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The endogenous circadian period of animals and humans is typically very close to 24 h. Individuals with much longer circadian periods have been observed, however, and in the case of humans, these deviations have health implications. Previously, we observed a line of Drosophila with a very long average period of 31.3 h for locomotor activity behavior. Preliminary mapping indicated that the long period did not map to known canonical clock genes but instead mapped to multiple chromosomes. Using RNA-Seq, we surveyed the whole transcriptome of fly heads from this line across time and compared it with a wild-type control. A three-way generalized linear model revealed that approximately two-thirds of the genes were expressed differentially among the two genotypes, while only one quarter of the genes varied across time. Using these results, we applied algorithms to search for genes that oscillated over 24 h, identifying genes not previously known to cycle. We identified 166 differentially expressed genes that overlapped with a previous Genome-wide Association Study (GWAS) of circadian behavior, strongly implicating them in the long-period phenotype. We tested mutations in 45 of these genes for their effect on the circadian period. Mutations in Alk, alph, CG10089, CG42540, CG6034, Kairos ( CG6123), CG8768, klg, Lar, sick, and tinc had significant effects on the circadian period, with seven of these mutations increasing the circadian period of locomotor activity behavior. Genetic rescue of mutant Kairos restored the circadian period to wild-type levels, suggesting it has a critical role in determining period length in constant darkness.

Hip and groin injuries are common in athletes who take part in high level sports. Adductor muscle tendon injuries represent a small but important number of these injuries. Avulsion of the tendons attached to the symphysis pubis has previously been described: these can be managed both operatively and non-operatively. We describe an uncommon variant of this injury, namely complete avulsion of the adductor sleeve complex: this includes adductor longus, pectineus and rectus abdominis. We go on to describe a surgical technique which promotes a full return to the pre-injury level of sporting activity. Over a period of ten years, 15 high-level athletes with an MRI-confirmed acute adductor complex avulsion injury (six to 34 days old) underwent surgical repair. The operative procedure consisted of anatomical re-attachment of the avulsed tissues in each case and mesh reinforcement of the posterior inguinal wall in seven patients. All underwent a standardised rehabilitation programme, which was then individualised to be sport-specific. One patient developed a superficial wound infection, which was successfully treated with antibiotics. Of the 15 patients, four complained of transient local numbness which resolved in all cases. All patients (including seven elite athletes) returned to their previous level of participation in sport. Cite this article: Bone Joint J 2015;97-B:1488–92.

RNA from various mouse organs was analyzed by Northern hybridization to determine the response of squalene synthase (SQS) mRNA to dietary cholesterol, or lovastatin and cholestyramine, administration. Two size-classes of highly abundant mouse SQS (mSQS) mRNAs of approximately 1.9 and 2.0 kb were found in testis. These transcripts were unresponsive to sterol regulation. A single size-class of liver mSQS mRNA of approximately 1.9 kb was sterol-regulated. Studies using primer extension and 5' rapid amplification of cDNA ends (RACE) indicated that the size differences in liver and testis mSQS transcripts were due to variations in the lengths of the 5' untranslated regions (UTRs). The longest testis 5' UTR extended approximately 106 nt 5' of the primary transcription initiation site in liver of mice fed lovastatin and cholestyramine. These results suggest that tissue-specific promoter elements control the transcriptional regulation of the promoters for the mSQS gene in liver and testis.

This study designed new quality scales to measure 2 prevention–appraisal–failure constructs, (1) product quality's strategic importance construct and (2) nonfinancial/financial disclosure constructs. Three of these scales exhibited consistently uniform dimensionality results, loading patterns, reliabilities, and convergent/divergent validity. Instead of relying on only 1 sample and 1 extraction method this study's results used 3 samples and 2 extraction methods. First, exploratory factor analysis was used, and results were compared against those produced by a nonlinear principal component technique rescaling the data as ordinal. The use of multiple samples and multiple extraction represented an effort to produce robust measurement results independent of a single sample's and a single extraction technique's characteristics.

