Arcsin Transformation Research Articles

A systematic review of research on illicit substance use and a meta-analysis of its prevalence among college students in Türkiye

In this study, we attempted to systematically review and meta-analysis the prevalence and risk factors for illicit substance use (ISU) in Türkiye. Based on the PRISMA 2020 statement and checklist, we considered only full-text research articles on ISU among college students (CS) in Türkiye. We searched relevant research via Web of Science, PubMed, MEDLINE, Scopus, ScienceDirect, Google Scholar, and ULAKBIM between 11.06.2022-12.31.2022. We applied double arcsin transformation to the prevalence values and used the LFK index and trim and fill method to measure publication bias. In addition, we generated doi and funnel plots of the prevalence values reported in the selected studies. Among systematically reviewed 53 studies, we identified and meta-analyzed ISU prevalence in 37 studies. The mean prevalence of ISU was determined to be 0.06 (95% CI: 0.04 to 0.07). We discovered that cannabis was the most commonly used substance, with an aggregate prevalence rate of 0.044 (95% CI: 0.026 to 0.066). Common risk factors for ISU were found to be gender, parental education, income, mental problems/suicide/attempted suicide, ISU in family members/relatives, smoking, and alcohol use. Despite the lower prevalence of ISU among students in Türkiye, the research findings emphasized that it has recently become a significant public health problem among the young.

In vitro culture of nodal cuttings in kiwifruit

This study aimed to determine the effects of plant growth regulators and different medium on in vitro propagation of ˈHaywardˈ and ˈZespri Goldˈ kiwifruits. Micro-cuttings were used as explant sources and cultured in MS and WPM medium containing different concentrations of BA and GA3. Subcultures were carried out at intervals of 4-6 weeks. Contamination rate (%), blackening rate (%), callus development rate (%), shoot rate (%), and propagation coefficient (plantlet/explant) were determined. The micropropagation experiment was carried out according to the randomized plot design. The arc-sin transformation was applied to the percentage values before analysis. Significance levels of the means were compared with the LSD test. It was determined that 0.1% mercury chloride application increased the sterilization efficiency. As the BA concentration in the medium increased, the rate of callus formation of the explants increased. While there was no difference in shoot rates of explants cultured on MS and WPM medium, the propagation rate was higher in MS medium. The propagation coefficient of the ˈHaywardˈ kiwi cultivar was 3.08 in MS medium containing 1.0 mgL-1 BA + 0.5 mgL-1 GA3. MS medium can be recommended as a basic medium for in vitro kiwif propagation.

Biological effectiveness of fungicides Rango, Tacora, Velficur and Opus for control of Karnal Bunt

Commercial fungicides Rango, Tacora, Velficur, and Opus were evaluated in the field, to determine their biological effectiveness to control karnal bunt (Tilletia indica) of wheat. A completely randomized design was used with four replications. Twenty heads of cultivar Tacupeto F2001 were inoculated during the boot stage with an allantoid sporidial suspension (10,000/mL). Commercial rates indicated in the containers of each product were followed. The first application was carried out ten days after inoculation (Zadoks stages 56-58), and the second one ten days later. Inoculated spikes were threshed by hand and the healthy and infected kernels were counted to determine the percentage of infection. Two hundred grains per treatment and per replication were weighed. The biological effectiveness of the products evaluated were Opus 91.6, Velficur 83.2, Tacora 80.2, and Rango 70.3%. The untreated inoculated check had a mean of 18.6% infection. There were no statistical differences for the products evaluated for level of infection after arcsin transformation (Tukey, p = 0.05) and the coefficient of variation was 16.2%. The average weight of 200 grains per treatment was 10.6, 10.5, 10.6, and 10.7 g, respectively, and 10.7 g for the untreated inoculated check. No phytotoxic effects of treatments applied to the wheat plant were observed.

Meningitis Risk in Patients with Inner Ear Malformations after Cochlear Implants: A Systematic Review and Meta-Analysis.

The objective of this study is to determine the rate of postoperative meningitis after cochlear implantation in those with inner ear malformations (IEMs) via meta-analysis. Medline, EMBASE, and the Cochrane Library. This study was reported following the preferred reporting items for systematic reviews and meta-analyses (PRISMA) checklist. Proportion meta-analysis was conducted through an inverse variance random-effect model based on arcsin transformation and presented as forest plots. Quality assessment of the included studies was performed through the National Institutes of Health Quality Assessment Tool. Overall, 38 of 2966 studies met the inclusion criteria and were included in the analysis. There were 10 cases of meningitis after cochlear implantation in 1300 malformed ears. The overall rate of meningitis after cochlear implantation in IEMs was 0.12% (95% confidence interval, 0.006-0.380%; I2 = 0%). Cases occurred in incomplete partition (n = 5), Mondini deformity (n = 2), common cavity (n = 2), and enlarged internal auditory canal (n = 1). Six of 10 cases of postoperative meningitis occurred with an intraoperative cerebrospinal fluid leak. In those with IEMs, the risk of meningitis after cochlear implantation is very low.

