Fluconazole resistance in Candida albicans is induced by Pseudomonas aeruginosa quorum sensing.

Microorganisms employ quorum sensing (QS) mechanisms to communicate with each other within microbial ecosystems. Emerging evidence suggests that intraspecies and interspecies QS plays an important role in antimicrobial resistance in microbial communities. However, the relationship between interkingdom QS and antimicrobial resistance is largely unknown. Here, we demonstrate that interkingdom QS interactions between a bacterium, Pseudomonas aeruginosa and a yeast, Candida albicans, induce the resistance of the latter to a widely used antifungal fluconazole. Phenotypic, transcriptomic, and proteomic analyses reveal that P. aeruginosa's main QS molecule, N-(3-Oxododecanoyl)-L-homoserine lactone, induces candidal resistance to fluconazole by reversing the antifungal's effect on the ergosterol biosynthesis pathway. Accessory resistance mechanisms including upregulation of C. albicans drug-efflux, regulation of oxidative stress response, and maintenance of cell membrane integrity, further confirm this phenomenon. These findings demonstrate that P. aeruginosa QS molecules may confer protection to neighboring yeasts against azoles, in turn strengthening their co-existence in hostile polymicrobial infection sites.

Antibiotic-induced inhibition of pheromone synthesis in a bark beetle.

Ingestion of diet containing streptomycin inhibited the conversion of myrcene, a host plant terpene, to the male-specific pheromones ipsenol and ipsdienol in 1ps paraconfusus. Synthesis of cis-verbenol, which is not a sex-specific pheromone, from the host plant terpene (-)-alpha-pinene and other metabolites from these two terpenes was not inhibited by the antibiotic.

Western pine beetle: field response to its sex pheromone and a synergistic host terpene, myrcene.

In the field, both sexes of the western pine beetle, Dendroctonus brevicomis, are attracted by the female-produced bicyclic ketal exo-brevicomin; this response is enhanced by myrcene (a constituent of the beetle's host, ponderosapine), which is not an attractant by itself. This synergism may be part of the phenomenon of the mass attack on its host. Temnochila virescens chlorodia, one of the principal insectan predators of this beetle, is attracted by exo-brevicomin alone.

Response of Ips confusus to synthetic sex pheromones in nature.

The first flight response of bark beetles to synthetic sex pheromones under natural conditions is reported. Two insect species predaceous on this bark beetle also responded. The synthetic compounds delivered in an airstream from a substrate of Carbowax 20M on Chromosorb A appear to elicit the same response as the natural attractant from male-infested bolts of ponderosa pine elicits.

Brevicomin: principal sex attractant in the frass of the female western pine beetle.

The principal component of the sex attractant produced by the female western pine beetle (Dendrotonus brevicomis) is exo-7-ethyl-5-methyl-6,8-dioxabicyclo[3.2.1]octane; the trivial name brevicomin is suggested.

Western pine beetle: specificity among enantiomers of male and female components of an attractant pheromone.

The flight response of both sexes of Dendroctonus brevicomis to the mixture of myrcene, racemic frontalin, and (1R,5S,7R)-(+)-exo-brevicomin and to the mixture of myrcene, (1S,5R)-(-)-frontalin and racemic exo-brevicomin was significantly greater than the response to the same mixtures in which the antipodes were substituted. The flight response to these two mixtures was also greater than the response to the ternary mixture of myrcene, racemic frontalin, and racemic exo-brevicomin (MFE). The walking response of both sexes to the mixture of myrcene, racemic frontalin, and (+)-exo-brevicomin was not different from the response to MFE. Substitution of the antipode lowered the response when compared to that of MFE. When evaporated with ponderosa pine turpentine, (-)-frontalin was active in the field while its antipode was not.

Regulation of mammary gland sensitivity to thyroid hormones during the transition from pregnancy to lactation.