Background Urinary incontinence is a common problem but, for those experiencing significant clinical symptoms, help seeking has been reported to be as low as 20%. As many of these people could be helped by simple interventions, there is a clear need to identify barriers to help seeking and develop interventions to overcome these barriers. Objectives This study explores help-seeking behaviour in people with urinary symptoms such as leakage, frequency, nocturia and urgency in order to identify barriers to service use. Method Thirty-one people who either had agreed to treatment as part of an intervention study or who were receiving treatment at a hospital out-patient clinic for their urinary problems took part in unstructured, taped interviews. Respondents were questioned about their views on help seeking for urinary problems and their personal experiences of consultations. The data were transcribed, coded and thematic analysis carried out. Results The most common theme to emerge was a lack of knowledge of the condition and of available treatments. Urinary symptoms frequently were considered a normal part of ageing or childbirth, or it was felt that these types of symptoms were inappropriate for medical intervention. Older people were not only more likely to accept symptoms but were also less likely to want to bother their GP. Patients did not always communicate their concerns about urinary symptoms to their GP, through either embarrassment or misconceptions of what is a 'medical problem'. Conclusions There is a clear need for health education and health promotion in this area. Future work should explore professionals' knowledge and views of these types of conditions with the aim of providing guidelines for management in primary care.

Mammalian squalene synthase (SQS) catalyzes the first reaction of the branch of the isoprenoid metabolic pathway committed specifically to sterol biosynthesis. SQS produces squalene in an unusual two-step reaction in which two molecules of farnesyl diphosphate are condensed head-to-head. Recent studies have advanced understanding of the reaction mechanism, the functional domains of the enzyme, and transcriptional regulation of the gene. Site-directed mutagenesis has identified conserved Asp, Tyr, and Phe residues that are essential for SQS activity. The Asp residues are hypothesized to be required for substrate binding; the Tyr and Phe residues may stabilize carbocation reaction intermediates. The elucidation of SQS crystal structure will most likely direct future research on the relationship between enzyme structure and function. SQS activity, protein, and mRNA levels are regulated by cholesterol status and by the cytokines TNF-alpha and IL-1beta. Activation of the SQS promoter in response to cholesterol deficit is mediated by sterol regulatory element binding proteins SREBP-1a and SREBP-2. The precise contributions made by individual SREBPs and accessory transcription factors to SQS transcriptional control, and the mechanisms underlying cytokine regulation of SQS are major foci of current research.

Farnesyl diphosphate synthase (FPPS: EC2.5.1.10), a key enzyme in isoprenoid metabolic pathways, catalyzes the synthesis of farnesyl diphosphate (FPP) an intermediate in the biosynthesis of both sterol and non-sterol isoprenoid end products. The localization of FPPS to peroxisomes has been reported ( Krisans, S. K., J. Ericsson, P. A. Edwards, and G. A. Keller. 1994. J. Biol. Chem. 269: 14165-14169). Using indirect immunofluorescence and immunoelectron microscopic tech- niques we show here that FPPS is localized predominantly in the peroxisomes of rat hepatoma H35 cells. However, the par- tial release of 60-70% of cellular FPPS activity is observed by selective permeabilization of these cells with digitonin. Under these conditions, lactate dehydrogenase, a cytosolic enzyme, is completely released whereas catalase, a known peroxisomal enzyme, is fully retained. Digitonin treatment of H35 cells dif- ferentially affects the release of other peroxisomal enzymes involved in isoprenoid metabolism. For instance, mevalonate kinase and phosphomevalonate kinase are almost totally re- leased (95% and 91%, respectively), whereas 3-hydroxy-3- methylglutaryl-CoA reductase is fully retained. Indirect immu- noflourescence studies indicate that FPPS is localized in peroxisomes of Chinese hamster ovary (CHO)-K1 cells but is dispersed in the cytosol of ZR-82 cells, a mutant that lacks per- oxisomes. Unlike in H35 cells, FPPS is completely released upon digitonin permeabilization of CHO-K1 and ZR-82 cells. In contrast, under the same permeabilization conditions, cata- lase is fully retained in CHO-K1 cells but completely released from ZR-82 cells. These studies indicate that FPPS and other enzymes in the isoprenoid biosynthetic pathways, in- volved in the formation of FPP, are differentially associated with peroxisomes and may easily diffuse to the cytosol. Based on these observations, the significance and a possible regula- tory model in the formation of isoprenoid end-products are discussed. —Gupta, S. D., R. S. Mehan, T. R. Tansey, H-T. Chen, G. Goping, I. Goldberg, and I. Shechter. Differential binding of proteins to peroxisomes in rat hepatoma cells: unique association of enzymes involved in isoprenoid metabo- lism. J. Lipid Res. 1999. 40: 1572-1584. Supplementary key words peroxisomescholesterolisoprenoids • immunofluorescencecellular permeabilizationfarnesyl diphosphate synthasecatalase