Effect of bovine seminal plasma Binder of Sperm Protein 1 on stallion sperm cryopreservation

Proteins present in seminal plasma directly interact with the plasma membrane of spermatozoa, being important for successful sperm maturation and fertilization. The aim of this work was to evaluate the effect of Binder of Sperm Protein 1 (BSP1) from the seminal vesicle of bulls on semen quality of stallions and its effect on freezability of this semen. The BSP1 protein was isolated using the ammonium sulfate technique and lyophilized. Protein presence was observed at the highest concentration of BSP1 (98%) confirmed by SDS-PAGE and Western blot. The effect of addition of BSP1 to stallion semen was tested at concentrations 0 (control), 10, 20 and 40 μg/mL by addition to a commercial freezing extender. Semen from 4 stallions with 5 ejaculates per stallions was used and cryopreserved using a programmable machine (TK 3000®). After thawing (37°C / 30 seconds), samples were submitted to sperm kinematic analysis by CASA (time 0 hour and Thermo Resistance Test - TRT of 1 hour at 37°C), integrity of plasma membranes (iMP) and acrosomal membrane integrity (iAC; epifluorescence microscopy. The percentage values were subjected to arcsin transformation and means were compared using T-test at 5% probability comparing time 0 vs. TRT of 1 hour. No difference was observed for iMP andiAC regardless of BSP1 concentration. At time 0h there was a difference at the highest BSP1 concentration (40 μg/mL) (P <0.05) for ALH and BCF (control 2,53 ± 0,08a and 40 μg 2,40 ± 0,07b). After 1h TRT, the concentration of 40 μg / mL continued to present a significant difference. Hyperactivation is characterized by an increase in average lateral head displacement (ALH, mm), and beat cross frequency (BCF, Hz), possibly due to an increase in calcium influx. Our results indicate exactly the opposite with a significant reduction in these two parameters. These results suggest that in the horse BSP1 acts modulating calcium influx. Mitochondrial function determined by JC-1 showed a significant difference at the concentration of 10 μg/mL of the protein, lower than the other groups. As the bovine has a higher fertility compared to stallion, these results, in addition to the novelty of the work, allow further studies using proteins from heterologous species with the aim of improving seminal quality in stallions. In conclusion, BSP1 may exert an antioxidant effect in relation to the mitochondrial potential of sperm and change sperm kinematics due to a modulation of calcium influx by BSP1 protein. Further investigations are required to evaluate intracellular calcium concentrations after addition of BSP1.

BIOLOGICAL EFFECTIVENESS OF FUNGICIDES PRIORI XTRA, FOLICUR, APPROACH PRIMA, AND MAXTROBIN XTRA FOR CONTROL OF KARNAL BUNT

Commercial fungicides Approach Prima CS, Priori Xtra CS, Maxtrobin Xtra CS, and Folicur EW were evaluated in the field, to determine their biological effectiveness to control karnal bunt (Tilletia indica) of wheat. A completely randomized design was used with four replications. Twenty heads of cultivar Tacupeto F2001 were inoculated during the boot stage with an allantoid sporidial suspension (10,000/mL). Commercial rates indicated in the containers of each product were followed. The first application was carried out ten days after inoculation {Zadoks 56-58, (Feekes 10.4-10.5)}, and the second one ten days later. Inoculated spikes were threshed by hand and the healthy and infected kernels were counted to determine the percentage of infection. The biological effectiveness of the products evaluated were Priori Xtra 99.2, Folicur 98.9, Approach Prima 98.5, and Maxtrobin Xtra 85.3%. The untreated inoculated check had a mean of 36.9% infection. There were no statistical differences for the products evaluated for level of infection after arcsin transformation (Tukey, p = 0.05) and the coefficient of variation was 6.1%. The a thousand grain weight was 55.3, 51.1, 54.8, and 55.1 g, respectively, and 53.6 g for the untreated inoculated check. No phytotoxic effects to the wheat plants were observed.

THE PREVALENCE OF HIV RECOMBINANT FORMS IN RUSSIA AND COUNTRIES OF THE CIS: SYSTEMATIC REVIEW AND METAANALYSIS

This review was to aimed to access the prevalence of HIV recombinant forms in Russia and countries of the CIS, which have close social and economic ties. We conducted a search in Russian Science Citation Index and PubMed for a depth of 8 years. We included 22 articles, which contained the results of 35 independent cross-sectional studies, in our review. Meta-analysis of HIV recombinant forms prevalence was conducted in Open Meta-analyst with the use of Der Simonian Laird method, arcsin transformation and correction factor for zero values. Subgroup analysis was used along with meta-regression (by date of collection). Pooled prevalence of HIV recombinant forms was 21.3% (95% CI 16.2–26.5) and was highly heterogeneous. Blood samples from 3,494 HIV patients living in various regions of the Russian Federation and CIS countries — Belarus, Kazakhstan, Kyrgyzstan, Uzbekistan, Tajikistan, Armenia, Georgia — were examined. Among subtypes of HIV-1, the main share was occupied by subtype A — 75.6% (n = 2643), subtype B — 5.5% (n = 193), C — 0.8% (n = 31), G — 0.2% (n = 7), F — 0.14% (n = 5). Among the recombinant forms (n = 616), the fraction of CRF02_ AG was 39.6% (n = 244), CRF02_AG/A was 32.9% (n = 203), CRF63_02A1 was 15.9% (n = 98), CRF03_AB — 5.2% (n = 32), CRF06_cpx — 2.1% (n = 13). Unique recombinant forms were 2.7% (n = 17), including URF63_A1 (2.5%). In Russia, the highest prevalence of recombinant forms of HIV-1 was registered in the Siberian Federal District (33.2% (95% CI 12.2– 54.1), the lowest in the Northwest Federal District — 1.6% (95% CI 0.9–2.3). In the CIS countries, the highest prevalence of recombinant forms of HIV-1 is found in a subgroup of the Central Asian republics — Uzbekistan, Tajikistan, Kyrgyzstan, Kazakhstan — 51.7% (95% CI 38.5–64.9). The significant upward trend was shown with a meta-regression. Our review is the first such research in Russia and thus is of significant interest, but the result should be applied with caution due to high risk of publication bias as well as significant heterogeneity of our results.

