Effect of rate of weight gain of steers during the stocker phase. IV. Rumen fermentation characteristics and expression of genes involved in substrate utilization for fatty acid synthesis in adipose tissues of growing-finishing beef cattle.

The objective of this study was to determine the impact of stocker production systems differing in growth rate on rumen fermentation characteristics and utilization of substrates for fatty acid synthesis in intramuscular (IM), subcutaneous (SC), and perirenal (PR) adipose tissues. Angus steers were assigned to 4 stocker cattle production systems in 2 consecutive years: 1) 1.0 kg/d of 40% CP cottonseed meal­based supplement while grazing dormant native range (CON), 2) ground corn/soybean meal­based supplement while grazing dormant native range fed at 1% of BW (CORN), 3) grazing wheat pasture at a high stocking rate to achieve a low rate of BW gain (LGWP), and 4) grazing wheat pasture at a low stocking rate for a high rate of BW gain (HGWP). Eight ruminally cannulated steers were used to determine rumen fermentation characteristics. Steers were harvested during the stocker phase at similar age (different carcass weight) in Exp. 1 (3 steers/treatment) or at similar carcass weight in Exp. 2 (4 steers/treatment). Adipose tissues were analyzed for mRNA expression of genes involved in glucose (solute carrier family 2, member 4 [GLUT4], glucose-6-phosphate dehydrogenase [G6PDH], phosphofructokinase, muscle [PFKM], and pyruvate kinase 2, muscle [PK2]), lactate (lactate dehydrogenase B [LDHB]), and acetate (acetyl-CoA synthetase, cytosol [ACSS2]) utilization for fatty acid synthesis. The acetate:propionate ratio was least (P < 0.05) for HGWP steers, intermediate for CORN and LGWP steers, and greatest for CON steers. At similar age, LGWP and HGWP steers tended (F-test; P < 0.15) to have greater (P < 0.10) G6PDH and ACSS2 mRNA expression than CON and CORN steers in SC and PR but not IM adipose tissue. Expression of PFKM and PK2 mRNA tended (F-test; P < 0.15) to be greater (P < 0.10) in HGWP than CON and LGWP steers in IM but not SC or PR adipose tissue. At similar HCW, expression of GLUT4 and G6PDH mRNA were greater (P < 0.10) in SC adipose tissue of LGWP and HGWP steers compared with CON and CORN steers but not in IM and PR adipose tissue. Expression of LDHB mRNA was lesser (P < 0.10) in SC adipose tissue but greater (P < 0.10) in PR adipose tissue of LGWP and HGWP steers compared with CON and CORN steers. These results indicate a shift toward glucose utilization in SC adipose tissue but a shift towards lactate utilization in PR adipose tissue. These results suggest that diet and changes in VFA profile can influence substrates utilized for fatty acid synthesis, but diet has a greater effect in SC than IM adipose tissue.

Effect of muscle type, sire breed, and time of weaning on fatty acid composition of finishing steers.