The autism spectrum disorders (ASDs) are a group of conditions characterized by impairments in reciprocal social interaction and communication, and the presence of restricted and repetitive behaviours 1. Individuals with an ASD vary greatly in cognitive development, which can range from above average to intellectual disability2. Although ASDs are known to be highly heritable ( ∼90%)3, the underlying genetic determinants are still largely unknown.Hereweanalysed the genome-wide characteristics of rare (

Although autism spectrum disorders (ASDs) have a substantial genetic basis, most of the known genetic risk has been traced to rare variants, principally copy number variants (CNVs). To identify common risk variation, the Autism Genome Project (AGP) Consortium genotyped 1558 rigorously defined ASD families for 1 million single-nucleotide polymorphisms (SNPs) and analyzed these SNP genotypes for association with ASD. In one of four primary association analyses, the association signal for marker rs4141463, located within MACROD2, crossed the genome-wide association significance threshold of P < 5 3 10-28. When a smaller replication sample was analyzed, the risk allele at rs4141463 was again over-transmitted; yet, consistent with the winner's curse, its effect size in the replication sample was much smaller; and, for the combined samples, the association signal barely fell below the P < 5 × 10-28 threshold. Exploratory analyses of phenotypic subtypes yielded no significant associations after correction for multiple testing. They did, however, yield strong signals within several genes, KIAA0564, PLD5, POU6F2, ST8SIA2 and TAF1C. © The Author 2010. Published by Oxford University Press. All rights reserved.

Background There is considerable variation in service use, with overuse in conditions such as upper respiratory tract infections and high levels of unmet need in older patients with embarrassing conditions such as incontinence. The reasons for this situation are varied but to facilitate appropriate accessing of services the decision processes involved in help-seeking require greater understanding. Objectives This study aimed to describe the decision making process for help-seeking in middle and older aged people with urinary storage symptoms. Methods In-depth qualitative interviews were carried out with 33 men and women who had sought medical care for urinary symptoms. Respondents were recruited from an epidemiological study of the prevalence of urinary symptoms in community dwelling adults aged 40 years and over. Questions were directed at help-seeking for urinary symptoms as well as help-seeking in general. Interviews were audio-taped and a thematic analysis carried out according to grounded theory methods. Results The steps in the decision process to seek medical care were drawn from the data. These consisted of appraisals of the presence and cause of symptoms, treatment options and the severity of symptoms and their impact on quality of life. Once the level of threat of the symptom was established, the costs and benefits of treatments and services were balanced against the perceived impact of symptoms on quality of life. These results were considered in relation to available models of health behaviour. Conclusions Interventions that increase individuals' active involvement in their health care and which encourage greater knowledge of symptoms and outcomes will facilitate appropriate help-seeking and service use.

Background: The introduction of intranet services in a district general hospital provided an opportunity to put evidence based national guidelines online to facilitate access and promote application of best practice in acute medical care. This study evaluated the effectiveness of this approach. Method: Local guidelines were made available online at ward terminals after they had been distributed in paper form. An interrupted time series design was used to evaluate the impact on compliance with three preselected guidelines, which addressed the management of suspected deep vein thrombosis, upper gastrointestinal bleeding, and stroke. This was supplemented by a qualitative assessment of the views of medical staff. Results: There was a significant increase in the adherence to the guidelines for stroke when they were made available online, but this was not demonstrable for deep vein thrombosis or upper gastrointestinal bleeding. Qualitative interviews with junior medical staff and consultants after the study was completed revealed that there was confusion regarding the application of the guidelines for deep vein thrombosis and little active support from the gastroenterologists for the guidelines for upper gastrointestinal bleeding. The stroke guidelines were actively promoted by their author and widely supported. Conclusion: Making guidelines available online will not be effective unless they are actively promoted and represent a consensus view.