22 Comparing Three Extenders: Hashi, Green Buffer and INRA 96, for Chilled Storage of Bactrian Camel Semen

Semen preservation remains challenging in the camel industry. The objective of the present study was to compare 3 different extenders for chilled storage of Bactrian camel semen. Semen (n = 9 ejaculates) was collected from camel bulls (n = 2) manually using artificial vagina. Good neat semen, as far as mass vibration concerns, was equally distributed into 3 double-wall vessels filled with 35°C water. The 3 extenders used in the present study were Hashi, Green buffer (IMV, L’Aigle, France), and INRA 96 (IMV). The Hashi extender consisted of Tris, 2.6%; citric acid, 1.35%; glucose, 0.9%; fructose, 0.9%; penicillin G sodium, 1000 IU mL−1; streptomycin sulfate, 1000 mg mL−1 supplemented with 20% plasma egg yolk and 20% camel skim milk; osmolality of 330 and pH of 6.9). Green buffer was supplemented with 20% plasma egg yolk (osmolality of 335 and pH of 6.9). The osmolality and pH of INRA 96 were 310 and 7, respectively. Extenders at a ratio of 1:3 were added to semen followed by pipetting 10 times with an automatic pipettor. The water-jacketed extended specimen was covered with foam and transferred to individual vaccine carrier equipped with 4 ice packs. This system of cooling not only allows the specimen to cool down slowly and reach 4°C after 7 h, but also reduces the viscosity of camel semen. The assessment was carried out 7 and 24 h after semen dilution, when the specimen reached 4°C. Semen viability parameters were assessed after short-term semen preservation in different extenders. Total motility and progressive forward motility were examined subjectively by single operator using Sperm Track (ISAS, Proiser, Spain) after diluting the specimen to achieve 25 × 106 sperm mL−1. Live percentage of sperm was estimated using Eosin B Fast Green staining method. Plasma membrane integrity was assessed using the hypo-osmotic swelling (HOS) test. Following arcsin transformation, data were analysed by GLM procedure followed by Tukey test in SAS (SAS Institute Inc., Cary, NC, USA). At 7 and 24 h, there were no differences among the 3 extenders in total motility of sperm (Hashi: 73 and 67.4%; Green buffer: 71.6 and 62.1%; INRA 96: 70 and 66.2%; P &gt; 0.05), live percentage of sperm (Hashi: 76 and 73%; Green buffer: 70.5 and 65.6%; INRA 96: 77.8 and 70.7%; P &gt; 0.05), or HOS test (Hashi: 52.4 and 45.2%; Green buffer: 49.6 and 40.6%; INRA 96: 57.3 and 51.1%; P &gt; 0.05). However, at the same times, progressive forward motility was similar between Hashi (47.7 and 28.6%) and Green buffer (40 and 23.5%; P &gt; 0.05) but was different between Hashi and INRA 96 (23.6 and 16.7%; P &lt; 0.05). In conclusion, Hashi and Green buffer could be considered suitable extenders to preserve Bactrian camel semen under chilled condition. Further studies with a larger number of bulls and ejaculates are warranted.

Hierarchical probabilistic models for multiple gene/variant associations based on next-generation sequencing data.

MotivationThe identification of genetic variants influencing gene expression (known as expression quantitative trait loci or eQTLs) is important in unravelling the genetic basis of complex traits. Detecting multiple eQTLs simultaneously in a population based on paired DNA-seq and RNA-seq assays employs two competing types of models: models which rely on appropriate transformations of RNA-seq data (and are powered by a mature mathematical theory), or count-based models, which represent digital gene expression explicitly, thus rendering such transformations unnecessary. The latter constitutes an immensely popular methodology, which is however plagued by mathematical intractability.ResultsWe develop tractable count-based models, which are amenable to efficient estimation through the introduction of latent variables and the appropriate application of recent statistical theory in a sparse Bayesian modelling framework. Furthermore, we examine several transformation methods for RNA-seq read counts and we introduce arcsin, logit and Laplace smoothing as preprocessing steps for transformation-based models. Using natural and carefully simulated data from the 1000 Genomes and gEUVADIS projects, we benchmark both approaches under a variety of scenarios, including the presence of noise and violation of basic model assumptions. We demonstrate that an arcsin transformation of Laplace-smoothed data is at least as good as state-of-the-art models, particularly at small samples. Furthermore, we show that an over-dispersed Poisson model is comparable to the celebrated Negative Binomial, but much easier to estimate. These results provide strong support for transformation-based versus count-based (particularly Negative-Binomial-based) models for eQTL mapping.Availability and implementationAll methods are implemented in the free software eQTLseq: https://github.com/dvav/eQTLseqSupplementary information Supplementary data are available at Bioinformatics online.