Thyroid hormones are galactopoietic and help to establish the mammary gland's metabolic priority during lactation. Expression patterns for genes that can alter tissue sensitivity to thyroid hormones and thyroid hormone activity were evaluated in the mammary gland and liver of cows at 53, 35, 20, and 7 days before expected parturition, and 14 and 90 days into the subsequent lactation. Transcript abundance for the three isoforms of iodothyronine deiodinase, type I (DIO1), type II (DIO2) and type III (DIO3), thyroid hormone receptors alpha1 (TRalpha1), alpha2 (TRalpha2) and beta1 (TRbeta1), and retinoic acid receptors alpha (RXRalpha) and gamma (RXRgamma), which act as coregulators of thyroid hormone receptor action, were evaluated by quantitative RT-PCR. The DIO3 is a 5-deiodinase that produces inactive iodothyronine metabolites, whereas DIO1 and DIO2 generate the active thyroid hormone, triiodothyronine, from the relatively inactive precursor, thyroxine. Low copy numbers of DIO3 transcripts were present in mammary gland and liver. DIO2 was the predominant isoform expressed in mammary gland and DIO1 was the predominant isoform expressed in liver. Quantity of DIO1 mRNA in liver tissues did not differ with physiological state, but tended to be lowest during lactation. Quantity of DIO2 mRNA in mammary gland increased during lactation (P < 0.05), with copy numbers at 90 days of lactation 6-fold greater than at 35 and 20 days prepartum. When ratios of DIO2/DIO3 mRNA were evaluated, the increase was more pronounced (>100-fold). Quantity of TRbeta1 mRNA in mammary gland increased with onset of lactation, whereas TRalpha1 and TRalpha2 transcripts did not vary with physiological state. Conversely, quantity of RXRalpha mRNA decreased during late gestation to low levels during early lactation. Data suggest that increased expression of mammary TRbeta1 and DIO2, and decreased RXRalpha, provide a mechanism to increase thyroid hormone activity within the mammary gland during lactation.

Gastric lipolysis of milk lipids in suckling rats.

Fatty acid composition of the major lipid classes in stomach contents of suckling rats at 1, 5, 10, 17 and 20 days of lactation was compared to that of milk lipids. In milk, 98% of fatty acids were in triacylglycerols at all lactation times. Medium-chain fatty acid concentrations increased from 8% in colostrum to 26% at day 5. Fatty acid composition of stomach acylglycerols at all lactation times was different from that of milk triacylglycerols, containing less medium-chain fatty acids, 8:0 and 10:0. This preferential hydrolysis was also shown by higher concentrations of medium-chain fatty acids in the free fatty acid fraction. The lipolysis of medium-chain fatty acids from triacylglycerols resulted in the appearance of di- and monoacylglycerols with 50-100% higher amounts of 14:0 and 16:0. The similar fatty acid composition of products suggests that considerable lipolysis occurred in stomachs of suckling rats even at 1 day of age. Although there was a 10-fold increase in milk consumption, the extent of lipolysis was similar throughout the suckling period because of a parallel rise in lingual lipase levels.

Escherichia coli induces apoptosis and proliferation of mammary cells.

Mammary cell apoptosis and proliferation were assessed after injection of Escherichia coli into the left mammary quarters of six cows. Bacteriological analysis of foremilk samples revealed coliform infection in the injected quarters of four cows. Milk somatic cell counts increased in these quarters and peaked at 24 h after bacterial injection. Body temperature also increased, peaking at 12 h postinjection. The number of apoptotic cells was significantly higher in the mastitic tissue than in the uninfected control. Expression of Bax and interleukin-1beta converting enzyme increased in the mastitic tissue at 24 h and 72 h postinfection, whereas Bcl-2 expression decreased at 24 h but did not differ significantly from the control at 72 h postinfection. Induction of matrix metalloproteinase-9, stromelysin-1 and urokinase-type plasminogen activator was also observed in the mastitic tissue. Moreover, cell proliferation increased in the infected tissue. These results demonstrate that Escherichia coli-induced mastitis promotes apoptosis and cell proliferation.

