Gonadal intersex in teleosts: Mechanisms, molecular biomarkers and diagnostic assays

Natural and synthetic estrogenic and androgenic compounds are continuously released into aquatic ecosystems. Exposure of teleost fishes to these contaminants can negatively impact sex differentiation and reproductive output. Specifically, development of gonadal intersex in gonochoristic (fixed sex) fish species has been studied extensively in relation to exposure to this class of compounds. The main objectives of this dissertation were to: 1) conduct field and laboratory studies to investigate the molecular signaling pathways behind the development of gonadal intersex; and 2) establish molecular biomarkers and assays for testing the ability of environmental pollutants to develop this condition using a battery of molecular, cellular and organ-level tools. First, we conducted a literature review to summarize all available articles reporting gonadal intersex in wild populations of gonochoristic fish (Chapter 1). We also included the limited information available on this topic in amphibians. We analyzed studies from across the globe, identified families and species with reported cases of gonadal intersex, and highlighted the contaminants often linked with this condition. In addition, we discussed the current knowledge of molecular signaling pathways behind the development of gonadal intersex and summarized molecular biomarkers tested and others that require further investigation. We then conducted a field study investigating the prevalence of testicular oocytes (TOs), the most prevalent form of gonadal intersex, in a sentinel freshwater fish species, the smallmouth bass Micropterus dolomieu, inhabiting the St. Joseph River and its tributaries in northern Indiana (Chapter 2). This constitutes the first study of this nature. Sites on this river were previously identified as having medium to high intersex induction potential based on contaminant quantification and estrogen equivalence estimations. We reported prevalence and severity of gonadal intersex reaching 100% in some sites, and significant decreases in prevalence and increases in severity of TOs occurrence after the spawning season. We evaluated changes in the transcription levels of several genes involved in sex differentiation and gonadal development. Significantly higher levels of vitellogenin ( vtg) transcripts were found in livers of males with TOs, but only when sampled in the spawning season. Further, we quantified contaminant levels in surface water to identify possible correlations between contamination levels and the observed prevalence of gonadal intersex. Multiple sites had detectable levels of endocrine disruptors, but no correlations with the prevalence or severity of TOs was recorded. In order to develop molecular biomarkers and assays that test contaminants’ ability to develop gonadal intersex, short-term laboratory exposures were performed using Japanese medaka Oryzias latipes, a fish model with a well-understood sex determination mechanism and high sensitivity to exogenous hormone exposure (Chapter 3). First, we identified a gene, ovary structure protein 1 (osp1), with strong female-specific expression during gonadal differentiation and observed a significant downregulation in its expression in females following short-term (10 d) exposure to a potent synthetic androgen, 17β-trenbolone. Importantly, this decrease in osp1expression was correlated with changes in ovarian phenotype, namely ovarian intersex, later in life. We decided to further utilize this promising molecular biomarker by incorporating it in a visual in vivo reporter assay for rapid detection of contaminants with estrogenic/androgenic potential (Chapter 4). For this purpose, we built a pOSP1-AcGFP (promoterOSP1-Aequorea coerulescens green florescence protein) Japanese medaka transgenic line with osp1 promoter region driving the expression of a reporter protein, AcGFP. After establishing this line, we tested its use in an in vivo visual reporter system for identifying estrogenic contaminants. Significant upregulation in fluorescence intensity was recorded in 30 d post hatch females following a 24 h exposure to 500 ng/L of a synthetic potent estrogen, ethinyl estradiol. Overall, our results support earlier findings suggesting that gonadal intersex is highly prevalent in impacted rivers across the US and that smallmouth bass are highly sensitive to developing this condition. We also conclude that hepatic and plasma VTG are promising biomarkers for diagnosing gonadal intersex, but only in males sampled during the spawning season. Our findings also support the hypothesis that molecular biomarkers, such as osp1 , are sensitive tools that can be used for early detection of the effects of contaminants with estrogenic and androgenic activity on fish and are ideal endpoints in wide-scale contaminant screening assays

Effect of nitrogen on seed yield and content and quality of oil of winter grown sunflower (Helianthus annuus L.)

ABSTRACT     Field trials were carried out at Wad Medani and Sennar locations during season 2008 / 09 to investigate the effects of nitrogen levels on growth, seed yield, oil content and quality of two sunflower cultivars. The treatments consisted of three nitrogen levels (86.7, 108 and 129 kg N/ha, applied as urea) and two cultivars (Damazin and Hysun33). Nitrogen at the higher rate (129 kg N/ha) gave higher seed yield. The oil content ranged from 36.3 to 42.9 %. Nitrogen, cultivar and location had no significant effects on oil content. Oleic and linoleic acids were the most dominant fatty acids in the oil. It is recommended to apply 129 kg N/ha to winter grown sunflower to obtain high seed yield

Studies on some physico-chemical factors of Kaptai lake

The present investigation dealt with the climatic and some physico-chemical conditions of the Kaptai lake with respect to their monthly variation. Air temperature was found always higher than water temperature. Vertical variation in temperature (0.8-4.7°C) was observed in all months. The water level fluctuates appreciably throughout the year. Wide seasonal fluctuations were also noted in water transparency. The lake was found to be slightly hard and alkaline pH. Dissolved oxygen (DO) (6.4-9.1 mg/l) and free carbondioxide (4.7- 6.0 mg/l) contents showed favourable condition for aquatic lives. DO at different depth has shown no wide variation (1.0-2.4 mg/1). Conductivity ranged between 91.9±7.1 and 106.4 ±5.2 mS/cm

A mixed culture trail of mud crab (Scylla serrata Forskal) with tilapia ( Oreocromis niloticus L.)