Sharing Local Public Health Services Across Jurisdictions: Comparing Practice in 2012 and 2014.

Describe cross-jurisdiction service sharing (CJS) by local and tribal health departments (LHD) in Wisconsin in 2014 compared to 2012. An online survey of 91 LHD directors in Wisconsin was conducted. Results were compared to the results of a 2012 survey. Characteristics of CJS arrangements and differences in results by population size, geographic region, and governance type were described. Standardized proportion differences (h) were estimated using the arcsin transformation. Confidence intervals were estimated using unconditional exact confidence intervals for the difference of proportions.8 A forest plot of the estimates and confidence intervals was generated to visualize change in CJS for each population category. Seventy-eight percent of respondents in 2014 reported currently sharing services compared to 71% of respondents in 2012. Positive effect sizes indicate increased sharing in year 2014 relative to 2012. CJS was more frequent for LHD serving smaller jurisdictions, consistent with both 2012 survey results and national findings. All governance types continue to engage in sharing public health services. Cross jurisdictional service sharing is widespread and increasing in Wisconsin, implying that it is a useful strategy for providing public health services under some circumstances. Educating public health practitioners and students about CJS strategies in public health is recommended.

Comment on ‘An improved score interval with a modified midpoint for a binomial proportion’

Yu et al. [An improved score interval with a modified midpoint for a binomial proportion. J Stat Comput Simul. 2014;84:1022–1038] propose a novel confidence interval (CI) for a binomial proportion by modifying the midpoint of the score interval. This CI is competitive with the various commonly used methods. At the same time, Martín and Álvarez [Two-tailed asymptotic inferences for a proportion. J Appl Stat. 2014;41:1516–1529] analyse the performance of 29 asymptotic two-tailed CI for a proportion. The CI they selected is based on the arcsin transformation (when this is applied to the data increased by 0.5), although they also refer to the good behaviour of the classical methods of score and Agresti and Coull (which may be preferred in certain circumstances). The aim of this commentary is to compare the four methods referred to previously. The conclusion (for the classic error α of 5%) is that with a small sample size (≤80) the method that should be used is that of Yu et al.; for a large sample size (n ≥ 100), the four methods perform in a similar way, with a slight advantage for the Agresti and Coull method. In any case the Agresti and Coull method does not perform badly and tends to be conservative. The program which determines these four intervals are available from the address http://www.ugr.es/local/bioest/Z_LINEAR_K.EXEhttp://www.ugr.es/local/bioest/Z_LINEAR_K.EXE.

208 CLONING AND CHARACTERIZATION OF BUFFALO INTERFERON-TAU AND EFFICACY OF RECOMBINANT BUFFALO INTERFERON-TAU FOR IN VITRO EMBRYO DEVELOPMENT

Interferon tau (IFN-tau) is known as maternal pregnancy recognition factor in ruminants. IFN-tau not only acts as a signalling molecule of pregnancy recognition but also performs various functions for successful implantation and pregnancy establishment. The aim of the present study was to produce recombinant buffalo interferon-tau (BuIFN-Tau) and observe if it has any effect on in vitro embryo development. The BuIFN-Tau gene was obtained through polymerase chain reaction (PCR) from hatched buffalo blastocysts and was cloned into pJET cloning vector. Screening of the recombinant colonies gave 8 distinct buffalo IFN-tau isoforms, out of which the predominant buffalo IFN-t tau1 isoform (gene bank accession number JX481984), was subcloned into expression vector pET22b without signal sequence. The recombinant plasmid was induced to express the recombinant protein by isopropyl b-D-1-thiogalactopyranoside. Analysis of the products of recombinant BuIFN-tau without signal sequence by SDS–PAGE revealed a new 20-kDa protein coinciding with the molecular weight of IFN-tau as reported earlier in literature. The purified recombinant BuIFN-tau was confirmed by Western blot using anti-HIS antibody and was subjected to three steps of large-scale purification using HIS affinity chromatography, anion exchange chromatography, and gel filtration chromatography. Finally, a relatively pure histidine-tagged recombinant protein, which had a purity of at least 90%, was generated as confirmed through SDS. The concentration of recombinant BuIFN-tau was 1 mg mL–1 by Bradford assay. The purified recombinant BuIFN-tau was used as supplement of the culture medium for IVF early buffalo embryos at the following concentrations: control, 1, 2, and 4 µg mL–1. Sixty oocytes each in 4 groups (with 20 oocytes/drop in three replicates for each group) were used for in vitro maturation. After 24 h, the matured oocytes were incubated with in vitro capacitated sperm cells for 18 h; thereafter, the presumptive zygotes were cultured in IVC medium supplemented with 0, 1, 2, or 4 µg mL–1 of the purified recombinant BuIFN-tau. The experiment was repeated 3 times. The data were analysed using SYSTAT 7.0 (SPSS Inc., Chicago, IL, USA) after arcsin transformation of percentage values. The differences were analysed by one-way ANOVA followed by Fisher's least significant difference test. Out of 3 concentrations of recombinant BuIFN-tau, the 2 µg mL–1 concentration significantly promoted the rate of blastocyst development, 45.55% against 31.1% (control; P &lt; 0.01). Blastocyst development rate for low and high concentrations was 29.97% and 10.18% respectively. It is concluded that the addition of 2 µg mL–1 of recombinant BuIFN-tau enhances the blastocyst development rate in buffalo, and hence there is some evidence that BuIFN-tau has not only a role in maternal recognition of pregnancy but also in embryonic development.