Thirty-three steer calves were used to determine the effect of sire breed (Angus or Charolais), time of weaning [normal weaned at approximately 210 d of age (NW) or late weaned at approximately 300 d of age (LW)], and muscle type [LM and semitendinosus muscle (STN)] on fatty acid composition. The whole plot consisted of a 2 (sire breed) × 2 (time of weaning) treatment arrangement, and the subplot treatment was muscle type. Body weights were recorded at 28-d intervals to determine animal performance. Muscle biopsies were collected on d 127 and 128 of finishing. All calves were slaughtered on d 138, and carcass data were collected. Angus-sired steers had lighter initial BW (271 vs. 298 kg; P = 0.02), and LW steers were heavier (351 vs. 323 kg; P = 0.03) on d 28, but no other differences in BW were noted. Charolais-sired steers had larger LM area (P = 0.03), reduced yield grades (P = 0.01), less 12th-rib fat (P < 0.01), and less marbling (P < 0.01) than Angus-sired steers. Carcass measures overall indicate Angus-sired steers were fatter. Hot carcass weight was heavier (348 vs. 324 kg; P = 0.04) in LW steers than NW steers. No other differences (P > 0.05) were observed for feedlot performance or carcass characteristics. Total lipids were extracted from muscle biopsies, derivatized to their methyl esters, and analyzed using gas chromatography. The LM had greater SFA (43.94 vs. 35.76%; P < 0.01) and decreased unsaturated fatty acids (UFA; 56.90 vs. 66.19%; P < 0.01) compared with the STN. Percent total MUFA was greater in STN than LM (51.05 vs. 41.98%; P < 0.01). Total SFA, UFA, and MUFA did not differ due to sire breed or time of weaning. Total PUFA differed (P = 0.04) due to a sire breed × time of weaning interaction but did not differ due to muscle type, with greater PUFA in NW Charolais than any other sire breed × time of weaning combination. Observed changes in percent MUFA may be a result of greater Δ(9)-desaturase activity. The calculated desaturase index suggests STN has a greater Δ(9)-desaturase activity than LM, but no differences (P > 0.05) between sire breed or time of weaning were observed. These results indicate that sire breed, time of weaning, and muscle type all affect fatty acid composition in beef. This information provides insight into factors for manipulation of beef fatty acids. More research is needed to identify beef cuts based on fatty acid profile and healthfulness.

Impact of feeding various amounts of wet and dry distillers grains to yearling steers on palatability, fatty acid profile, and retail case life of longissimus muscle.

Due to increased production of ethanol, abundance of distillers grains (DG) is increasing. Steers (n = 176) were assigned to 1 of 5 treatment groups: steam-flaked corn (SFC), 10% dry DG (DDG), 10% wet DG (WDG), 20% WDG, or 30% WDG. The objectives were to determine the effects of feeding greater amounts of WDG, or DDG on meat quality. Steaks, 2.54 cm, were cut from strip loins and identified for simulated retail display, Warner-Bratzler shear force analysis, palatability, and fatty acid composition. Steaks from cattle fed 10% WDG and 30% WDG had smaller (P < 0.05) Warner-Bratzler shear force values than steaks from cattle fed 20% WDG. Trained sensory panelists found no differences (P > 0.05) in overall tenderness and off-flavors. No differences were found in total SFA and MUFA composition among treatments; however, 20% and 30% WDG had a greater proportion of PUFA and n-6 fatty acids than 10% WDG. No differences were found during simulated retail display between various amounts of WDG. Further research needs to be conducted to evaluate methods that aid in increasing shelf life of steaks from cattle fed greater rates of WDG.

Pyrosequencing and characterization of immune response genes from the American dog tick, Dermacentor variabilis (L.).

Ticks continue to be a threat to animal and human health, and new and novel control strategies are needed for ticks and tick-borne pathogens. The characterization of the tick-pathogen interface and the tick immune response to microbial infections is fundamental toward the formulation of new control strategies for ticks and the pathogens they transmit. Our overall hypothesis for this research is that the tick immune system manages the maintenance of pathogens. Therefore, discovery of tick immune response genes may provide targets for novel control strategies directed toward reducing vector competency and pathogen transmission. In these studies, 454 pyrosequencing, a high-throughput genomic sequencing method was used to discover tick genes expressed in response to bacterial and fungal infections. Expressed sequence tags (ESTs) were analysed from Dermacentor variabilis ticks that had been injected with bacteria (Escherichia coli, Bacillus subtilis, Micrococcus luteus) or fungi (Saccharomyces cerevisiae and Candida albicans) and ticks that were naturally infected with the intracellular bacterium, Anaplasma marginale. By this approach, ESTs were assembled into 5995 contigs. Contigs fell into the five main functional categories of metabolism, genetic information processing, environmental information processing, cellular processes and human diseases. We identified more than 30 genes that are likely to encode for proteins involved in tick immune function. We further analysed by reverse transcriptase PCR (RT-PCR) the expression of 22 of these genes in each of our bacterial or fungal treatment groups and found that seven were up-regulated. Up-regulation of these seven genes was confirmed for bacterial, but not fungal treatment by quantitative PCR (qPCR). One of these products was novel, encoding a new tick defensin. Our results clearly demonstrate the complexities of the tick immune system and mark new directions for further study and characterization of proteins that modulate microbial infections in the American dog tick.