Electrophysiologic testing in the upright position: improved evaluation of patients with rhythm disturbances using a tilt table.

Patients are traditionally evaluated in the supine position in the electrophysiology laboratory, although symptoms due to a cardiac rhythm disturbance are often maximal clinically during standing. The assumption of the upright position results in dependent displacement of blood, followed by prompt vasoconstriction to maintain arterial pressure. This normal response may aggravate tachyarrhythmias by increasing catecholamine levels or may precipitate vasodepressor syncope if the vasoconstrictor response is absent. The use of a tilt table during electrophysiologic testing was evaluated over a 12 month period in 104 patients having a mean age of 60 years (range 37 to 81): 59 with supraventricular tachycardia, 6 with vasovagal syncope and 39 with carotid sinus hypersensitivity. Twenty-three patients (22%) had significant abnormalities when upright that were not present when supine: eight patients with supraventricular tachycardia who had their clinical syndromes of palpitation and syncope reproduced when upright, but only minimal symptoms when supine; two patients with supraventricular tachycardia who had sustained atrioventricular reentry when upright, but only two to eight beats of tachycardia when supine; six patients with syncope and a normal cardiac evaluation before electrophysiologic testing who had their typical spells only after being placed upright during a vasovagal event and seven patients with carotid sinus hypersensitivity who had their clinical syndromes reproduced with carotid sinus massage only when upright, developing hypotension despite maintaining their heart rate with sinus rhythm or pacing (vasodepressor response). In 22% of patients, electrophysiologic testing in the upright position provided clinically important information that was not evident during standard testing in the supine position.

Cardiac histologic findings in patients with life-threatening ventricular arrhythmias of unknown origin.

Percutaneous endomyocardial biopsy (right ventricle in 10, left ventricle in 2) was performed in 12 patients, aged 9 to 57 years, with serious ventricular arrhythmias occurring in the setting of normal cardiac anatomy and mechanical function. Light microscopic examination of tissue revealed histologic abnormalities in 11 patients, including myocardial cellular hypertrophy in 7, interstitial fibrosis in 5, endocardial fibrosis in 2, myocardial degenerative changes in 1 and increased interstitial cellularity in 1. One patient had histologic evidence of acute lymphocytic myocarditis. Thus, a majority of patients with serious ventricular arrhythmias and normal cardiac anatomy had histologic abnormalities, bringing into question the concept of primary electrical heart disease or idiopathic ventricular tachycardia.

Symptomatic "isolated" carotid sinus hypersensitivity: natural history and results of treatment with anticholinergic drugs or pacemaker.

Because syncope may occur intermittently in patients with carotid sinus hypersensitivity, a knowledge of its natural history is needed as a basis for interpreting the usefulness of therapy. Fifty-six consecutive patients are described (47 men and 9 women; mean age 61 years) with carotid sinus hypersensitivity and syncope in whom 24 hour ambulatory monitoring and intracardiac electrophysiologic study revealed no other cause for the syncope. The mean duration of symptoms was 44 months (range 1 to 480) and the mean number of episodes was 4.0 (range 1 to 20). During a follow-up period of 6 to 120 months (median 40), syncope recurred in 3 of 13 patients who received no treatment, in 2 of 23 patients who received a pacemaker and in 4 of 20 patients who received anticholinergic drugs (incidences corrected for totals available at follow-up: 27, 9 and 22%, respectively). Two-thirds of the patients receiving no treatment were asymptomatic compared with all nine of the patients with syncope and a pure cardioinhibitory response to carotid sinus massage who received an atrioventricular (AV) sequential pacemaker. Although pacing was effective in abolishing syncope, its use should be reserved for recurrent episodes because of the high rate of spontaneous remission of symptoms.

Double-blind study of intravenous propafenone for paroxysmal supraventricular reentrant tachycardia.