151 A NOVEL METHOD TO INCREASE THE DEVELOPMENTAL POTENTIAL OF ACTIVATED OOCYTES BY USING THE Zn2+ CHELATOR TPEN [N,N,N',N'-TETRAKIS(2-PYRIDYLMETHYL)ETHYLENEDIAMINE

An increase in intracellular Ca2+ concentration is essential for oocyte activation. Thus most artificial oocyte-activation methods focus on increasing the Ca2+ concentration in the oocytes. Recently, full-term development was reported in mice when oocytes were activated with no increase in intracellular Ca2+ using the Zn2+ chelator N,N,N′,N′-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN). During oocyte maturation, Zn2+ is responsible for Cdc25c recognition. Once attached, the Cdc25c-zinc complex dephosphorylates maturation-promoting factor (MPF)/cyclin-dependent kinase 1 (cdk1), thus activating the Cdc25c-MPF positive feedback loop and keeping the oocyte suspended in metaphase II. The TPEN can inhibit the Cdc25c-MPF positive feedback loop, indirectly giving TPEN the power to degrade MPF, allowing the oocyte to exit metaphase II. First, we tested if incubation of porcine oocytes with TPEN could induce oocyte activation. Second, we examined whether the combination of TPEN with conventional activation methods could increase the developmental potential of activated oocytes. Last, based on the results, somatic cell nuclear transfer (SCNT) embryos were further activated with the optimum condition of TPEN to produce clones to confirm developmental competence. Frequencies of blastocyst formation were recorded and analysed by using ANOVA following arcsin transformation. Total cell numbers in blastocysts were counted and compared by using the Student's t-test. Differences at P &lt; 0.05 were considered significant. When oocytes were incubated with a high concentration of TPEN (100–250 μM) for 10 to 120 min, blastocyst formation was comparable with conventional activation methods; however, the total cell number in the blastocysts was significantly lower (31.3 ± 3.1 v. 24.8 ± 1.9). When oocytes were activated with conventional methods and then incubated with the high concentration of TPEN, embryo development was drastically decreased; no blastocyst development was achieved from TPEN-treated oocytes. Interestingly, when activated oocytes were incubated with a low concentration of TPEN (5–10 μM), surprisingly, the TPEN-treated group showed higher developmental potential compared with the control group. Specifically, the average percent blastocyst formation of TPEN-treated oocytes (5 μM for 30 min) was 27.2 ± 1.7%, but only 10.6 ± 2.5% developed to blastocyst in the control group. Moreover, the average cell number in blastocysts was significantly higher in TPEN-treated oocytes compared with the control group (33.1 ± 2.6 v. 28.2 ± 2.1, respectively). When 290 chemically activated SCNT embryos were treated with 5 μM TPEN for 30 min and transferred into a surrogate, 2 healthy piglets were born. The results indicate that incubation of oocytes with TPEN alone can activate porcine oocytes. Also, when activated oocytes are incubated with the right concentration of TPEN, it can increase embryo quality in vitro. Embryo transfer results also show that TPEN-incubated SCNT embryos are developmentally competent. Additional studies would guide us to develop more efficient way to use TPEN in the activation of SCNT embryos.

103 ESHERICHIA COLI LIPOPOLYSACCHARIDE STIMULATES PROLIFERATION OF BOVINE UTERINE EPITHELIAL CELLS

Lipopolysaccharide (LPS) is a component of the outer membrane of gram-negative bacteria involved in the pathogenic processes leading to postpartum metritis and endometritis in dairy cows. It also causes inflammation of the endometrium and implantation failures in many animal species. An increase in cell proliferation by LPS has been reported in human epithelial and immune cells (Martin et al. 2000 J. Immunol. 165, 139–147) but not in endometrial cells. The aim of this study was to characterise the proliferative response of bovine endometrial cells following exposure to Escherichia coli LPS. In vitro cultures of bovine endometrial epithelial cells (EEC) and fibroblasts were performed following collection of bovine endometrium at the slaughterhouse. Endometrial epithelial cells and fibroblasts were separated before primary culture (Charpigny et al. 1999 J. Reprod. Fertil. 117, 315–324). On passages 4 to 6, EEC issued from 5 cows (total 25 replicates) were challenged with 2, 4, 8, 12, 16, or 24 μg mL–1 of LPS. At the time of challenge and 72 h later, the numbers of attached cells were counted for control and LPS-treated groups. The variation in cell numbers after the challenge was calculated as the ratio of the number of LPS-treated cells minus the number of untreated control cells to the number of untreated control cells. The variation in cell numbers over time was analysed by ANOVA (SAS 9.1, PROC GLM; SAS Institute Inc., Cary, NC, USA) following arcsin transformation of percentages. The effect of passage number, initial number of cells at the time of challenge, treatment group, and corresponding interactions were included in the model. A significant increase in cell numbers was observed for cells treated with 2, 4, 8, and 12 μg mL–1 of LPS (+21 ± 5%, +30 ± 5%, +41 ± 4%, and +14 ± 5% over the control, respectively; P ≤ 0.001), whereas a nonsignificant effect was observed for 16 and 24 μg mL–1 of LPS (+4 ± 5%, and –5 ± 14%, respectively). Effects of passage number and initial number of cells at the time of challenge were nonsignificant and no interactions of those factors with treatment effects were observed. For fibroblasts, preliminary results (2 replicates from 2 cows) suggest that the response to LPS is much less important than for EEC (+3.6 and +1.3% for 2 μg mL–1 of LPS; +5.8 and +4% for 8 μg mL–1 of LPS). These findings indicate that E. coli LPS stimulates proliferation of bovine endometrial epithelial cells, and that the effect of LPS is dose dependent but not linear under the range of concentrations tested.