Identification of SNARE and cell trafficking regulatory proteins in the salivary glands of the lone star tick, Amblyomma americanum (L.).

Prostaglandin E(2) (PGE(2)) stimulates secretion of tick salivary gland proteins via a phosphoinositide signaling pathway and mobilization of intracellular Ca(2+) (). Highly conserved intracellular SNARE (soluble NSF attachment protein receptors) complex proteins are associated with the mechanism of protein secretion in vertebrate and invertebrate neuronal and non-neuronal cells. Proteins in the salivary glands of partially fed female lone star ticks cross-react individually with antibodies to synaptobrevin-2 (vesicle (v)-SNARE), syntaxin-1A, syntaxin-2 and SNAP-25 (target (t)-SNAREs), cytosolic alpha/beta SNAP and NSF (N-ethylmaleimide-sensitive fusion protein), Ca(2+) sensitive synaptotagmin, vesicle associated synaptophysin, and regulatory cell trafficking GTPases Rab3A and nSec1. V-SNARE and t-SNARE proteins form an SDS-resistant, boiling sensitive core complex in the salivary glands. Antibodies to SNARE complex proteins inhibit PGE(2)-stimulated secretion of anticoagulant protein in permeabilized tick salivary glands. We conclude that SNARE and cell trafficking regulatory proteins are present and functioning in the process of PGE(2)-stimulated Ca(2+) regulated protein secretion in tick salivary glands.

Identity and synthesis of prostaglandins in the lone star tick, Amblyomma americanum (L.), as assessed by radio-immunoassay and gas chromatography/mass spectrometry.

High concentrations of PGE(2) and PGF(2alpha) were identified by radio-immunoassay (RIA) and/or gas chromatography/mass spectrometry (GC/MS) in the hemolymph, salivary glands and saliva of the lone star tick Amblyomma americanum (L.). Binding studies indicated that PGE(2) was free and not bound to any proteins in the hemolymph. A small amount of 6-keto-PGF(1alpha) (breakdown product of PGI(2); prostacyclin) was also found in the salivary glands but not in the hemolymph or saliva. Neither PGD(2) nor PGA(2)/B(2) was detected in any tick material investigated. Although PGE(2) was found in the gut contents, only small amounts of label crossed the gut into the hemolymph during artificial feeding with labeled PGE(2), indicating that the high amounts of PGE(2) in hemolymph and salivary glands are not sequestered from the host blood meal. Isolated salivary glands and salivary gland homogenates demonstrated robust synthesis of PGE(2) at high concentrations of exogenous arachidonic acid. Synthesis by the salivary glands was monitored by measuring increasing PGE(2) with increasing arachidonic acid by RIA, GC/MS and labeled PGE(2) in the presence of labeled arachidonic acid. Synthesis was inhibited in a dose-dependent manner by indomethacin indicating that the cyclooxygenase synthesizing prostaglandins in ticks shares similarities to the enzyme found in mammals.

Prostaglandin E(2)-stimulated secretion of protein in the salivary glands of the lone star tick via a phosphoinositide signaling pathway.

