally exist in relatively low concentrations in the soil environment

Do wood-grazing fishes partition their niche?: morphological and isotopic evidence for trophic segregation in Neotropical Loricariidae

Summary 1. Morphic detritus, including coarse particulate organic matter such as terrestrial tree leaves and wood, is consumed by few fishes in temperate stream systems but is ingested by abundant and diverse groups of specialized fishes in tropical rivers; physiological assimilation and partitioning of morphic detritus by fishes remain poorly understood. 2. This study examines seven species of Neotropical suckermouth-armored catfishes (Loricariidae) that live among and feed on coarse woody debris. Five species represent two unrelated evolutionary lineages showing convergent morphological specializations for gouging into and eating wood, small particles of which fill their guts. Two morphologically distinct species unrelated to wood-eaters and to each other forage along the surface of wood. 3. We examined six jaw functional morphological characteristics of each loricariid species as well as C and N stable isotope ratios of blood plasma, red blood cells and fin tissue of three wood-eating species and muscle tissues of all seven species. Consumer isotopic signatures were compared among species and with isotopic signatures of potential food resources, including biofilm, seston and both bulk wood and holocellulose extracted from bulk wood. 4. Wood-eating species had robust jaws specialized for gouging wood, d 13 C signatures consistent with assimilation of cellulosic wood carbon (not bulk wood carbon or lignin) and elevated d 15 N values (&gt;5AE8&amp;) relative to wood that were consistent with assimilation of N from intermediate microbial decomposers in the environment rather than direct assimilation of N from wood or from endosymbiotic N-fixers. Two non-wood-eating species occupied divergent regions of jaw functional morphospace, and isotopic signatures were consistent with assimilation of C from biofilm and seston, respectively, and N from enriched sources such as microbes, macroinvertebrates or seston. 5. Food resources associated with the surfaces of coarse woody debris in Neotropical rivers are partitioned among at least three guilds of loricariid consumers with divergent jaw morphologies specialized for wood gouging, surface grazing and macroinvertebrate probing. Direct consumption of morphic detritus by specialized Neotropical fishes constitutes a potentially important but poorly understood component of detritus processing and nutrient cycling in tropical rivers

Phenotypic plasticity of gut structure and function during periods of inactivity in Apostichopus japonicus.

Apostichopus japonicus is a common sea cucumber that undergoes seasonal inactivity phases and ceases feeding during the summer months. We used this sea cucumber species as a model in which to examine phenotypic plasticity of the digestive tract in response to food deprivation. We measured the body mass, gross gut morphology and digestive enzyme activities of A. japonicus before, during, and after the period of inactivity to examine the effects of food deprivation on the gut structure and function of this animal. Individuals were sampled semi-monthly from June to November (10 sampling intervals over 178 days) across temperature changes of more than 18°C. On 5 September, which represented the peak of inactivity and lack of feeding, A. japonicus decreased its body mass, gut mass and gut length by 50%, 85%, and 70%, respectively, in comparison to values for these parameters preceding the inactive period. The activities of amylase, cellulase and lipase decreased by 77%, 98%, and 35% respectively, in comparison to mean values for these enzymes in June, whereas pepsin activity increased two-fold during the inactive phase. Alginase and trypsin activities were variable and did not change significantly across the 178-day experiment. With the exception of amylase and cellulase, all body size indices and digestive enzyme activities recovered and even surpassed the mean values preceding the inactive phase during the latter part of the experiment (October-November). Principal Component Analysis (PCA) utilizing the digestive enzyme activity and body size index data divided the physiological state of this cucumber into four phases: an active stage, prophase of inactivity, peak inactivity, and a reversion phase. These phases are all consistent with previously suggested life stages for this species, but our data provide more defined characteristics of each phase. A. japonicus clearly exhibits phenotypic plasticity (or life-cycle staging) of the digestive tract during its annual inactive period

Substrate concentration constraints on microbial decomposition

Soil organic carbon is chemically heterogeneous, and microbial decomposers face a physiological challenge in metabolizing the diverse array of compounds present in soil. Different classes of polymeric compounds may require specialized enzymatic pathways for degradation, each of which requires an investment of microbial resources. Here we tested the resource allocation hypothesis, which posits that decomposition rates should increase once substrate concentrations are sufficient to overcome biochemical investment costs. We also tested the alternative hypothesis that mixing different substrates increases resource acquisition through priming effects involving generalist enzymes. Using a microcosm approach, we varied the soil concentration of seven distinct substrates individually and in mixture. We found that the percent carbon respired from starch, cellulose, chitin, and the mixture was significantly reduced at the lowest substrate concentration. The activities of β-glucosidase and N-acetyl-glucosaminidase that target cellulose and chitin, respectively, were also significantly lower at the lowest concentrations of their target substrates. However, we did not observe parallel declines in enzyme activity with starch or the mixture. Some enzymes, such as β-xylosidase, were consistent with specialist strategies because they showed the highest activity in the presence of their target substrate. Other enzymes were more generalist, with activity observed across multiple substrates. Together, these results suggest that the costs of biochemical machinery limit microbial decomposition of substrates at low concentration. The presence of enzymes with low substrate specificity was not sufficient to overcome this constraint for some substrates. Concentration constraints driven by microbial allocation patterns may be common in mineral soil and could be represented in new biogeochemical models based on microbial physiology