Squalene synthase (SQS) catalyzes the first reaction of the branch of the isoprenoid metabolic pathway committed specifically to sterol biosynthesis. Regulation of SQS is thought to direct proximal intermediates in the pathway into either sterol or nonsterol branches in response to changing cellular requirements. The importance of SQS in cholesterol metabolism has stimulated research on the mechanism, structure, and regulation of the enzyme. SQS produces squalene, a C30 isoprenoid, in a two-step reaction in which two molecules of farnesyl diphosphate are condensed head to head. Site-directed mutagenesis of rat SQS has identified conserved Tyr, Phe, and Asp residues that are essential for function. The aromatic rings of Tyr and Phe are postulated to stabilize carbocation intermediates of the first and second half-reactions, respectively; the acidic Asp residues may be required for substrate binding. SQS activity, protein level, and gene transcription are strictly and coordinately regulated by cholesterol status, decreasing with cholesterol surfeit and increasing with cholesterol deficit. The human SQS (hSQS) gene has an unusually complex promoter with multiple binding sites for the sterol regulatory element binding proteins SREBP-1a and SREBP-2, and for accessory transcription factors known to be involved in the control of other sterol-responsive genes. SREBP-1a and SREBP-2 require different subsets of hSQS regulatory DNA elements to achieve maximal promoter activation. Current research is directed at elucidating the precise contribution made by individual SREBPs and accessory transcription factors to hSQS transcriptional control.

Squalene synthase (SQS) catalyzes the first reaction of the branch of the isoprenoid metabolic pathway committed specifically to sterol biosynthesis. Regulation of SQS is thought to direct proximal intermediates in the pathway into either sterol or nonsterol branches in response to changing cellular requirements. The importance of SQS in cholesterol metabolism has stimulated research on the mechanism, structure, and regulation of the enzyme. SQS produces squalene, a C 30 isoprenoid, in a two-step reaction in which two molecules of farnesyl diphosphate are condensed head to head. Site-directed mutagenesis of rat SQS has identified conserved Tyr, Phe, and Asp residues that are essential for function. The aromatic rings of Tyr and Phe are postulated to stabilize carbocation intermediates of the first and seconds half-reactions, respectively; the acidic Asp residues may be required for substrate binding. SQS activity, protein level, and gene transcription are strictly and coordinately reagulated by cholesterol status, decreasing with cholesterol surfeit and increasing with cholesterol deficit. The human SQS (hSQS) gene has an unusually complex promoter with multiple binding sites for the sterol regulatory element binding proteins SREBP-1a and SREBP-2, and for accessory transcription factors known to be involved in the control of other sterol-responsive genes. SREBP-1a and SREBP-2 require SREBP-2 required different subsets of hSQS regulatory DNA elements to achieve maximal promoter activation. Current research is directed at elucidating the precise contribution made by individual SREBPs and accessory transcription factors to hSQS transcriptional control.

Farnesyl diphosphate synthase (FPPS: EC2.5.1.10), a key enzyme in isoprenoid metabolic pathways, catalyzes the synthesis of farnesyl diphosphate (FPP) an intermediate in the biosynthesis of both sterol and non-sterol isoprenoid end products. The localization of FPPS to peroxisomes has been reported (Krisans, S. K., J. Ericsson, P. A. Edwards, and G. A. Keller. 1994. J. Biol. Chem. 269: 14165;-14169). Using indirect immunofluorescence and immunoelectron microscopic techniques we show here that FPPS is localized predominantly in the peroxisomes of rat hepatoma H35 cells. However, the partial release of 60;-70% of cellular FPPS activity is observed by selective permeabilization of these cells with digitonin. Under these conditions, lactate dehydrogenase, a cytosolic enzyme, is completely released whereas catalase, a known peroxisomal enzyme, is fully retained. Digitonin treatment of H35 cells differentially affects the release of other peroxisomal enzymes involved in isoprenoid metabolism. For instance, mevalonate kinase and phosphomevalonate kinase are almost totally released (95% and 91%, respectively), whereas 3-hydroxy-3-methylglutaryl-CoA reductase is fully retained. Indirect immunoflourescence studies indicate that FPPS is localized in peroxisomes of Chinese hamster ovary (CHO)-K1 cells but is dispersed in the cytosol of ZR-82 cells, a mutant that lacks peroxisomes. Unlike in H35 cells, FPPS is completely released upon digitonin permeabilization of CHO-K1 and ZR-82 cells. In contrast, under the same permeabilization conditions, catalase is fully retained in CHO-K1 cells but completely released from ZR-82 cells. These studies indicate that FPPS and other enzymes in the isoprenoid biosynthetic pathways, involved in the formation of FPP, are differentially associated with peroxisomes and may easily diffuse to the cytosol. Based on these observations, the significance and a possible regulatory model in the formation of isoprenoid end-products are discussed.