Propafenone was administered during electrophysiologic testing to determine its efficacy and safety for terminating and preventing reinduction of paroxysmal supraventricular reentrant tachycardia. Four men and 10 women (mean age 50 years, range 28 to 69) were studied. Five patients had Wolff-Parkinson-White syndrome with orthodromic atrioventricular (AV) reentrant tachycardia, three had a concealed accessory pathway with AV reentrant tachycardia and six had tachycardia due to reentry within the AV node. In the five patients with Wolff-Parkinson-White syndrome, propafenone terminated reentrant tachycardia in three (the tachycardia was reinducible in one) and had no effect in two. In the three patients with a concealed accessory pathway, propafenone terminated reentrant tachycardia in all three and prevented reinduction of the tachycardia in two. In the six patients with tachycardia due to reentry within the AV node, propafenone terminated and prevented reinduction of reentrant tachycardia. Propafenone had no effect on blood pressure, heart rate, PA interval, AV node refractoriness or rate of reentrant tachycardia. Propafenone significantly (p less than 0.05) prolonged the AH, HV, QRS and ventriculoatrial intervals and decreased the AV node Wenckebach rate. Of the nine patients receiving long-term oral propafenone therapy, eight had a reduction of at least 90% in reentrant tachycardia during a mean follow-up period of 14.5 months (range 11 to 22); all eight patients had had noninducible reentrant tachycardia after intravenous propafenone. One patient had increased frequency of reentrant tachycardia; this patient had had inducible reentrant tachycardia after intravenous propafenone. In conclusion, intravenously administered propafenone terminated reentrant tachycardia in 85% of patients and prevented reinduction in 71%, with no adverse hemodynamic effects.

Echocardiographic findings in systemic amyloidosis: spectrum of cardiac involvement and relation to survival.

One hundred thirty-two patients with biopsy-proven systemic amyloidosis underwent echocardiographic examination to define the spectrum of cardiac involvement. Echocardiographic abnormalities were then correlated with clinical variables and survival at follow-up. Patients were subgrouped by left ventricular wall thickness: Group I, mean wall thickness 12 mm or less; Group II, mean wall thickness greater than 12 mm but less than 15 mm; Group III, mean wall thickness 15 mm or greater; or Group IV, atypical features such as wall motion abnormalities or left ventricular dilation. Patients with greater wall thickness had a higher frequency of associated echocardiographic abnormalities such as left atrial enlargement or granular sparkling appearance on two-dimensional examination and, more commonly, reduced systolic function. The occurrence of clinical congestive heart failure was strongly correlated with greater wall thickness and multiple other echocardiographic abnormalities. Survival was negatively influenced both by greater wall thickness and reduced systolic function. The median survival of the entire group was 1.1 years. Echocardiographic examination is an important tool for establishing the presence of cardiac amyloid involvement and may be useful in estimating prognosis in such patients.

Adenosine: potential modulator for vasovagal syncope.

OBJECTIVES: This study examined the hypothesis that adenosine could provoke a vasovagal response in susceptible patients. Mechanisms of the vasovagal response were further explored by studying the adenosine-mediated reactions. BACKGROUND: Increased sympathetic activity is frequently observed before vasovagal syncope. Recent studies have demonstrated that adenosine, in addition to its direct bradycardiac and vasodilatory effects, can increase sympathetic discharge by activating cardiovascular afferent nerves. METHODS: The effects of adenosine and head-up tilt-table testing with or without isoproterenol were prospectively evaluated in 85 patients examined for syncope after negative results of electrophysiologic testing (51 men and 34 women, mean [+/- SD] age 61 +/- 17 years). Adenosine bolus injections of 6 mg and 12 mg were sequentially administered to patients in the upright position. The same protocol was implemented in 14 normal control subjects (7 men and 7 women, mean [+/- SD] age 38 +/- 10 years). RESULTS: Transient hypertension or tachycardia was observed in 57 (67%) and 20 (24%) patients after administration of 6 mg and 12 mg of adenosine, respectively, during the immediate phase (first 15 s), suggesting direct sympathetic activation. Hypotension and reflex tachycardia were observed in all patients during the delayed phase (15 to 60 s after adenosine injection), suggesting baroreceptor unloading. A vasovagal response was induced in 22 (26%) and 29 (34%) patients after adenosine administration and during tilt-table testing. Inducibility of a vasovagal response by these two methods was comparable (p = 0.12). Of the control subjects, one (7%) had a vasovagal response after adenosine administration and one (7%) had a positive response during tilt-table testing. CONCLUSIONS: These observations support the idea that adenosine is an endogenous modulator of the cardiac excitatory afferent nerves. Sympathetic activation by adenosine can be direct (i.e., cardiac excitatory afferent nerves) and indirect (i.e., vasodilation and reflex sympathetic activation). Adenosine could be an important modulator in triggering a vasovagal response in susceptible patients during examination for syncope.