Previous studies identified a prostaglandin E(2) (PGE(2)) receptor in the salivary glands of partially fed female lone star ticks, Amblyomma americanum (L.). In the present studies, protein secretion from dispersed salivary gland acini was shown to be specific for PGE(2), as compared with PGF(2alpha) or the thromboxane analog U-46619, in accordance with their respective binding affinities for the PGE(2) receptor. Furthermore, the selective PGE(2) EP1 receptor agonist, 17-phenyl trinor PGE(2), was as effective as PGE(2) in stimulating secretion of anticoagulant protein. Calcium ionophore A-23187 (1 to 100 microM) stimulated secretion of anticoagulant protein in a dose-dependent manner but the voltage-gated Ca(2+)-channel blocker verapamil (1 to 1000 microM) and the receptor-mediated Ca(2+)-entry antagonist, SK&F 96365 (1 and 10 microM), and 5mM ethylene glycol bis(beta-aminoethyl ether)-N,NN', N'-tetraacetic acid (EGTA) had no appreciable effect on inhibiting PGE(2)-stimulated secretion of anticoagulant protein. PGE(2) (0.1 microM) and the non-hydrolyzable analog of guanosine triphosphate (GTP), GTPgammaS (10 microM), directly activated phospholipase C (PLC) in a membrane-enriched fraction of the salivary glands after PLC was first incubated with the PGE(2) EP1 receptor antagonist AH-6809, which presumably antagonized endogenous PGE(2) (0.3 microM) in the broken-cell-membrane-enriched fraction. TMB-8, an antagonist of intracellular inositol trisphosphate (IP(3)) receptors, inhibited PGE(2)-stimulated secretion. The results support the hypothesis that PGE(2) stimulates secretion of tick salivary gland protein via a phosphoinositide signaling pathway and mobilization of intracellular Ca(2+).

Phospholipase A2 activity in salivary glands and saliva of the lone star tick (Acari: Ixodidae) during tick feeding.

Phospholipase A2 (PLA2) activity levels in tick [Amblyomma americanum (L.)] salivary glands and saliva were examined during tick feeding by using 14C-phosphatidylcholine as the substrate. Saliva produced by stimulating female ticks to salivate with dopamine contains PLA2 (ts-PLA2) activity. The ts-PLA2 activity level in saliva did not change significantly during tick feeding except for a decrease in the last rapid feeding phase (> 200 mg) and in replete ticks. Phospholipase A2 activity was higher in salivary glands of fed than unfed ticks, both in males and females; activity increased during tick feeding correlating with salivary secretory rates during tick feeding suggesting that much of the PLA2 activity measured in whole salivary glands is synthesized for subsequent secretion. During the time course of in vitro salivation, the first 10 microliters of saliva contained higher ts-PLA2 activity than saliva secreted thereafter. Phospolipase A2 was identified in the saliva of artificially fed ticks indicating that ts-PLA2 is a physiological component of tick saliva.

Prostaglandin E2 in the salivary glands of the female tick, Amblyomma americanum (L.): calcium mobilization and exocytosis.

A cholera toxin-sensitive, prostaglandin E2 (PGE2) specific receptor has been identified in the plasma membrane fraction of tick salivary glands. In the present study, we report that stimulation of dispersed salivary glands of the lone star tick Amblyomma americanum (L.) with 1 nM to 10 microM PGE2 increased the intracellular concentration of inositol trisphosphate (IP3) in a dose-dependent manner. Incubation of dispersed tissue with 1 nM to 10 microM PGE2 also stimulated release of 45Ca2+ from preloaded tissue. PGE2 (10 microM) did not stimulate an influx of 45Ca2+. Therefore, the PGE2 receptor in the salivary glands appears to activate a phosphoinositide phospholipase C signalling pathway to increase formation of intracellular IP3 and, thus, mobilize Ca2+ from intracellular stores. Incubation of dispersed salivary glands with 1 nM to 1 microM PGE2 stimulated secretion of anticoagulant protein, but not at < 1 nM or > 1 microM PGE2. In addition, the mammalian PGE2 EP1 receptor antagonist AH-6809 affected secretion of anticoagulant by dispersed salivary gland tissue at a low concentration supporting the hypothesis that the PGE2 receptor in tick salivary glands is EP1-like. We propose that a major function for PGE2 in tick salivary glands is to mobilize Ca2+ and stimulate secretion (exocytosis) of bioactive proteins into the tick's saliva during feeding.

A novel phospholipase A2 activity in saliva of the lone star tick, Amblyomma americanum (L.).