Measuring phenol oxidase and peroxidase activities with pyrogallol, l-DOPA, and ABTS: Effect of assay conditions and soil type

Microbial phenol oxidases and peroxidases mediate biogeochemical processes in soils, including microbial acquisition of carbon and nitrogen, lignin degradation, carbon mineralization and sequestration, and dissolved organic carbon export. Measuring oxidative enzyme activities in soils is more problematic than assaying hydrolytic enzyme activities because of the non-specific, free radical nature of the reactions and complex interactions between enzymes, assay substrates, and the soil matrix. We compared three substrates commonly used to assay phenol oxidase and peroxidase in soil: pyrogallol (PYGL, 1,2,3-trihydroxybenzene), l-DOPA (l-3,4-dihydroxyphenylalanine), and ABTS (2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid). We measured substrate oxidation in three soils across a pH gradient from 3.0 to 10.0 to determine the pH optimum for each substrate. In addition, we compared activities across 17 soils using the three substrates. In general, activities on the substrates followed the trend PYGL&gt;. l-DOPA&gt;ABTS and were inversely related to substrate redox potential. PYGL and ABTS were not suitable substrates at pH&gt;5, and ABTS oxidation often declined with addition of peroxide to the assay. Absolute and relative oxidation rates varied widely among substrates in relation to soil type and assay pH. We also tested whether autoclaved or combusted soils could be used as negative controls for the influence of abiotic factors (e.g., soil mineralogy) on oxidative activity. However, neither autoclaving nor combustion produced reliable negative controls because substrate oxidation still occurred; in some cases, these treatments enhanced substrate oxidation rates. For broad scale studies, we recommend that investigators use all three substrates to assess soil oxidation potentials. For focused studies, we recommend evaluating substrates before choosing a single option, and we recommend assays at both the soil pH and a reference pH (e.g., pH 5.0) to determine the effect of assay pH on oxidase activity. These recommendations should contribute to greater comparability of oxidase potential activities across studies. © 2013 Elsevier Ltd

Microbial enzymatic responses to drought and to nitrogen addition in a southern California grassland

Microbial enzymes play a fundamental role in ecosystem processes and nutrient mineralization. Therefore understanding enzyme responses to anthropogenic environmental change is important for predicting ecosystem function in the future. In a previous study, we used a reciprocal transplant design to examine the direct and indirect effects of drought and nitrogen (N) fertilization on litter decomposition in a southern California grassland. This work showed direct and indirect negative effects of drought on decomposition, and faster decomposition by N-adapted microbial communities in N-fertilized plots than in non-fertilized plots. Here we measured microbial biomass and the activities of nine extracellular enzymes to examine the microbial and enzymatic mechanisms underlying litter decomposition responses to drought and N. We hypothesized that changes in fungal biomass and potential extracellular enzyme activity (EEA) would relate directly to litter decomposition responses. We also predicted that fungal biomass would dominate the microbial community in our semi-arid study site. However, we found that the microbial community was dominated by bacterial biomass, and that bacteria responded negatively to drought treatment. In contrast to patterns in decomposition, fungal biomass and most potential EEA increased in direct response to drought treatment. Potential EEA was also decoupled from the decomposition response to N treatment. These results suggest that drought and N alter the efficiencies of EEA, defined as the mass of target substrate lost per unit potential EEA. Enzyme efficiencies declined with drought treatment, possibly because reduced water availability increased enzyme immobilization and reduced diffusion rates. In the N experiment, the efficiencies of β-glucosidase, β-xylosidase, and polyphenol oxidase were greater when microbes were transplanted into environments from which they originated. This increase in enzymatic efficiency suggests that microbial enzymes may adapt to their local environment. Overall, our results indicate that drought and N addition may have predictable impacts on the efficiencies of extracellular enzymes, providing a means of linking enzyme potentials with in-situ activities

Inside the guts of wood-eating catfishes: can they digest wood?