The tropomyosin I(TmI) gene of Drosophila melanogaster encodes two isoforms of tropomyosin. The Ifm-Tml isoform is expressed only in indirect flight and jump muscles; the Scm-TmI isoform is found in other muscles of the larva and adult. The level of Ifm-TmI is severely reduced in the flightless mutant Ifm(3)3, which also is unable to jump. To explore the functional significance of tropomyosin isoform diversity in Drosophila, we have used P element-mediated transformation to express Scm-TmI in the indirect flight and jump muscles of Ifm(3)3 flies. Transformants gained the ability to jump and fly. The mechanical properties of isolated indirect flight muscle myofibres, and the ultrastructure of indirect flight and jump muscles from the transformants were comparable to wildtype. Thus, the Scm-TmI isoform can successfully substitute for Ifm-TmI in the indirect flight and jump muscles of the Ifm(3)3 strain.

We have studied the structure and function of muscle fibers in which tropomyosin stoichiometry has been reduced by genetic mutation. We used a Drosophila melanogaster flightless mutant Ifm(3)3 and a genetic cross of this mutant with wild type flies to achieve a gradation of tropomyosin gene dosage. We measured the flight ability and wingbeat frequency of the live insects and the ultrastructure and mechanochemistry of isolated single flight muscle fibers. Flight ability is impaired when tropomyosin gene dosage is reduced. Wingbeat frequency also depends upon gene dosage as well as the severity of myofilament lattice disruption and the number of myofilaments in the organized core of the myofibrils. A reduction in number of myofilaments appears to result in a reduction in active muscle stiffness without resulting in an appreciable change in kinetics of force production. Ifm(3)3 is trapped in a relaxed state and cannot generate active force. However, tight-binding rigor cross-bridges are able to form; in the absence of ATP, Ifm(3)3 muscle fibers have high stiffness and force.

The steady decline in plasma progesterone level that occurs during the last week of pregnancy in the normal rat (Wiest, '70) provides good opportunity to study the effect of withdrawal of progesterone on uterine differentiation. Evidence is presented that tissue monocytes, heterophils, and eosinophils are regular components of the normal late gestational uterus and that their number increases as term approaches. Uterine monocytes and heterophils are located in the endometrial and myometrial stroma as well as within the basal intercellular compartment of the luminal epithelium. Stromal monocytes are distributed throughout the attenuated endometrium of late gestation, but are more common immediately beneath the luminal epithelium. In the myometrium, monocytes and heterophils occur, often as perivascular clusters, in the connective tissue septum that separates the two layers of smooth muscle. Eosinophils are present especially in the deep endometrial and myometrial stroma, and increase in number as plasma estrogen rises immediately before parturition. A small population of lymphocytes is regularly present. An important feature of the prepartum uterine stroma is the sparseness of macrophages. Near term, however, the beginnings of monocytic-macrophagic transformation are noticeable as the cell surface becomes more irregular and organelles associated with endocytic activity arise. The prepartum monocytes are positioned in the same histological sites that during the postpartum period of regression will be occupied by macrophages (Padykula and Campbell, '76). Since it is generally accepted that monocytes are precursors of macrophages, this spatial correlation raises the possibility that cellular preparations for regression commence before birth. The possible significance of prepartum monocytic infiltration is discussed in relation to the effect of changing plasma and uterine concentrations of progesterone on uterine collagenase activity. The steady increase in uterine tissue leucocytes which occurs concomitantly with decreasing uterine binding capacity for progesterone supports the hypothesis by Siiteri et al. ('77) that progesterone in high local concentrations has an antiinflammatory effect.

