MALDI-TOF "fingerprint" phospholipid mass spectra allow the differentiation between ruminantia and feloideae spermatozoa

The detailed comparative analysis of sperm lipids could essentially contribute to a better understanding of membrane function in the context of fertilization and, moreover, of sperm preservation. The application of sensitive analytical methods is particularly necessary for endangered species as the available amount of spermatozoa (and, accordingly, extractable lipids) is strongly limited. It will be shown that matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is a fast, simple and sensitive method for the determination of the phospholipid composition of spermatozoa from several ruminantia (cattle, roe deer, Klipspringer) and feloideae species (domestic cat, Siberian tiger, fosa). Characteristic “fingerprints” are obtained from the positive ion spectra that allow the differentiation between both animal groups. In contrast to the lipid extracts of ruminantia spermatozoa which predominantly contain ether lipids including essential amounts of plasmalogens, the more complex phospholipid composition of feloideae spermatozoa is clearly dominated by diacyl phospholipids and contains only marginal amounts of plasmalogens. It will also be shown that the lipid compositions of ejaculated, electroejaculated and cauda epididymal spermatozoa of the same species are very similar and give comparable data. Therefore, the analysis of ejaculated spermatozoa is not an absolute must.     

MALDI-TOF mass spectrometry as a simple tool to determine the phospholipid/glycolipid composition of sperm: pheasant spermatozoa as one selected example

Cellular membranes are composed of highly variable lipid molecules, mainly cholesterol and phospholipids (PLs). The cholesterol moiety and the saturation degree of the fatty acyl residues of PL determine the fluidity of the membrane, which is particularly important for sperm because they have to undergo characteristic membrane-dependent processes (acrosomal exocytosis and fusion with the oocyte). Glycolipids are an essential part of the membrane surface acting as key mediators in the interactions of sperm with components of the female genital tract. Although the lipid composition of many mammalian spermatozoa has already been determined, the lipid composition of avian spermatozoa has scarcely been investigated. Using spermatozoa extracts of the ring-necked pheasant (Phasianus colchicus) as a selected example, this work demonstrates that matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a simple and fast method to determine spermatozoal lipid compositions. The lipid compositions of pheasant spermatozoa have not yet been investigated. In addition to common membrane (primarily diacyl) PL (sphingomyelin, phosphatidylcholine, phosphatidylinositol and phosphatidylethanolamine), remarkable variation of different sulfoglycolipids (sulfogalactocerebrosides) was identified. This is in strong contrast to all other animal species investigated so far which nearly exclusively contain the sulfoglycolipid seminolipid (sulfogalactoalkylacylglycerol). We emphasize that the MALDI MS approach allows the characterization of sulfoglycolipids of sperm within a few minutes without the necessity for previous chromatographic separation.     

Thermal performance curves identify seasonal and site-specific variation in the development of Ecklonia radiata (Phaeophyceae) gametophytes and sporophytes

Rapid ocean warming is affecting kelp forests globally. While the sporophyte life stage has been well studied for many species, the microscopic life stages of laminarian kelps have been understudied, particularly regarding spatial and temporal variations in thermal tolerance and their interaction. We investigated the thermal tolerance of growth, survival, development, and fertilization of Ecklonia radiata gametophytes, derived from zoospores sampled from two sites in Tasmania, Australia, throughout a year, over a temperature gradient (3–30°C). For growth we found a relatively stable thermal optimum at ~20.5°C and stable thermal maxima (25.3–27.7°C). The magnitude of growth was highly variable and depended on season and site, with no consistent spatial pattern for growth and gametophyte size. Survival also had a relatively stable thermal optimum of ~17°C, 3°C below the optimum for growth. Gametophytes grew to single cells between 5 and 25°C, but sporophytes were only observed between 10 and 20°C, indicating reproductive failure outside this range. The results reveal complex effects of source population and season of collection on gametophyte performance in E. radiata, with implications when comparing results from material collected at different localities and times. In Tasmania, gametophytes grow considerably below the estimated thermal maxima and thermal optima that are currently only reached during summer heatwaves, whereas optima for survival (~17°C) are frequently reached and surpassed during heatwaves, which may affect the persistence and recruitment of E. radiata in a warmer climate.     

Fungal biomass associated with decaying leaf litter in a stream

Summary Fungal biomass, measured as ergosterol content, was determined on alder leaf litter incubated during autumn in a softwater Pyrenean stream. The ergosterol content of the leaf litter increased rapidly to a maximum of 462 μg/g detrital dry mass. Ergosterol contents of aquatic Hyphomycetes grown in shake culture were typically ≤5 mg/g mycelial dry mass. Using the corresponding ergosterol-to-biomass conversion factor of 200, peak fungal mass accounted for 9.2% of total system mass, or 10.2% of leaf dry mass. For the period of highest activity (incubation days 7–28), net fungal production on leaf litter was estimated as 2.3 mg d⁻¹ g⁻¹ leaf mass. A conservative estimate of the growth efficiency for the same period was 105 mg mycelial mass per gram leaf mass degraded, assuming that non-leaf organic matter did not constitute an important carbon source supporting fungal production.     

Changes of murine sperm phospholipid composition during epididymal maturation determined by MALDI-TOF mass spectrometry

After leaving the testis, spermatozoa undergo several important steps of biochemical maturation during the passage through the epididymis, increasing their motility and fertilizing ability. These changes comprise (among others) the modification of the phospholipid composition of the sperm membrane. This process is thought to be important for the achievement of motility and fertilizing capacity. The lipids of the sperm membrane are characterized by a significant content of unsaturated fatty acyl residues, resulting in a high sensitivity against oxidative stress. This is evidenced by the appearance of lysolipids, for example, lysophosphatidylcholine, which acts like a detergent and is normally present in only very small amounts in biological membranes. The epididymis represents a tubular system comprising three main parts (caput, corpus, and cauda), through which the spermatozoa are consecutively transported undergoing distinct maturation stages. Using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, we established three striking differences in the lipid composition of murine spermatozoa from the different epididymal regions: in comparison to the caput sperm, sperm from the cauda are characterized by (1) a higher degree of unsaturation (PC 18:0/22:5 and 18:0/22:6 vs. 18:0/20:4 and 18:0/18:1), (2) an enhanced plasmalogen content, and (3) an enhanced content of lysolipids. These changes are likely to be of physiological relevance and potentially useful as diagnostic markers of sperm maturation and acquisition of motility.