To better understand the structure and function of the gastrointestinal (GI) tracts of wood-eating catfishes, the gross morphology, length, and microvilli surface area (MVSA) of the intestines of wild-caught Panaque nocturnus, P. cf. nigrolineatus “Marañon”, and Hypostomus pyrineusi were measured, and contrasted against these same metrics of a closely related detritivore, Pterygoplichthys disjunctivus. All four species had anatomically unspecialized intestines with no kinks, valves, or ceca of any kind. The wood-eating catfishes had body size-corrected intestinal lengths that were 35% shorter than the detritivore. The MVSA of all four species decreased distally in the intestine, indicating that nutrient absorption preferentially takes place in the proximal and mid-intestine, consistent with digestive enzyme activity and luminal carbohydrate profiles for these same species. Wild-caught Pt. disjunctivus, and P. nigrolineatus obtained via the aquarium trade, poorly digested wood cellulose (<33% digestibility) in laboratory feeding trials, lost weight when consuming wood, and passed stained wood through their digestive tracts in less than 4 h. Furthermore, no selective retention of small particles was observed in either species in any region of the gut. Collectively, these results corroborate digestive enzyme activity profiles and gastrointestinal fermentation levels in the fishes’ GI tracts, suggesting that the wood-eating catfishes are not true xylivores such as beavers and termites, but rather, are detritivores like so many other fishes from the family Loricariidae

Do herbivorous minnows have “plug-flow reactor” guts? Evidence from digestive enzyme activities, gastrointestinal fermentation, and luminal nutrient concentrations

Few investigations have empirically analyzed fish gut function in the context of chemical reactor models. In this study, digestive enzyme activities, levels of gastrointestinal fermentation products [short chain fatty acids (SCFA)], luminal nutrient concentrations, and the mass of gut contents were measured along the digestive tract in herbivorous and carnivorous minnows to ascertain whether their guts function as “plug-flow reactors” (PFRs). Four of the species, Campostoma anomalum, C. ornatum, C. oligolepis, and C. pauciradii, are members of a monophyletic herbivorous clade, whereas the fifth species, Nocomis micropogon, is a carnivore from an adjacent carnivorous clade. In the context of a PFR model, the activities of amylase, trypsin and lipase, and the concentrations of glucose, protein, and lipid were predicted to decrease moving from the proximal to the distal intestine. I found support for this as these enzyme activities and nutrient concentrations generally decreased moving distally along the intestine of the four Campostoma species. Furthermore, gut content mass and the low SCFA concentrations did not change (increase or decrease) along the gut of any species. Combined with a previous investigation suggesting that species of Campostoma have rapid gut throughput rates, the data presented here generally support Campostoma as having guts that function as PFRs. The carnivorous N. micropogon showed some differences in the measured parameters, which were interpreted in the contexts of intake and retention time to suggest that PFR function breaks down in this carnivorous species

Alzheimer disease models and human neuropathology: similarities and differences

Animal models aim to replicate the symptoms, the lesions or the cause(s) of Alzheimer disease. Numerous mouse transgenic lines have now succeeded in partially reproducing its lesions: the extracellular deposits of Aβ peptide and the intracellular accumulation of tau protein. Mutated human APP transgenes result in the deposition of Aβ peptide, similar but not identical to the Aβ peptide of human senile plaque. Amyloid angiopathy is common. Besides the deposition of Aβ, axon dystrophy and alteration of dendrites have been observed. All of the mutations cause an increase in Aβ 42 levels, except for the Arctic mutation, which alters the Aβ sequence itself. Overexpressing wild-type APP alone (as in the murine models of human trisomy 21) causes no Aβ deposition in most mouse lines. Doubly (APP × mutated PS1) transgenic mice develop the lesions earlier. Transgenic mice in which BACE1 has been knocked out or overexpressed have been produced, as well as lines with altered expression of neprilysin, the main degrading enzyme of Aβ. The APP transgenic mice have raised new questions concerning the mechanisms of neuronal loss, the accumulation of Aβ in the cell body of the neurons, inflammation and gliosis, and the dendritic alterations. They have allowed some insight to be gained into the kinetics of the changes. The connection between the symptoms, the lesions and the increase in Aβ oligomers has been found to be difficult to unravel. Neurofibrillary tangles are only found in mouse lines that overexpress mutated tau or human tau on a murine tau −/− background. A triply transgenic model (mutated APP, PS1 and tau) recapitulates the alterations seen in AD but its physiological relevance may be discussed. A number of modulators of Aβ or of tau accumulation have been tested. A transgenic model may be analyzed at three levels at least (symptoms, lesions, cause of the disease), and a reading key is proposed to summarize this analysis