The amino-terminal domain of ORF149 of koi herpesvirus is preferentially targeted by IgM from carp populations surviving infection.

Recombinantly expressed fragments of the protein encoded by ORF149 (pORF149), a structural protein from the common- and koi-carp-infecting cyprinid herpesvirus-3 (CyHV-3) that was previously shown to be antigenic, were used to obtain evidence that its amino-terminal part contains immunodominant epitopes in fish populations that survived the infection. To obtain such evidence, nonspecific binding of carp serum tetrameric IgM had to be overcome by a novel ELISA protocol (rec2-ELISA). Rec2-ELISA involved pre-adsorption of carp sera with a heterologous recombinant fragment before incubation with pORF149 fragments and detection with anti-carp IgM monoclonal antibodies. Only in this way was it possible to distinguish between sera from uninfected and survivor carp populations. Although IgM from survivors recognised pORF149 fragments to a lesser degree than whole virus, specificity was confirmed by correlation of rec2- and CyHV-3-ELISAs, inhibition of rec2-ELISA by an excess of frgIIORF149, ELISA using IgM-capture, Western blotting, and reduction of reactivity in CyHV-3-ELISA by pre-adsorption of sera with frgIIORF149. The similarity of IgM-binding profiles between frgIORF149 (amino acid residues 42-629) and frgIIORF149 (42-159) and their reactivities with previously described anti-CyHV-3 monoclonal antibodies confirmed that most pORF149 epitopes were localised in its amino-terminal part.

Review of brucellosis cases from laboratory exposures in the United States in 2008 to 2011 and improved strategies for disease prevention.

Five laboratory-acquired brucellosis (LAB) cases that occurred in the United States between 2008 and 2011 are presented. The Centers for Disease Control and Prevention (CDC) reviewed the recommendations published in 2008 and the published literature to identify strategies to further prevent LAB. The improved prevention strategies are described.

A two-part model for reference curve estimation subject to a limit of detection.

Reference curves are commonly used to identify individuals with extreme values of clinically relevant variables or stages of progression which depend naturally on age or maturation. Estimation of reference curves can be complicated by a technical limit of detection (LOD) that censors the measurement from the left, as is the case in our study of reproductive hormone levels in boys around the time of the onset of puberty. We discuss issues with common approaches to the LOD problem in the context of our pubertal hormone study, and propose a two-part model that addresses these issues. One part of the proposed model specifies the probability of a measurement exceeding the LOD as a function of age. The other part of the model specifies the conditional distribution of a measurement given that it exceeds the LOD, again as a function of age. Information from the two parts can be combined to estimate the identifiable portion (i.e. above the LOD) of a reference curve and to calculate the relative standing of a given measurement above the LOD. Unlike some common approaches to LOD problems, the two-part model is free of untestable assumptions involving unobservable quantities, flexible for modeling the observable data, and easy to implement with existing software. The method is illustrated with hormone data from the Third National Health and Nutrition Examination Survey.

Orbital haematoma after orbital fracture repair using silicone, polytetrafluorethylene, and poly-L-lactic acid/hydroxyapatite implants.

The purpose of this paper was to report the incidence of orbital haematoma formation following the repair of orbital fractures with silicone, polytetrafluorethylene (PTFE), and poly-L-lactic acid/hydroxyapatite (PLLA/HA) implants. This retrospective review examined 234 patients (235 sides) who underwent repair of orbital wall fractures over a six-year period. Of these, 36 patients received a silicone sheet; 49 patients received a silicone sheet with superimposed PTFE implant; and 149 patients (150 sides) received PTFE with or without an underlying PLLA/HA implant. Orbital haematomas were documented in 13 out of 36 patients (36.1%) who underwent fracture repair with a silicone sheet; seven out of 49 patients (14.3%) who had a silicone sheet with a superimposed PTFE implant; and three out of 150 sides (2.0%) with PTFE with or without underlying PLLA/HA. The difference in incidence of orbital haematoma formation was significant among the groups (p < 0.001, Pearson's chi squared test). In the silicone sheet group, haematomas formed within two weeks of surgery in 11 patients (84.6%). In the silicone sheet with superimposed PTFE implant group, five patients (71.4%) developed orbital haematomas no earlier than after two weeks postoperatively. This study showed that the incidence of orbital haematoma formation following orbital fracture repair is different among the implant materials. The use of PTFE and PLLA/HA implants may minimise this complication.