Effect of biostimulation and social organization on the interval from calving to resumption of ovarian cyclicity in postpartum Angus cows

The objective was to assess the effect of biostimulation by the male presence and social organization on the interval from calving to resumption of ovarian cyclicity (ICR). Thirty Angus cows were allocated according parity into three groups (10 per group); two groups were exposed to bulls, and a third group not exposed to bulls served as a control. Dominance values (with subsequent arc-sin transformation) were calculated from daily recorded agonistic interactions and later organized into dominance order comprising three social categories as follows: dominant (D), intermediate (I), and subordinates (S). The ICR was established by determining presence of luteal tissue and a rise of blood progesterone concentration above 1 ng/mL using ultrasonography and a solid-phase, nonextraction radioimmunoassay (Coat-a-Count; Diagnostics Products Corporation, Los Angeles, CA, USA), respectively. The effect of biostimulation, dominance order, and treatment by dominance order on ICR was statistically analyzed applying ANOVA using PROC GLM of SAS (2010). The ICR was influenced by biostimulation (P < 0.002) and dominance order (P < 0.004). The ICR increased as dominance order decreased (D = 34.5 ± 6 days; I = 45.0 ± 6; S = 53.1 ± 4 days; P < 0.01). However, when comparing cows within social categories, ICR was reduced in the group exposed to bulls (D = 26.3 ± 8.2 days; I = 42.0 ± 6.4 days; S = 46.1 ± 4.1 days) compared with those not exposed to bulls (D = 43.0 ± 8.2 days; I = 48.0 ± 10.1 days; S = 60.2 ± 6.4 days) cows. In conclusion, biostimulation and social dominance influenced the ICR.

28 EFFECT OF TRICHOSTATIN A ON IN VITRO EMBRYO DEVELOPMENT OF INTERSPECIES NUCLEAR TRANSFER EMBRYOS RECONSTRUCTED FROM CAT DONOR NUCLEI AND BOVINE CYTOPLASM

Interspecies somatic cell nuclear transfer (iSCNT) is an invaluable tool for studying nucleus-cytoplasm interactions and may provide an alternative for cloning endangered animals whose oocytes are difficult to obtain. The developmental ability of iSCNT embryos decreases with increases in taxonomic distance between the donor and recipient species. The development of cat-bovine iSCNT embryos is reportedly blocked at the 8-cell stage (Thongphakdee et al. 2008 J. Reprod. Dev. 54, 142–147). Abnormal epigenetic reprogramming, such as DNA methylation or histone modifications, may cause low iSCNT efficiencies. The present study was conducted to evaluate the effect of the histone deacetylase inhibitor trichostatin A (TSA), previously used to enhance nuclear reprogramming following SCNT, on the developmental ability of cat iSCNT embryos using bovine oocytes matured in vitro. The matured bovine oocyte was enucleated by the glass needle and the domestic cat fetal fibroblast used as the donor nuclei was then placed into the perivitelline space adjacent to the plasma membrane of the oocyte. Couplets with bovine ooplasm were fused and activated simultaneously with a single DC pulse of 2.3 kV cm–1 for 30 µs, respectively, using an electro cell fusion generator followed by cycloheximide treatment. Reconstructed cat-bovine embryos were treated with 0, 25, 50, and 100 nM concentrations of TSA for 24 h following fusion. The percentages of embryos cleaved and embryos developed to the blastocyst stage were subjected to arc sin transformation before ANOVA. The TSA treatment at 50 nM contributed significantly higher rates of cleavage and blastocyst formation (n = 139; 84.3 and 4.6%, respectively) compared with untreated embryos (n = 187; 63.8 and 0%, respectively) and embryos treated with 100 nM TSA (n = 172; 71.4 and 0%, respectively; P &lt; 0.05). Development to the morula stage of iSCNT embryos was observed in the TSA treatment groups, whereas no embryos developed beyond the 16-cell stage in the untreated group. In conclusion, our results indicate that TSA treatment for 24 h following fusion improves the development of iSCNT embryos. Specifically, 50 nM TSA treatment provides a beneficial effect on cleavage and development to the blastocyst stage of cat iSCNT embryos using bovine oocytes matured in vitro as recipients and domestic cat fibroblasts as donor nuclei.