Rapid collagen breakdown in the postpartum rat uterus is accompanied by rising collagenase activity (Jeffrey and Gross, '70) and a transient infiltration of the stroma by heterophils, eosinophils, monocyte-macro-phages, lymphocytes, and plasma cells (Padykula and Campbell, '76), cells usually associated with inflammatory response. This uterine catabolism is initiated soon after birth while blood estrogen and progesterone levels are low. To investigate the hormonal factors involved in regulation of this postpartum stromal differentiation, we analyzed the cytological effects of experimentally elevating progesterone and estradiol levels in the peripartum period by following the protocol of biochemical experiments that have demonstrated inhibition of collagenase activity by progesterone (Koob and Jeffrey, '74) and estradiol (Ryan and Woessner, '74). Prolonged gestation (progesterone, 10 mg/day starting on day 19 gestation) was used as a condition to prevent the prenatal drop in blood progesterone; this treatment was the most effective in blocking postpartum stromal differentiation. It preserved the state of prepartum uterine differentiation and most importantly it prevented monocytic-macrophagic conversion. Progesterone (40 mg/day) given at birth delayed but did not block stromal differentiation during the first 48 hours; by 72 hours collagen loss was extensive in both control and progesterone-treated rats and numerous macrophages were present. Estradiol (100 μg/day) given at birth caused a greater delay in stromal differentiation than progesterone given at birth; for approximately 48 hours the number of eo-sinophils, heterophils, and macrophages was less than normal. By day 3 the number and distribution of the macrophages resembled that of the day 1 control uteri. Overall these experiments indicate that the low estrogen and progesterone levels at birth are essential for normal stromal regression. Since these transient cells originate from the blood, the temporal pattern of their emigration into the uterus may be under hormonal control. Experimental disturbance of this pattern influences the course of collagen resorption.

The extent to which veterinary pathogenic and human opportunistic species of thermophilic and thermotolerant fungi disseminate in aerosols from heated effluents of nuclear production reactors of the Savannah River Plant (SRP), South Carolina, has been measured. Aerosol samples were taken at 140 sites, from directly over thermal effluents to more than 100 kilometers from the SRP boundary. Sampling methods included settle plates (134 sites, 1, 103 plates for a total of 2, 805 hrs.), liquid impingement (78 sites, 1 cubic meter each), filtration (79 sites, 1 cubic meter each), and a particle sizing cascade impactor (Andersen Sampler) (17 sites, 1 cubic meter each). Soils, foams, and microbial mats from thermal effluents (a total of 191 samples), and vegetation (560 samples) were sampled to study distribution of particular species. Sampling was done under a variety of conditions: hot weather and cold, wet and dry, day and night, windy and calm, reactor(s) operating and not, disturbed vegetation and undisturbed. At 102 of the aerosol sampling sites, sophisticated meterological analyses were used to allow sampling of air in the plume which originated from thermal effluents. Soil, foam, microbial mat, vegetation, and aerosol samples were quantitatively plated for detection of viable units; filters were halved and then both plated and observed microscopically. Significant dissemination of thermophilic and thermotolerant fungi from thermal effluents was not detected. Thermophilic and thermotolerant fungi were widely distributed in soil, air, and on vegetation. Dactylaria gallopava, the indicator species and dominant fungus in microbial mats lining SRP thermal effluents and the cause of epidemic fatal phaeohyphomycosis in flocks of turkeys and chickens in South Carolina, Georgia, and elsewhere, was isolated from air at a maximum of 50 meters from effluents (8 viable units per cubic meter of air in the plume of an effluent during a strong wind).

Thermophilic and thermotolerant fungi were isolated from samples of domestic and municipal compost piles, self-heated industrial wood chip piles, alligator nests, sun-heated plant debris and soil, ...

Four monoclonal antibodies (MAbs) were produced by immunizing mice with a disrupted yeast cell homogenate of Histoplasma capsulatum. MAbs 1 and 2 reacted only with the yeast cell antigens of H. capsulatum and Blastomyces dermatitidis, whereas MAbs 3 and 4 showed broader cross-reactivity. MAb 3 cross-reacted with B. dermatitidis, Paracoccidioides brasiliensis, Sporothrix schenckii, and Candida albicans, and MAb 4 cross-reacted with B. dermatitidis, C. albicans, Coccidioides immitis, Aspergillus fumigatus, and Mycobacterium tuberculosis. All four MAbs exhibited unique specificity when reacted with three different strains of H. capsulatum (G217B, A811, and P-IN). MAb 1 belonged to the IgG2b subclass, MAb 3 belonged to the IgG1 subclass, and MAbs 2 and 4 belonged to the IgG3 subclass. MAbs 1, 2, and 3 formed bands in the Western immunoblot assay; the two dominant distinct bands had apparent molecular masses of 72 and 62 kilodaltons.