Mars Extant Life: What's Next? Conference Report.

On November 5-8, 2019, the "Mars Extant Life: What's Next?" conference was convened in Carlsbad, New Mexico. The conference gathered a community of actively publishing experts in disciplines related to habitability and astrobiology. Primary conclusions are as follows: A significant subset of conference attendees concluded that there is a realistic possibility that Mars hosts indigenous microbial life. A powerful theme that permeated the conference is that the key to the search for martian extant life lies in identifying and exploring refugia ("oases"), where conditions are either permanently or episodically significantly more hospitable than average. Based on our existing knowledge of Mars, conference participants highlighted four potential martian refugium (not listed in priority order): Caves, Deep Subsurface, Ices, and Salts. The conference group did not attempt to reach a consensus prioritization of these candidate environments, but instead felt that a defensible prioritization would require a future competitive process. Within the context of these candidate environments, we identified a variety of geological search strategies that could narrow the search space. Additionally, we summarized a number of measurement techniques that could be used to detect evidence of extant life (if present). Again, it was not within the scope of the conference to prioritize these measurement techniques-that is best left for the competitive process. We specifically note that the number and sensitivity of detection methods that could be implemented if samples were returned to Earth greatly exceed the methodologies that could be used at Mars. Finally, important lessons to guide extant life search processes can be derived both from experiments carried out in terrestrial laboratories and analog field sites and from theoretical modeling.

Mesoscopic conductors and correlations in laser speckle patterns.

Submicrometer disordered metallic systems at low temperatures display interesting conductance fluctuation effects, owing to the interference of coherent electron wave transmission through the system. This quantum interference phenomenon also gives rise to the striking experimental observation that the 1/f noise power (where f is frequency) in disordered conductors increases as the temperature is decreased, contrary to common intuition. The theoretical techniques developed for this problem can be applied to the transmission of light through a random medium, and this yields novel predictions for the correlations in the laser speckle patterns and the possibility of studying the positions and motions of scatterers in a medium which multiply scatter the probing light.

Localization, interactions, and the metal-insulator transition.

Recent advances in our understanding of electronic conduction have pointed up deficiencies in traditional thinking. For a metal at a sufficiently low temperature, it is known both theoretically and experimentally that the conventional picture in terms of the Boltzmann theory breaks down. Improved understanding of both electron localization and the effects of electron-electron interactions in a disordered medium has led to experimentally verifiable predictions. These effects have an important influence on the nature of the metal-insulator transition.

Subglacial Lake Whillans microbial biogeochemistry: a synthesis of current knowledge.

Liquid water occurs below glaciers and ice sheets globally, enabling the existence of an array of aquatic microbial ecosystems. In Antarctica, large subglacial lakes are present beneath hundreds to thousands of metres of ice, and scientific interest in exploring these environments has escalated over the past decade. After years of planning, the first team of scientists and engineers cleanly accessed and retrieved pristine samples from a West Antarctic subglacial lake ecosystem in January 2013. This paper reviews the findings to date on Subglacial Lake Whillans and presents new supporting data on the carbon and energy metabolism of resident microbes. The analysis of water and sediments from the lake revealed a diverse microbial community composed of bacteria and archaea that are close relatives of species known to use reduced N, S or Fe and CH4 as energy sources. The water chemistry of Subglacial Lake Whillans was dominated by weathering products from silicate minerals with a minor influence from seawater. Contributions to water chemistry from microbial sulfide oxidation and carbonation reactions were supported by genomic data. Collectively, these results provide unequivocal evidence that subglacial environments in this region of West Antarctica host active microbial ecosystems that participate in subglacial biogeochemical cycling.

Depot leuprolide acetate for treatment of precocious puberty.

Leuprolide acetate (D-Leu 6 des-Gly-NH2(10), Pro-ethylamide9) for depot suspension has been shown to be effective in suppressing gonadotropins in 6 patients. Serum LH and FSH response to exogenous LHRH was obliterated after 4 weeks and weekly urinary gonadotropin levels during depot therapy were suppressed. This form of this drug should be effective in the treatment of precocious puberty.

