Mined applying a microbalance (Mettler Toledo XP2U; 0.1 g).Aliquots of
Mined applying a microbalance (Mettler Toledo XP2U; 0.1 g).Aliquots of food suspensions were ETB Accession filtered onto precombusted glass fibre filters (Whatman GFF, 25 mm diameter) and analyzed for particulate organic carbon (POC) and nitrogen employing an EuroEA3000 elemental analyzer (HEKAtech GmbH, Wegberg, Germany). For the determination of particulate phosphorus, aliquots were collected on acid-rinsed polysulfone filters (HT-200; Pall, Ann Arbor, MI, USA) and digested having a answer of 10 potassium peroxodisulfate and 1.5 per cent sodium hydroxide for 60 min at 121 . Soluble reactive phosphorus was determined working with the molybdate-ascorbic acid method [54].Fatty acidsFor the evaluation of fatty acids in the prepared meals suspensions about 1 mg POC had been filtered onto pre-combusted GFF filters (Whatman, 25 mm). Total lipids had been ALK7 medchemexpress extracted three occasions from filters with dichloromethanemethanol (two:1, vv). Pooled cell-free extracts have been evaporated to dryness below a nitrogen stream. For the analysis of fatty acids in the liposomes, aliquots from the liposome stock solutions had been evaporated to dryness straight. The lipid extracts have been transesterified with 3 M methanolic HCl (60 , 20 min). Subsequently, fatty acid methyl esters (FAMEs) had been extracted 3 instances with 2 ml of iso-hexane. The lipid-containing fraction was evaporated to dryness under nitrogen and resuspended in a volume of 20 L iso-hexane. Lipids were analyzed by gas chromatography on a HP 6890 GC equipped having a flame ionization detector (FID) in addition to a DB-225 (J W Scientific, 30 m 0.25 mm ID 0.25 mm film) capillary column to analyse FAMEs. Information of GC configurations for the analysis of FAMEs are given elsewhere [27]. FAMEs were quantified by comparison with an internal normal (C23:0 ME) of known concentration, utilizing multipoint normal calibration curves determined previously with lipid standards (Sigma-Aldrich). FAMEs were identified by their retention times and their mass spectra, which have been recorded using a gas chromatograph-mass spectrometer (Agilent Technologies, 5975C) equipped having a fused-silica capillary column (DB-225MS, J W). Spectra had been recorded amongst 50 and 600 Dalton in the electron effect ionization mode. The limit for quantitation of fatty acids was 20 ng. The absolute quantity of each and every fatty acid was related towards the POC.Data analysis and statisticsInfection efficiencies were analyzed employing a generalized linear model (GLM) with logit function as the hyperlink function for binominal distribution. Treatment effects had been evaluated by assessing deviation in the grand imply. Numbers of offspring developed on the distinct foodSchlotz et al. BMC Ecology 2013, 13:41 http:biomedcentral1472-678513Page 9 ofregimes had been analyzed employing a GLM with log function because the link function for quasi-Poisson distribution. To compensate for overdispersion the model was fitted using quasi-Poisson errors [55]. To specify variations amongst food regimes the subsets “control” and “infected” had been analyzed separately. For each GLMs, multiple comparisons amongst food regimes were conducted using the `multcomp package’ in R (R Development Core Team, 2010) utilizing common linear hypotheses testing as an implementation with the framework for simultaneous inference based on Hothorn et al. [56]. To test for differences in within-host reproduction in the parasite in between food treatments one-way analyses of variance (ANOVA) have been carried out followed by numerous comparisons (Tukey’s HSD); assumptions for ANOVA have been met.