Saliva from female lone star ticks, Amblyomma americanum, contained a novel phospholipase A2 (PLA2) activity that hydrolyzed 14C-arachidonate from 14C-arachidonyl phosphatidylcholine. The tick saliva PLA2 (ts-PLA2) was active over a broad pH range (4.5-11.5) with two distinct pH optima of pH 5.5 and 9.5. Though extracellular PLA2s are reported to be activated by millimolar Ca2+, ts-PLA2 was sensitive to submicromolar Ca2+ and was half-maximally activated by 3.5 microM Ca2+. Tick saliva contains > 500 microM Ca2+ and the feeding lesion in the host is expected to contain millimolar Ca2+. Saliva exhibited a single peak of PLA2 activity corresponding to a molecular weight of 55.7 +/- 1.3 kDa by size exclusion chromatography. The ts-PLA2 was unaffected by a variety of compounds known to inhibit either secreted or cytosolic PLA2s from other sources. However, ts-PLA2 was inhibited by the substrate analog, oleyloxyethyl phosphorylcholine (IC50 = 1.4 microM), and the end product, arachidonic acid (IC50 = 38 microM). Low concentrations of dithiothreitol did not greatly affect ts-PLA2, but activity was reduced at higher concentrations. The PLA2 activity found in A. americanum salivary glands showed many similarities to ts-PLA2, but also some distinct differences. Secreted at the tick-host interface, ts-PLA2 is thought to play an important, but unknown, role during the prolonged tick feeding.

Isolation and characterization of americanin, a specific inhibitor of thrombin, from the salivary glands of the lone star tick Amblyomma americanum (L.).

A thrombin (EC 3.4.21.5) inhibitor (americanin) was isolated from the salivary glands of the lone star tick Amblyomma americanum (L.) using reversed-phase chromatography and anion-exchange chromatography. Americanin did not inhibit any other protease tested, including factor Xa, plasmin, trypsin, chymotrypsin, elastase, papain, pepsin, and carboxypeptidase. The inhibition of thrombin by americanin decreased dramatically with dilution of the reaction mixture including thrombin, its substrate, and americanin. When thrombin assays were performed in the presence of americanin, the reaction curve showed a time-dependent inhibition. Significant inhibition was observed when americanin concentration was approximately equal to that of thrombin, with a Ki of 0.073 nM. The results suggest that americanin is a specific, reversible, competitive, slow, tight-binding inhibitor of thrombin.

A specific prostaglandin E2 receptor and its role in modulating salivary secretion in the female tick, Amblyomma americanum (L.).

Prostaglandins of the 2-series (e.g. PGE2) are typically synthesized from arachidonic acid (AA) after AA is released from cellular phospholipids after activation of an intracellular phospholipase A2 (PLA2). Treatment of isolated salivary glands with PLA2 inhibitor oleyloxyethyl phosphorylcholine (OPC) or prostaglandin synthetase inhibitors reduced dopamine-induced fluid secretion and cyclic AMP (cAMP) levels in isolated salivary glands. PGE2 and its analog, 17-phenyl trinor PGE2, partly reversed the inhibition of secretion and cAMP level by OPC, suggesting that prostaglandins may have an autocrine effect in modulating tick salivary gland function. A specific PGE2 receptor was identified in the plasma membrane fraction of the salivary glands. The receptor exhibits a single, high affinity PGE2 binding site with a KD approximately 29 nM, is saturable, reversible, and specific for PGE2 and coupled to a cholera toxin-sensitive guanine nucleotide regulatory protein. Assay of adenylate cyclase activity in salivary gland membranes showed that PGE2 neither stimulated nor inhibited adenylate cyclase activity, indicating that the PGE2 effects on cAMP levels and possibly secretion are indirect, and that the PGE2 receptor stimulates an alternate "second messenger" pathway.

Identification of hemolytic activity in saliva of the lone star tick (Acari:Ixodidae).

Hemolytic activity was identified in the saliva of Amblyomma americanum (L.) when red blood cells from sheep were incubated with tick saliva in the presence of phosphatidylcholine and sodium deoxycholate. The hemolytic activity was destroyed by boiling or treating with trypsin. The hemolytic activity in tick saliva was calcium-dependent, and inhibited by a phospholipase A2 inhibitor oleyloxyethyl phosphorylcholine. Phosphatidylserine could replace phosphatidylcholine in the hemolytic assays but phosphatidylethanolamine and phosphatidylinositol were ineffective. Size exclusion chromatography of tick saliva revealed one peak of hemolytic activity, which correlated with the activity of tick salivary phospholipase A2, both having a molecular weight approximately 55,000 daltons. These results suggest that the hemolytic activity in tick saliva results from salivary phospholipase A2. The hemolytic activity in tick saliva may play a role in lysing host red blood cells, thus facilitating the tick digestive process.