A Feulgen staining procedure was used to stain nuclei of sporangiospores ofRhizomucor miehei and R. pusillus. Most of the sporangiospores of all of the 18 different strains of R. miehei, and some ofthe 12 strains of R. pusillus examined, contained one nucleus, but occasional (sometimes rare) sporangiospores of all strains contained more than one (usually two). Some strains of R. pusillus, however, consistently produced sporangiospores having more than one nucleus each. Effects on nucleus number of growth temperature, age of cultures, growth media, single-sporing by micromanipulation, repeated sampling with different protocols, origin of strains, mating system of strains, and treatment with a potential haploidizing agent are also described.

Rhizomucor miehei (Cooney & Emerson) Schipper, R. pusillus (Lindt) Schipper, Rhizopus rhizopodiformis (Cohn) Zopf, and Absidia corymbifera (Cohn) Sacc. & Trotter were isolated from numerous materials immersed in YpSs agar or 2% malt extract agar containing 50 μg/ml Benlate, 100 U/ml penicillin G, and 40 μg/ml streptomycin sulfate; agar plates were incubated at 45 °C. Alternative media and procedures useful for special circumstances are described. These four pathogenic thermophilic or thermotolerant species were readily isolated from spices, herbal teas, sunflower seeds, and other materials.

We report the performance of an optically pumped submillimeter wave laser incorporating a Michelson interferometric output coupler consisting of three mirrors and a metal mesh beam splitter. 10.33-μm radiation from a pulsed CO 2 TEA laser was used to pump NH 3 for generating submillimeter wave radiation at 281 μm. Optimum output was achieved by coupling out 59 percent of the radiation per pass using a 250-lines-per-inch mesh at a phase angle of 49°. The submillimeter laser operated in the TEM 00 mode and was tunable over its free spectral range of about 350 MHz. Translation of the coupling mirror allowed optimization of laser output.

Thermophilic fungi isolated from partially decomposed vegetation from the interior of a recently used alligator (Alligator mississipiensis) nest included Chaetomium thermophile var. coprophile Cooney & Emerson, C. thermophile var. dissitum Cooney & Emerson, Humicola lanuginosa (Griffon & Maublanc) Bunce, Talaromyces thermophilus Stolk, and Thermoascus aurantiacus Miehe sensu Apinis. The thermotolerant fungi Aspergillus fumigatus Fresenius and Burgoa-Papulaspora sp. were also isolated. The ontogeny of bulbils of the latter isolate is described. Alligator nesting material provides a habitat suitable for the thermophilic fungi, where the heat required for growth of these fungi is not provided by the body heat of the nest builder. These data suggest that habitats suitable for the origin and evolutionary survival of obligately thermophilic fungi have existed for long periods of time.

Thermophilic fungi isolated from self-heated wood chips stored at a paper factory included Chaetomium thermophile var. coprophile Cooney & Emerson, C. thermophile var. dissitum Cooney & Emerson, Hu...

SUMMARY Acid spring effluents are often covered with mats of the eucaryotic phycocyanin-containing alga. Cyanidium caldarium. The primary bacterial component of such mats is an acidophilic strain of Bacillus coagulans, and the primary fungal component is Dactylaria gallopava. Because of the limited species diversity, C. caldarium mats appeared to be an excellent system for studying algal excretion and various microbial interactions in nature. From 2 to 6% of the NaH14CO3 taken up by natural or laboratory populations of the alga was excreted as 14C-labeled materials. The maximum excretion occurred at temperature, light, and pH values optimum for NaH14CO3 uptake. However, when excretion was expressed as a percentage of NaH14CO3 uptake, a higher percentage of the radioactivity was excreted at nonoptimal conditions for NaH14CO3 uptake. Fungal biomass was directly proportional to algal density, but bacterial numbers varied widely and did not correlate with algal numbers. The bacterial and fungal components could be grown in mixed culture with either growing C. caldarium cultures or in an extract prepared, by healing algal cells.

Rates of growth of seven strains of Chaetomium thermophile var. dissitum and one strain of C. thermophile var. coprophile were determined over a wide range of temperature by measurement of increase in colony diameter on Emerson YpSs Agar. After a lag phase, increase in diameter occurred at a constant rate. Growth rates and cardinal points differed among the strains. Ability to grow, form mycelial pigmentation, and produce immature and mature perithecia at extreme temperatures differed significantly among the strains for a given character, and within a strain for different characters. The range of temperature suitable for sexual reproduction was small and was much less than that for vegetative growth and pigmentation. The two taxonomic varieties differed significantly from one another in their responses to extreme temperatures, and one strain of variety dissitum had lower growth rates and more restricted morphogenetic capacity than other strains of the variety.

