Mass spectrometry and animal science: protein identification strategies and particularities of farm animal species

Proteomic approaches are gaining increasing importance in the context of all fields of animal and veterinary sciences, including physiology, productive characterization, and disease/parasite tolerance, among others. Proteomic studies mainly aim the proteome characterization of a certain organ, tissue, cell type or organism, either in a specific condition or comparing protein differential expression within two or more selected situations. Due to the high complexity of samples, usually total protein extracts, proteomics relies heavily on separation procedures, being 2D-electrophoresis and HPLC the most common, as well as on protein identification using mass spectrometry (MS) based methodologies. Despite the increasing importance of MS in the context of animal and veterinary science studies, the usefulness of such tools is still poorly perceived by the animal science community. This is primarily due to the limited knowledge on mass spectrometry by animal scientists. Additionally, confidence and success in protein identification is hindered by the lack of information in public databases for most of farm animal species and their pathogens, with the exception of cattle (Bos taurus), pig (Sus scrofa) and chicken (Gallus gallus). In this article, we will briefly summarize the main methodologies available for protein identification using mass spectrometry providing a case study of specific applications in the field of animal science. We will also address the difficulties inherent to protein identification using MS, with particular reference to experiments using animal species poorly described in public databases. Additionally, we will suggest strategies to increase the rate of successful identifications when working with farm animal species.     

Estuarine colonization, population structure and nursery functioning for 0-group sea bass (Dicentrarchus labrax), flounder (Platichthys flesus) and sole (Solea solea) in a mesotidal temperate estuary

The function of the Mondego estuary as a fish nursery habitat was investigated from June 2003 to June 2004 by comparing the timing of estuarine colonization with juveniles of sea bass Dicentrarchus labrax, flounder Platichthys flesus, and sole Solea solea, while also analysing their population structure, growth and diet composition. Differences in the onset of estuarine colonization were observed, since sole juveniles were the first to enter the estuary (in January), followed by flounder in April and sea bass in June. The estuarine population of these species consisted of several age‐groups, although the majority of individuals belonged to age‐groups 0 and 1. The growth rates determined for 0‐group fish were within the range of those reported for other European estuarine systems. Some differences were also recognized regarding the timing of estuarine colonization and the length of the growing season. Diet of 0‐group sea bass consisted mainly of Crustacea, Polychaeta and Mollusca. Flounder juveniles fed chiefly on Amphipoda (especially Corophium spp.), with Polychaeta, Isopoda and Decapoda also being common prey. The diet of 0‐group sole was dominated by Polychaeta, with Amphipoda, Mollusca and Decapoda ranking highest, with other important benthic organisms also being present. Dietary overlap among these species was relatively low.     

Spring feeding by pink-footed geese reduces carbon stocks and sink strength in tundra ecosystems

Tundra ecosystems are widely recognized as precious areas and globally important carbon (C) sinks, yet our understanding of potential threats to these habitats and their large soil C store is limited. Land‐use changes and conservation measures in temperate regions have led to a dramatic expansion of arctic‐breeding geese, making them important herbivores of high‐latitude systems. In field experiments conducted in high‐Arctic Spitsbergen, Svalbard, we demonstrate that a brief period of early season belowground foraging by pink‐footed geese is sufficient to strongly reduce C sink strength and soil C stocks of arctic tundra. Mechanisms are suggested whereby vegetation disruption due to repeated use of grubbed areas opens the soil organic layer to erosion and will thus lead to progressive C loss. Our study shows, for the first time, that increases in goose abundance through land‐use change and conservation measures in temperate climes can dramatically affect the C balance of arctic tundra.     

Isolation and characterization of microsatellite markers from the entomopathogenic fungus Metarhizium anisopliae

Fourteen polymorphic microsatellite markers were isolated from the entomopathogenic fungus, Metarhizium anisopliae, based on enriched genomic libraries. In order to assess allelic variability, the microsatellite loci were analysed in a collection of 34 isolates sampled from across Switzerland. The number of detected alleles in 14 loci ranged from two to eight and expected heterozygosity from 0.265 to 0.808. Because of the high expected heterozygosity, the 14 microsatellite loci are very useful for ecological studies and analysis of population diversity, and to identifying, monitoring, and tracking M. anisopliae strains applied as biological control agents.     

Sensitivity of respiratory metabolism and efficiency to foliar nitrogen during growth and maintenance

Scaling of respiration from the leaf to the canopy level currently depends on identification of physiological parameters that are tightly linked to respiration and that can readily be determined. Several recent studies have helped provide guides to predicting whole canopy respiration on the basis of foliar nitrogen (N). This approach is potentially powerful owing to the well‐described patterns of allocation of N that follow interception of radiation. In the present study, we investigated the sensitivity of the N–respiration correlation to environmental and developmental factors, in order to evaluate its usage for attempts to scale respiration to the organism and ecosystem level. We studied fully expanded, 1 and 2‐year‐old, and current‐year needles from canopies of Pinus radiata that had been treated (unthinned, thinned and thinned+fertilized treatments) in ways likely to induce a wide range of growth and respiratory responses. We examined respiration in detail during the growth period in spring and again at the end of summer, using calorespirometric methods (combined measurements of CO₂ and heat rates) to determine the respiration rates , instantaneous enthalpic growth rates (R_SGΔH_B, a measure of the conservation of electrons in anabolic products) and the enthalpy conversion efficiency (η_H) of needles differing in age. A general linear model revealed that was positively correlated with needle N, but this correlation was strongly dependent on the season and the needle age – indicating an important physiological difference between expanding young needles and fully expanded old needles. Furthermore, the strength of the correlation between needle N and respiration was comparatively weak for the current year, expanding foliage, indicating that factors other than foliage N significantly influenced the respiration of young needles. The analysis of instantaneous growth rates revealed two general processes. Older, nonexpanding foliage showed considerable rates of enthalpic growth (increases in enthalpy) that was mainly caused by the increment of lignin during secondary growth. Secondly, canopy development appeared dynamic and to be optimized according to environmental drivers and constraints – such as light and water availability. In late spring, needle extension slowed in the upper, but not the lower canopy, because the upper canopy appeared to be affected first by the onset of drought stress in late spring. Growth rates were reduced in the upper canopy despite greater rates of respiration, indicating higher demand of ATP for the maintenance of protein and for export of sugars. Consequently, the enthalpy conversion efficiency and enthalpic N productivity (enthalpic growth per unit N) were comparatively poor indicating advanced development of needles in the upper canopy. We suggest that the growth and maintenance paradigm of respiration is, at best, only moderately useful when applied to whole trees, and is not valid at the cellular level or that of the plant organ. A different concept, namely that of respiratory efficiency, seems a more suitable way to represent respiration in carbon (C) balance models and should help provide a better mechanistic understanding of how respiration affects the C conversion efficiency of plants, and ultimately the net primary productivity of ecosystems.