Identification and characterization of anticoagulant activities in the saliva of the lone star tick, Amblyomma americanum (L.).

Anticoagulant activities against both the extrinsic and intrinsic coagulation pathways were identified in the saliva of partially fed female lone star ticks, Amblyomma americanum (L.). The activities of factor Xa and thrombin in the common pathway of the coagulation cascade were inhibited by tick saliva. The greatest anticoagulant activities were found in the saliva of ticks weighing more than 200 mg. The anticoagulant activities in tick saliva could be detected without preincubation of tick saliva with sheep plasma, but preincubation significantly increased the activities. Tick saliva anticoagulant activities were abolished by boiling for 15 min or being treated with trypsin for 1 hr. Phosphatidylcholine (3 mM) and phospholipase A2 inhibitor oleyloxyethyl phosphorylcholine (0.2 mM) did not affect the anticoagulant activities significantly, suggesting that the phospholipase A2 activity found in tick saliva does not contribute to the anticoagulant activities. Size exclusion high performance liquid chromatography revealed that the molecular weights of the anticoagulant activities were approximately 16,000 D. The anticoagulant activities in tick saliva are believed to play an important role in facilitating tick feeding by helping overcome the host hemostatic system.

Tick salivary prostaglandins: Presence, origin and significance.

Prostaglandins (PGs) are oxygenated metabolites of polyunsaturated fatty acids, most notably arachidonic acid, that act as 'local hormones', regulating a plethora of physiological processes in mammals and other vertebrates. For a long time, PGs were reported only in higher vertebrates, but more recently they have been reported in lower organisms such as bacteria, yeasts and protozoa, and much information is now available on PGs in insects. Prostaglandins are increasingly reported to exist at the host-parasite interface and are thought to aid the parasite by modulating the inflammatory and immune response. Ticks secrete saliva containing extremely high concentrations of PGs into the host, and in this article Alan Bowman, Jack Dillwith and John Sauer provide a synopsis of the information, to date, on the presence, synthesis and proposed roles for these tick salivary PGs.

Dietary modification of host blood lipids affect reproduction in the lone star tick, Amblyomma americanum(L.).

The feeding and reproductive performance of female lone star ticks (Amblyomma americanum (L.)) infesting guinea pigs on diets containing 15% fish oil (FO) or safflower oil (SO) were investigated. Replete ticks fed on FO-fed guinea pigs weighed approximately 30% less than those on the SO-fed guinea pigs. The lower engorged weight resulted in a similar decrease in the mass and number of eggs laid and number of larvae hatching. No effect of host dietary treatment was observed upon the reproductive efficiency index, egg weight, or hatchability. Guinea pig blood on the FO-diet contained high levels of eicosapentaenoic acid, which has previously been shown to inhibit the accumulation of arachidonic acid in the tick salivary gland. It is suggested that the ticks on the FO-fed guinea pigs have impaired production and secretion of dienoic prostaglandins in the saliva resulting in poorer feeding performance, possibly by altering the amount of host blood present in the feeding lesion.

Alteration of arachidonate levels in tick salivary glands by dietary modification of host blood lipids.