Transcription of a gene cluster coding for two aminoacyl-tRNA synthetases and an initiation factor in Escherichia coli.

The alpha and beta subunits of phenylalanyl-tRNA synthetase are encoded by the pheS and pheT genes, respectively. These genes are clustered closely together with the genes for threonyl-tRNA synthetase (thrS) and translation initiation factor IF3 (infC); the gene order is thrS infC pheS pheT. We have used two methods to study the transcription pattern within this cluster. The first was the in vitro transcription of DNA restriction fragments with purified RNA polymerase, followed by fractionation of the RNA products by polyacrylamide gel electrophoresis. The second method was the mapping of promoters by means of the "abortive initiation" reaction of McClure and co-workers. This procedure consists of the incubation of RNA polymerase with DNA restriction fragments plus one nucleoside monophosphate and one [alpha-32P]nucleoside triphosphate; the polymerase synthesizes dinucleotide products of known sequence at promoter sites in the DNA. We found that transcription initiated at an internal site within infC (designated P1), and at two promoter sites between infC and pheS (designated P2 and P3). Transcription terminated at two sites about 200 nucleotides apart, located just before pheS. The initiation and termination signals were arranged so as to yield a nested set of overlapping transcripts. At the P1 promoter, transcription initiated with G-C, at P2 with A-C and sometimes A-G, and at P3 with G-U. Promoter activity was also found in a 3000-base interval that includes the start of the thrS gene; eight or nine transcripts (not mapped in detail) were observed, which started with at least four different dinucleotides. All major initiation sites in the gene cluster represented purine starts, although some pyrimidine initiation was observed in trace amounts. No promoter activity was found between pheS and pheT with either of the two techniques; this observation supports the conclusion that these genes are co-transcribed. No evidence was found for any promoter between the termination sites and the beginning of the pheS gene. It is suggested that one of the terminators is an attenuation site controlling the extension of transcription into pheS and pheT. Attenuation may explain the observed regulation of phenylalanyl-tRNA synthetase by the amino acid supply.

Chromosomal mapping and quantitative analysis of estrogen-related receptor alpha-1, estrogen receptors alpha and beta and progesterone receptor in the bovine mammary gland.

Steroid receptors are key transcriptional regulators of mammary growth, development and lactation. Expression of estrogen receptors alpha (ERalpha) and beta (ERbeta), progesterone receptor (PR), and estrogen-related receptor alpha-1 (ERRbeta) have been evaluated in bovine mammary gland. The ERRalpha is an orphan receptor that, in other species and tissues, appears to function in the regulation of estrogen-response genes including lactoferrin and medium chain acyl-CoA dehydrogenase and in mitochondrial biogenesis. Expression of ERalpha, ERbeta, PR and ERRalpha was characterized in mammary tissue obtained from multiple stages of bovine mammary gland development using quantitative real-time RT-PCR. Expression was evaluated in prepubertal heifers, primigravid cows, lactating non-pregnant cows, lactating pregnant cows and non-lactating pregnant cows (n=4 to 9 animals/stage). In addition, ERalpha, ERbeta, PR and ERRalpha were mapped to chromosomes 9, 10, 15 and 29 respectively, by linkage and radiation hybrid mapping. Results indicated that expression of ERalpha, PR and ERRalpha was largely coordinately regulated and they were present in significant quantity during all physiological stages evaluated. In contrast, ERbeta transcripts were present at a very low concentration during all stages. Furthermore, no ERbeta protein could be detected in bovine mammary tissue by immunohistochemistry. The ERalpha and PR proteins were detected during all physiological states, including lactation. Our results demonstrate the presence of ERalpha, PR and ERRalpha during all physiological stages, and suggest a functional role for ERRalpha and a relative lack of a role for ERbeta in bovine mammary gland development and lactation.