A new thermophilic species of Thielavia Zopf, T. australiensis, isolated from nesting material of an incubator bird (mallee fowl, Leipoa ocellata Gould), is described and illustrated.

The patterns of proteins synthesized in developing Spisula embryos and larvae were compared with in vitro translation products by one-dimensional gel electrophoresis. Major changes in the in vivo pattern occur at fertilization; these are regulated at the translational level (Rosenthal, Hunt, and Ruderman, 1980, Cell 20, 487-494). The pattern is further altered by midcleavage, and subsequent development is accompanied by frequent changes in the kinds of proteins made. By midcleavage many of the in vivo changes are paralleled by alterations in mRNA levels. Three cDNA clones containing developmentally regulated, nonmitochondrial sequences were isolated from a library constructed from veliger larval RNA. Clone 3v4 encodes alpha-tubulin. Clone 12v4 encodes a 35,000-D protein of unknown function. The protein product of clone 10v8 has not been identified. The concentration of alpha-tubulin RNA is relatively low through midcleavage, increases by the swimming gastrula stage, and is maintained at a moderately high level throughout larval development. 10v8 and 12v4 RNAs first appear in trochophore larvae; their concentrations peak 10-12 hr later, and then decline. The proportions of alpha-tubulin and 10v8 RNA that are translated vary with developmental stage. During early cleavage very little alpha-tubulin RNA is on polysomes; in swimming gastrulae 64% of this mRNA is polysomal. Seventy percent of 10v8 RNA is translated in the trochophore larva, while only approximately 40% is polysomal in the 21-hr veliger. These results show that translational regulation may be superimposed on changes in cytoplasmic mRNA concentrations to determine the level of gene expression during embryogenesis.

Fertilization of Spisula oocytes causes a rapid change in the overall pattern of protein synthesis. This change occurs independently of any new mRNA transcription: it is controlled entirely at the translational level. cDNA clones complementary to several translationally regulated mRNAs have been isolated and used to directly investigate fertilization-triggered changes in mRNA activity and structure. Several mRNAs gain a long poly(A) tail right after fertilization, whereas others lose their poly(A) tails. In general, there is a good correlation between possession of a poly(A) tail and translational activity in vivo. Except for these changes in poly(A), these mRNAs show no significant structural alterations. This result strongly suggests that the inactivity of maternal mRNAs in the oocyte is not due to their being stored as translationally incompetent larger precursor forms. Evidence for a role of masking components is also discussed.

Studies of biological responses to moonlight as distinct from lunar phases are warranted. Fungi are especially suitable organisms for this research. Moonlight is quantitatively and qualitatively similar to light which causes photoresponses of fungi.

A dramatic change in the pattern of protein synthesis occurs within ten minutes after fertilization of Spisula oocytes. This change is regulated entirely at the translational level. We have used DNA clones complementary to five translationally regulated messenger RNAs to follow shifts in mRNA utilization at fertilization and to characterize alterations in mRNA structure that accompany switches in translational activity in vivo . Four of the mRNAs studied are translationally inactive in the oocyte. After fertilization two of these mRNAs are completely recruited onto polysomes, and two are partially recruited. All four of these mRNAs have very short poly(A) tracts in the oocyte; after fertilization the poly(A) tails lengthen considerably. In contrast, a fifth mRNA, that encoding α-tubulin mRNA, is translated very efficiently in the oocyte and is rapidly lost from polysomes after fertilization. Essentially all α-tubulin mRNA in the oocyte is poly(A) + and a large portion of this mRNA undergoes complete deadenylation after fertilization. These results reveal a striking relationship between changes in adenylation and translational activity in vivo . This correlation is not perfect. however. Evidence for and against a direct role for polyadenylation in regulating these translational changes is discussed. Changes in poly(A) tails are the only alterations in mRNA sizes that we have been able to detect. This indicates that, at least for the mRNAs studied here, translational activation is not due to extensive processing of larger translationally incompetent precursors. We have also isolated several complementary DNA clones to RNAs encoded by the mitochondrial genome. Surprisingly, the poly(A) tracts of at least two of the mitochondrial RNAs also lengthen in response to fertilization.