Tick saliva contains prostaglandins of the 2-series, believed to facilitate bloodmeal acquisition. Because ticks cannot synthesize the prostaglandin precursor, arachidonic acid, investigations were undertaken to study the uptake, incorporation, and distribution of arachidonic acid in the salivary glands of the lone star tick in vitro and in vivo. Uptake of [3H]arachidonate by isolated salivary glands was reduced in the presence of low concentrations of arachidonic or eicosapentaenoic acids, but much higher, non-physiological concentrations of oleic and linoleic acids were required to inhibit [3H]arachidonate uptake. The incorporation of [3H]arachidonate into triglycerides increased at high concentrations of arachidonic or eicosapentaenoic acid, but not at any concentration of oleic or linoleic acid. Eicosatetraynoic acid greatly inhibited [3H]arachidonic acid. Guinea pigs fed hydrogenated coconut oil, safflower/primrose oil, or fish oil exhibited altered blood lipids; notably increased levels of eicosapentaenoic acid when fed fish oil. Salivary gland lipids in ticks fed on these hosts were also altered. Ticks parasitizing fish oil-fed guinea pigs contained high levels of eicosapentaenoic acid with a 30% reduction in arachidonate levels. The results demonstrated that eicosapentaenoic acid in the host diet had profound effects on arachidonate assimilation by tick salivary glands, which could lead to altered prostaglandin content in tick saliva.

Biosynthesis of salivary prostaglandins in the lone star tick, Amblyomma americanum.

Dopamine-induced saliva from ticks fed [3H]arachidonic acid contained the radiolabelled prostaglandins E2, F2 alpha, D2, and B2, the latter probably derived from PGE2 owing to the alkalinity of tick saliva. Prostaglandin synthetase (PGS) activity in the salivary gland homogenate from the lone star tick, Amblyomma americanum, could not be detected by standard radiometric methodologies successfully employed for tissues from many animal species, including numerous arthropods. Modifications to the assay conditions had no effect. The presence of a PGS-inhibitor in the salivary glands was ruled out. It is postulated that the PGS in A. americanum salivary glands may be considerably different from that found in other animals, including vertebrate hosts.

Uptake, incorporation and redistribution of arachidonic acid in isolated salivary glands of the lone star tick.

The ability of isolated salivary glands from the lone star tick, Amblyomma americanum, to take up, incorporate and redistribute [3H]arachidonic acid was examined. Uptake of arachidonic acid was concentration dependent--a single salivary gland incorporated up to approximately 2.8 micrograms arachidonic acid in 60 min. Over 90% of the [3H]arachidonate entering the glands was esterified and found only in the phospholipid (approximately 80%) and triglyceride (approximately 10%). Essentially no radioactivity was associated with the diglyceride fraction and none with phosphatidic acid indicating de novo phospholipid synthesis was negligible. Phospholipid synthesis via acylation of lysophospholipids (the Lands pathway) was indicated by the rapidity of the synthesis (< 2 min) and the sensitivity to sulfhydryl-blocking agents. Within the phospholipids, [3H]arachidonate was incorporated only into phosphatidylcholine (PC) and phosphatidylethanolamine (PE). Initially [3H]arachidonate was incorporated primarily into PC, but as the incubation proceeded PE contained an increasing proportion of the label. The proportion of [3H]arachidonate incorporated into triglyceride increased at higher media concentrations of arachidonic acid. The roles of lysophosphatide acyltransferase, transacylase and diglycerol acyltransferase in the distribution of arachidonate in tick salivary glands are discussed.

Origin of arachidonic acid in the salivary glands of the lone star tick, Amblyomma, americanum.

The contribution of synthesis and dietary sequestration to the high arachidonate content of the lone star tick, Amblyomma americanum, salivary glands was investigated by assessing the salivary metabolites of various radiolabeled fatty acid substrates administered to partially fed females. A portion of each of the fatty acids studied was incorporated into the fatty acid moiety of the phospholipid fraction. [14C]acetate was metabolized only into myristic, palmitic, palmitoleic, steric, and oleic acids. [3H]oleic acid, [14C]linoleic acid, [14C]gamma-linolenic acid and [14C]eicosatrienoic acids were incorporated into salivary gland phospholipids but underwent little change including elongation and/or desaturation to arachidonate. Ingested [3H]arachidonic acid was readily taken up by the salivary gland and distributed among the lipid classes in a pattern markedly different from that of the other fatty acids tested. We conclude that ticks are unable to synthesize arachidonic acid for incorporation into the salivary glands, but rather sequester it from the host bloodmeal.