Evidence for partial 21-hydroxylase deficiency among heterozygote carriers of congenital adrenal hyperplasia.

Concentrations of 17-hydroxyprogesterone are significantly greater in heterozygous carriers of CVAH than in controls 30 and 60 minutes after an infusion of 25 units of synthetic ACTH 1-24 and 2 hours after beginning a 4-hour infusion of 50 units ACTH. The majority of carriers were clearly above the control range at these collection times. Hence, heterozygous carriers have a partial enzyme deficiency although all cannot be diagnosed based on 17-hydroxyprogesterone levels after ACTH stimulation.

Puberty in boys: correlation of plasma levels of gonadotropins (LH, FSH), androgens (testosterone, androstenedione, dehydroepiandrosterone and its sulfate), estrogens (estrone and estradiol) and progestins (progesterone and 17-hydroxyprogesterone).

Mean serum concentration dehydroepiandrosterone (DHA), DHA sulfate (DHAS), progesterone (P), 17-hydroxyprogesterone (17-OH-P), estrone (E1), estradiol (E2), and androstenedione (A) were compared from 43 boys followed longitudinally for as long as 4 yr during puberty. These data were also compared with serum levels of LH, FSH, and testosterone. Elevation is recognized early in puberty for DHAS, late in puberty for P and A, and gradually throughout puberty for E1, 17-OH-P, and DHA. When compared by age, the same general pattern is apparent with adult levels of E1 reached at age 12, DHAS and E2 by 13, and DHA, P, 17-OH-P, and A not until after age 15. Significant elevations of DHA occurred with the onset of pubic hair and voice change; elevations of DHAS occurred with the onset of genital and axillary hair growth; and testosterone increased with the onset of genital and pubic hair growth and voice change.

Gonadotropin and sex steroid response to luteinizing hormone-releasing hormone in patients with premature adrenarche.

Luteinizing hormone-releasing hormone (100 mug iv) produced a response of LH, FSH, testosterone, and estradiol in children with premature pubarche that was not different from that among normal prepubertal children. Hence, this study provides no evidence that hypothalamic-pituitary-gonadal maturation occurs with the onset of premature sexual hair.

Depot leuprolide acetate dosage for sexual precocity.

Twenty-one children with early puberty have been evaluated to determine adequate dosage of depot leuprolide acetate. The minimal dosage of depot leuprolide acetate required, using intervals of 1.5 mg or less when given every 28 days, was determined. This dosage, as determined by suppression of gonadotropin responses to GnRH stimulation, was 4.14 +/- 1.33 mg (mean +/- SD) and 0.15 +/- 0.07 mg/kg.28 days. The dosage correlates with bone age and pubertal stage and is larger among patients with more advanced puberty. Five patients required a larger dosage from 5-10 months after initial suppression. Therefore, to monitor suppression, GnRH testing should be repeated at least at 6-month intervals. Samples 20 and 40 min after GnRH stimulation are sufficient to indicate adequacy of treatment, so an abbreviated test could be used.

Lack of correlation between gonadotropin and adrenal androgen levels in agonadal children.

To evaluate the relationship between the secretion of gonadotropins and adrenal androgens, patients with gonadal agenesis were evaluated by (a) administering human luteinizing hormone (hLH) for 5 days with or without estrogen pretreatment to agonadal patients who had prepubertal LH levels; (b) correlating circulating gonadotropin levels with adrenal androgens in 45 patients; and (c) comparing adrenal androgens with gonadotropins after long-term administration of estrogen or androgens. Results are as follows: (a) No alteration in serum concentrations of dehydroepiandrosterone (DHA), dehydroepiandrosterone sulfate (DHAS), estrone (E1), testosterone (T), or in excretion of urinary 17-ketosteroid (17 KS) occurred after the administration of hLH. (b) No clearcut relationship between endogenous level of LH or FSH and DHA OR DHAS was demonstrated although a coincident increase of all hormones with age occurred. (c) Administration of estrogen to patients with gonadal agenesis did not affect their levels of DHA and DHAS although those patients given androgen developed higher DHAS, but not DHA, levels. Hence, increasing gonadotropin concentrations would not appear to be a primary etiologic factor in the maturation of the adrenal.

Treatment of central precocious puberty: comparison of urinary gonadotropin excretion and gonadotropin-releasing hormone (GnRH) stimulation tests in monitoring GnRH analog therapy.