96 GRANULOCYTE–MACROPHAGE COLONY-STIMULATING FACTOR INCREASES DEVELOPMENTAL POTENTIAL OF PORCINE EMBRYOS IN VITRO

There have been significant improvements in the culture of porcine embryos in vitro; however, it is still suboptimal. Improvements in porcine embryo culture would benefit utilisation of porcine embryos for a variety of purposes. Granulocyte-macrophage colony-stimulating factor (GM-CSF) is known to be expressed in the female reproductive tract and the level of its expression is high between conception and implantation. Previous studies show supplementing GM-CSF in embryo culture promotes embryonic development in human and bovine embryos. The aim of this study was to investigate the effect of GM-CSF on the culture of porcine embryos derived from somatic cell nuclear transfer (SCNT) and IVF. Different concentrations of recombinant porcine GM-CSF (0, 2, 10 ng mL–1) were introduced into Porcine Zygote Medium 3 from Day 1 to 6. Frequencies of cleaved embryos and blastocyst formation were recorded and analysed by using ANOVA following arcsin transformation. Total cell number in blastocysts from each group were counted and compared by using the Student's t-test. Differences at P &lt; 0.05 were considered significant. A total of 563 SCNT embryos from 6 different donor cell lines on 11 different days were produced for the study. Incubation of SCNT embryos with GM-CSF did not affect the frequency of cleaved embryos. Frequencies of cleaved embryos in control (0 ng mL–1), 2 ng mL–1 GM-CSF and 10 ng mL–1 GM-CSF were 64.2%, 68.1% and 65.0%, respectively. Interestingly, both concentrations of GM-CSF significantly increased the frequency of blastocyst formation as compared with the control. In 2 ng mL–1 and 10 ng mL–1 of GM-CSF groups, 30.8% and 32.3% of embryos reached blastocyst respectively, whereas only 22.4% of embryos reached blastocyst in the control group. A significant increase in total cell number in blastocysts was observed when GM-CSF was introduced into embryo culture. An average of 28.8 ± 0.9 cells was recorded in the control group, whereas 31.9 ± 1.1 and 31.8 ± 1.1 were observed in 2 ng mL–1 and 10 ng mL–1 of GM-CSF groups, respectively. Similar effects were observed when GM-CSF was introduced to the culture of IVF embryos. For IVF study, 525 embryos were generated on 10 different days and embryos cultured in the presence of GM-CSF tended to show higher blastocyst formation (P = 0.1). Frequencies of blastocyst per cleaved in the 3 groups were 55.7% (control), 65.7% (2 ng mL–1 GM-CSF) and 66.7% (10 ng mL–1 GM-CSF). In addition, culture of IVF embryos with GM-CSF significantly increased total cell number in Day 6 blastocysts. Total cell number in blastocysts in 2 ng mL–1 GM-CSF (34.2 ± 0.8) and 10 ng mL–1 GM-CSF (34.4 ± 1.2) were significantly higher compared with control (27.3 ± 1.2). Our results indicate that introducing GM-CSF into embryo culture media can increase the quality of blastocyst stage embryos. An increase in the frequency of blastocyst formation and total cell number in blastocysts suggests that GM-CSF can be used to produce better-quality embryos in vitro. Currently, effects of GM-CSF on implantation of SCNT embryos are under investigation. Further studies would elucidate the specific mechanism of GM-CSF on porcine embryos.

361 EFFECT OF HORMONES, EGF AND β-MERCAPTOETHANOL ON IN VITRO MATURATION OF CAPRINE OOCYTES

In vitro maturation of oocytes is an integral part of in vitro culture system. In almost all studies of mammalian in vitro maturation, the basic medium is supplemented with serum and hormones. The maturation medium and selection of protein supplements, growth factors, antioxidants and hormones for IVM play an important role in subsequent in vitro fertilization and in vitro embryo development. The objective of the present experiment was to study the effect of exogenous addition of hormones, epidermal growth factor, insulin and β-mercaptoethanol to the maturation medium for in vitro maturation of caprine oocytes. A total of 1540 oocytes were collected from slaughtered goat of 1.5 to 2.5 year of age and randomly divided in to treatment groups. Group 1; COCs were matured in TCM-199 medium containing 10% calf serum (NCS) and 3 mg mL-1 BSA (used as a base medium) for control, Group 2; COCs were matured in a base medium supplemented with hormones (5 μg mL-1 FSH, 5 μg mL-1 LH and 1 μg mL-1 estradiol-17β), Group 3β COCs were matured in a base medium supplemented with 50 ng mL-1 insulin, Group 4β COCs were matured in a base medium supplemented with hormones and 10 ng mL-1 EGF, Group 5; COCs were matured in a base medium supplemented with hormones and 50 mM β-mercaptoethanol and Group 6; COCs were matured in a base medium supplemented with hormones and 50 ng mL-1 insulin. These COCs were matured at 38.5°C in 5% CO2 in air for 27 h. After the maturation, oocytes were separated from cumulus and corona cells by treatment with 0.1% hyaluronidase and by passing through a fine pipette. They are them fixed in 2.5% glutaraldehyde, stained with DAPI and observed under fluorescent microscope for evidence of nuclear maturation. The maturation rates in groups 1 to 6 were 33.6%, 38.0%, 39.7%, 60.0%, 37.4%, and 44%, respectively. Statistical analysis (ANOVA) after arcsin transformation revealed that the maturation rate in group 4 was statistically significant (P &lt; 0.05) as compared to those in groups 1, 2, 3, 5, and 6. The results suggest that the supplementation of EGF in maturation medium significantly enhances the in vitro maturation rate of caprine oocytes.