Cold pressor test as a predictor of hypertension.

To determine the usefulness of the cold pressor test as a predictor of hypertension, we compared the blood pressure recordings available from 142 patients in 1979 with readings obtained during performance of two cold pressor tests, the first in 1934 when these subjects were children, and the second in 1961. Forty-eight subjects were hyperreactors to the tests in either 1934 or 1961, and 94 were normoreactors. At last follow-up, blood pressures in 14 of the hyperreactors were between 140 and 160 mm Hg systolic or 90 and 100 mm Hg diastolic (Stratum 1) and in 20 exceeded 160 mm Hg systolic or 100 mm Hg diastolic (Stratum 2). Ten normoreactors had casual blood pressures in Stratum 1 and eight in Stratum 2. Hypertension had thus occurred in 71% of the hyperreactors and 19% of the normoreactors. Fifteen hyperreactors were receiving antihypertensive therapy, and this reduced the severity of the casual blood pressure elevation in most patients to Stratum 1. Antihypertensive therapy had been started in three normoreactors. The duration of follow-up, 45 years, and the mean age at follow-up, almost 57 years, were greater in this study than in any previously reported study. Early hyperreactivity was related to future hypertension in enough subjects to suggest that an abnormal response to an external cold stimulus may be useful as an indicator of future hypertension.

Comparison of the phospholipid composition of breast milk from mothers of term and preterm infants during lactation.

Phospholipids were determined in milk on postpartum day 3 (colostrum) and days 7, 21, 42, and 84 from mothers of 18 very premature (26 to 30 wk gestation age), 28 premature (31 to 36 wk), and 6 term (37 to 40 wk) infants. Lipids were analyzed by thin-layer and gas-liquid chromatography. Total fat content increased during lactation whereas phospholipids and cholesterol declined. Phospholipids were separated from neutral lipids by column chromatography and distributed by preparative thin-layer chromatography into classes, sphingomyelin, phosphatidyl choline, serine, inositol, and ethanolamine for fatty acid analysis. Phospholipids exhibited a remarkable constancy in class percentages in milks from mothers giving birth prematurely or at term. Changes were observed in fatty acid composition within each of the phospholipid classes as secretion progressed from colostrum (3d) to transitional (7d) to mature milk (21, 42, 84d). These changes in phospholipid fatty acid composition occurred only during the first 3 wk of lactation. Mature milk was found to be relatively constant in phospholipid composition.

Milk composition and volume during the onset of lactation in a diabetic mother.

Milk volume and composition were examined in a diabetic mother on days 3-7 postpartum. By day 5 milk volume produced and concentrations of sodium, potassium, chloride, lactose, protein, calcium, magnesium, and citrate were within limits of a reference population. Fat content of the milk was slightly lower. Free fatty acids were 2% of total lipid on day 3 but increased to 23% on days 4-7, suggesting impaired esterification in the mammary gland. Total milk lipoprotein lipase increased approximately fourfold during days 4-5. Other changes were 1) low cholesterol content, only one-fifth of normal milk; 2) decreased medium-chain fatty acids, suggesting impairment of fatty acid synthesis in the mammary gland; 3) increased oleic acid; and 4) high concentrations of polyunsaturated fatty acids, suggesting increased chain elongation. These results suggest that diabetes produces changes in lipid metabolism in the mammary gland that alter the composition of milk produced by the diabetic mother.