GnRH analogs (GnRH-a) have proven to be efficacious and have become the standard treatment for central precocious puberty (CPP). To confirm the diagnosis of CPP and to monitor the adequacy of hvpothalamic-pituitary-gonadal (HPG) axis suppression, GnRH stimulation testing has been essential. To determine whether 24-h urinary gonadotropin excretion could adequately assess HPG axis suppression, we compared the results of simultaneous GnRH stimulation tests and 24-h urinary gonadotropin determinations in 18 girls with CPP who were receiving GnRH-a therapy (leuprolide acetate, Depot-Lupron, TAP Pharmaceuticals). HPG axis suppression was defined as the absence of significant LH and FSH responses to GnRH stimulation. Simultaneous GnRH stimulation tests and urinary gonadotropin determinations had a concordance rate of 68% (42 of 62). The sensitivity and specificity of urinary LH determinations to detect inadequate HPG suppression were 75% and 64%, respectively. For urinary FSH determinations, the sensitivity and specificity were 90% and 28%, respectively. Hence, single timed urine collections lacked the sensitivity and specificity to assess HPG axis suppression and, thus, cannot replace GnRH stimulation tests for monitoring the adequacy of the GnRH-a dose.

Puberty in girls: correlation of serum levels of gonadotropins, prolactin, androgens, estrogens, and progestins with physical changes.

Twenty-seven girls aged 8 to 18 were studied in a longitudinal prospective fashion. Serum samples were collected at 6 month intervals up to 4 years and radioassayed for hormones of pituitary, ovarian, and adrenal origin. A progressive elevation of luteinizing hormone (LH), follicle-stimulating hormone/FSH), estradiol (E2), dehydroepiandrosterone (DHA), and androstenedione (delta4) occurred during puberty and continued until menarche. The onset of puberty occurred concomitantly with an elevation of estrone (E1) dehydroepiandrosterone (DHA), dehydroepiandrosterone sulfate (DHAS), and 17-hydroxyprogesterone (17-OH-P). Prolactin (Prol) and progesterone (Prog) concentrations did not change during puberty until after menarche. After menarche, levels of LH and FSH were comparable with menstruating adult females. Concentrations of E2 and Prog were lower during the second half of the cycle among most regularly menstruating subjects than expected during the luteal phase. LH and Prog levels indirectly suggest that ovulation occurs in a few girls within months after menarche.

Inhibin B: comparison with indexes of fertility among formerly cryptorchid and control men.

Infertility may be a consequence of cryptorchidism. We previously reported, using a large study cohort, that 38% of formerly bilateral cryptorchid men, 10% of unilateral cryptorchid men, and 5% of the control group were infertile. Men from this cohort donated blood and semen samples for inhibin B, FSH, LH, testosterone, free testosterone, and semen analyses. Results are reported comparing the entire group; some comparisons are based on normal or low sperm density. Data are also presented for men who had fathered children or had unsuccessfully attempted paternity. Mean (+/-SD) inhibin B levels were lower for the cryptorchid men (109 +/- 59 pg/mL) than the control men (153 +/- 60; P < 0.001), and FSH levels were higher (7.4 +/- 6.2 and 4.0 +/- 3.2; P < 0.0001). Inhibin B levels correlated with all other parameters for the cryptorchid group; however, correlations for the control group were only found with gonadotropins. Among the cryptorchid men, levels were significantly greater among men with normal sperm counts than men with low sperm counts (124 +/- 47 vs. 75 +/- 48 pg/mL; P < 0.0001). No difference was present for the control group (155 +/- 61 vs. 149 +/- 63 pg/mL). When the fertile group (based on paternity) vs. the infertile group (based on attempted paternity) were compared, significant differences were found for the cryptorchid group (117 +/- 62 vs. 73 +/- 52 pg/mL; P < 0.03), but not the control group (163 +/- 62 vs. 146 +/- 73 pg/mL). These data reveal relationships not apparent among the control group of men, which includes infertile men. Inhibin B data suggest that a larger portion of formerly cryptorchid men have compromised testicular function than indicated by paternity data. Low levels of inhibin B among individuals are an indication of diminished seminiferous tubule function and thus compromised potential for fertility. Low inhibin B levels together with elevated FSH levels and decreased sperm density are indicative of a high risk of infertility.