Analysis of the Variability of a Two-Dimensional Finite Strain Estimate

Finite strain estimation is a widely used technique for the study of rock deformation in structural geology. One particular algorithm proposed by Shimamoto and Ikeda uses the ‘average shape matrix’ of deformed markers. This paper provides a detailed error analysis for resulting strain estimates in two dimensions. When the number of markers exceeds 100, estimators of components of the strain tensor are shown to have an approximately Gaussian distribution with variances that increase with their mean. Equal variance estimators are obtained by applying a log transform for the elongation and an arcsin transformation for the orientation estimates. Confidence interval formulae for strain tensor components are proposed. Lithology specific constants arising in these formulae are estimated from undeformed samples. The results are validated by application to simulated data as well as observational data from thin sections of sandstone sampled from SE Ireland.

STATUS OF ENVIRONMENTAL PROTECTION AS A SOURCE OF FINANCE FOR REGIONAL ECONOMIC DEVELOPMENT: MEASUREMENT OF ENVIRONMENTAL AND REGIONAL POLICY WITH THE FISHER FUNCTION/ APLINKOS APSAUGOS PADĖTIS KAIP FINANSINIS IŠTEKLIUS REGIONINEI EKONOMINEI PLĖTRAI: MATAVIMAI, SUSIJĘ SU APLINKOS IR REGIONINE POLITIKA NAUDOJANT FISCHER FUNKCIJĄ/ПРИРОДООХРАННЫЙ СТАТУС КАК ФИНАНСОВЫЙ ИСТОЧНИК РЕГИОНАЛЬНОГО РАЗВИТИЯ: ОЦЕНКА ПРИРОДООХРАННОЙ И РЕГИОНАЛЬНОЙ ПОЛИТИКИ С ПРИМЕНЕНИЕМ ФУНКЦИИ ФИШЕРА

The present paper analyses spatial distribution of EU financial support for territories, where environmental protection is of high importance, and tests the measuring method based on the t‐statistic of sign frequency in order to apply the R. Fisher function's arcsin transformation. The measuring of EU fund distribution is based on the analysis of sign frequency in sample groups with sign frequency in all the municipalities of Latvia. There are 530 municipalities in Latvia, and in 100 of them environmental protection is of high importance. The author has concluded that Latvian municipalities with high environmental protection levels have significantly higher possibilities to obtain EU funds for environmental improvement. A compact group of municipalities with high environmental protection levels have higher and significantly higher possibilities to obtain EU funding for environmental improvement and local business development support. Participation in environmental protection projects did not worsen the economic development scenario. Santrauka Analizuojama Europos Sąjungos finansinės paramos teritorijoms, kuriose aplinkos apsaugai teikiama daug dėmesio, erdvinis pasiskirstymas ir tikrinamas matavimo metodas, kurio pagrindas yra reikšmių santykinio dažnio t-statistika, siekiant pritaikyti R. Fisher funkcijos arcsin transformaciją. ES lėšų pasiskirstymo matavimai remiasi reikšmių santykinio dažnio tiriamose grupėse su reikšmių santykiniu dažniu visose Latvijos savivaldybėse analize. Latvijoje iš viso yra 530 savivaldybių, o iš jų 100 savivaldybių aplinkos apsaugai teikiama daug dėmesio. Autorius padarė išvadą, kad Latvijos savivaldybės, kuriose yra aukštas aplinkos apsaugos lygis, turi reikšmingai didesnių galimybių gauti ES fondų paramą aplinkai gerinti. Šiose savivaldybėse kompaktinės grupės turi didesnių ir reikšmingesnių galimybių gauti ES lėšų paramą aplinkai gerinti ir vietiniam verslui plėtoti. Dalyvavimas aplinkos apsaugos projektuose nepablogino ekonominės plėtros scenarijaus. Резюме Проводится анализ пространственного распределения структурных фондов Евросоюза (ЕС) для территорий с высокой значимостью природоохранного статуса и проверяется метод, основанный на т-статистике относительной частоты и применении арксинусной трансформации функции Рональда Фишера. Оценка распределения фондов ЕС основывается на анализе относительной частоты в выборке по сравнению с относительной частотой в Латвии. Из 530 самоуправлений Латвии в 100 из них природоохранные участки занимают значительную часть территории. Автор делает вывод о том, что самоуправления с высокой значимостью природоохранного статуса имеют статистически значимо большие возможности в получении фондов ЕС для природоохранных целей. Компактные группы самоуправлениий с высокой значимостью природоохранного статуса, к примеру, территория Северо-Видземского биосферного заповедника, имеют статистически значимо большие возможности в получении фондов ЕС для природоохранных целей и развития местного бизнеса. Участие самоуправлений в разработке природоохранных проектов весьма перспективно для экономичеекого развития.