Insulin-like 3/relaxin-like factor gene mutations are associated with cryptorchidism.

Cryptorchidism is a common anomaly of male sexual differentiation. Two phases of testicular descent are recognized, transabdominal and inguinoscrotal. With evidence that androgens and Müllerian inhibitory hormone were not completely responsible for testicular descent, the existence of a third testicular hormone mediating testicular descent was postulated. Insulin-like 3 (INSL3) [also known as relaxin-like factor (RLF) and Leydig insulin-like protein (LEY I-L)] is a member of the insulin/relaxin hormone superfamily that is highly expressed in Leydig cells. The phenotype of transgenic mice with targeted deletion of the Insl3 gene was bilateral cryptorchidism with morphological evidence of abnormal gubernacular development. With this implicit evidence that Insl3 mediates testicular descent in mice, we performed mutation detection analysis of the coding regions of the 2 exon INSL3 gene in genomic DNA samples obtained from 145 formerly cryptorchid patients and 36 adult male controls. Single-strand conformational polymorphism analysis was used for the mutation detection studies. Two mutations, R49X and P69L, and several polymorphisms were identified. Both mutations were located in the connecting peptide region of the protein. The frequency of INSL3/RLF gene mutations as a cause of cryptorchidism is low, because only 2 of 145 (1.4%) formerly cryptorchid patients were found to have mutations.

Premature pubarche: etiological heterogeneity.

Premature pubarche is characterized by pubic hair, adult type body odor, acne, and axillary hair before 8 yr of age in girls and 9.5 yr of age in boys. Causes of this premature virilization include premature adrenarche, mild errors of steroidogenesis, precocious puberty, and adrenal and gonadal tumors. To determine whether any clinical parameters are helpful in distinguishing which children should undergo further evaluation for mild congenital adrenal hyperplasia, we performed ACTH stimulation tests in 69 children with premature pubarche and 8 pubertal controls. Patients were categorized as having typical (pubic hair with or without axillary hair and body odor) or atypical (pubic hair and genital enlargement) premature pubarche. Blood samples, before and 30 min after iv bolus administration of synthetic ACTH, were obtained for progesterone, 17-hydroxypregnenolone, 17-hydroxyprogesterone, dehydroepiandrosterone, androstenedione, 11-deoxycortisol, and cortisol measurements. The patients were divided into 4 groups based on their individual responses to ACTH stimulation: premature adrenarche (no apparent defect in steroidogenesis), possible decreased 21-hydroxylase activity, possible decreased 3 beta-hydroxysteroid dehydrogenase activity, and indeterminate responses. Five of 11 (45%) children with atypical premature pubarche and 7 of 58 (12%) children with typical premature pubarche were found to have evidence of mild defects in steroidogenesis. Similar to previous reports in postpubertal women, only responses to ACTH stimulation allowed accurate classification of these patients.

Evidence for a heterozygote advantage in congenital adrenal hyperplasia due to 21-hydroxylase deficiency.

21-Hydroxylase deficiency is one of the most common inherited disorders, with carrier frequencies of approximately 10% in all world populations studied to date. The high prevalence of the mutant gene is probably due to a flanking pseudogene serving as a reservoir for mutations. Despite the potential for a high rate of de novo mutations, a founder effect for specific gene conversions is observed in most populations. We hypothesized that there was a survival advantage to 21-hydroxylase heterozygotes, and here we report endocrinological and molecular investigations to test this hypothesis. We defined 28 carriers and 22 mutation-negative controls by molecular genotyping and determined ACTH-stimulated adrenal hormone responses. We found significantly elevated cortisol responses in the carriers compared to controls (30 min cortisol levels: normal, 24.2 +/- 4.6 micrograms/dL; carrier, 28.1 +/- 4.2 micrograms/dL; P < 0.005). Cortisol has a crucial role in maintaining homeostasis, influencing differentiation, suppressing inflammation, and effecting cross-talk among the immune, nervous, and endocrine systems. The brisk cortisol response we have documented in carriers of 21-hydroxylase may enable a rapid return to homeostasis in response to infectious, inflammatory, or other environmental stresses and may protect from inappropriate immune responses, such as autoimmune diseases.