A ten-month study on mixed culture of mud crab Scylla serrata with tilapia (Oreochromis niloticus) was performed in brackish water earthen ponds using live tilapia fry as the only feed for crab. The monthly growth rate varied from 9.07-19.llg among four treatments. Treatment T 1, cw: 0.68cm (±0.72) and bw: 19.11 gm (±12.97) showed highest performance which was followed by T 3, cw: 0.62cm (±0.60) and bw: 13.42 gm (±10.51), T4, cw: 0.65cm (±0.64) and bw: 13.20 gm (±9.89) and T2, cw: 0.36cm (± 0.25) and bw: 9.07 gm (±8.05). Highest survivability of crabs was also recorded in T 1 (21.5%) which was followed by T 2 (15.65%), T4 (14.95%) and T 3 (14.15%). In terms of survivability, significant differences (p<0.05) were observed among the treatments whereas these were recorded as insignificant difference (p<0.05) in final weight, weight gain and production of crabs and tilapia. Mixed culture of mud crab with tilapia could make more rewarding than crab monoculture but the study suggests that only tilapia fry can not fulfill the feed requirement of crabs in respects of survivability, final body weight and weight gain. Besides, existed salinity level of 4-12 ppt during experimental period might be the another key factor for low survivability and weight gain

Androgen-responsive non-coding small RNAs extend the potential of HCG stimulation to act as a bioassay of androgen sufficiency

Background: It is unclear whether a short-term change in circulating androgens is associated with changes in the transcriptome of the peripheral blood mononuclear cells (PBMC). Aims &amp; Methods: To explore the effect of hCG-stimulation on the PBMC-transcriptome, 12 boys with a median age (range) of 0.7yrs (0.3, 11.2) who received intramuscular hCG 1500u on 3 consecutive days as part of their investigations underwent transcriptomic array analysis on RNA extracted from peripheral blood mononuclear cells before and after hCG stimulation. Results: Median pre and post hCG testosterone for the overall group was 0.7nmol/l (&lt;0.5,6) and 7.9nmol/l (&lt;0.5, 31.5), respectively. Of the 12 boys, 3 (25%) did not respond to hCG stimulation with a pre and post median serum testosterone of &lt;0.5nmol/l and &lt;0.5nmol/l, respectively. When corrected for gene expression changes in the non-responders to exclude hCG effects, all 9 of the hCG responders consistently demonstrated a 20% or greater increase in the expression of piR-37153 and piR-39248, non-coding PIWI-interacting RNAs (piRNAs). In addition, of the 9 responders, 8, 6 and 4 demonstrated a 30%, 40% and 50% rise, respectively in a total of 2 further piRNAs. In addition, 3 of the responders showed a 50% or greater rise in the expression of another small RNA, SNORD5. On comparing fold change in serum testosterone with fold change in the above transcripts, a positive correlation was detected for SNORD5 (p=0.01). Conclusions: The identification of a dynamic and androgen-responsive PBMC-transcriptome extends the potential value of the hCG test for assessment of androgen sufficiency

Comparison of two hormone preparations on the reproductive performance of air breathing catfish Clarias batrachus (Lin.)

Two hormone preparations viz. Human Chorionic Gonadotropin (HCG) and pituitary gland (PG) suspension were compared for their comparative efficacy on the breeding performance of a air breathing catfish Clarias batrachus. It was found that HCG induced fish gave better ovulation response than PG. Both fertilization and hatching of eggs were significantly (p<O.Ol) higher in HCG treated fish than PG. On all consideration, HCG was found more suitable for induced breeding of C batrachus over PG

Post impoundment changes in the fish fauna of Kaptai reservoir, Bangladesh

Based on the present investigation and reviewing the published and unpublished documents critically, this communication considers the post impoundment changes in the fish fauna of Kaptai reservoir. Investigation reveals that a total of 73 species of fish belongs to 47 genera, 25 families and 2 species of prawn are present in the reservoir. Of them, 31 are commercially important, 6 and 9 species are newly identified

A study on damage caused to crustacean and finfish larvae during collection of Penaeus monodon (Fab.) postlarvae in the estuaries of Barguna, Bangladesh

A year round investigation in the estuaries of Barguna district revealed that for each Penaeus monodon postlarvae (PL), about 37 larvae of other shrimp species, 12 finfishes and 10 macrozooplankters are destroyed during the process of shrimp seed collection. Although abundance of P. monodon PL was not recorded throughout the year, a significant number of other shrimp spp., fin fishes including macrozooplankters are being damaged by the shrimp seed collectors. This indiscriminate destruction of aquatic organisms during P. monodon PL collection is serious threat to aquatic biodiversity

HIGHLIGHTS ON THE BENEFICIATION TRIALS OF THE EGYPTIAN PHOSPHATE ORES

introduction : Phosphate rocks are vital non-renewable resources and are essential components in agricultural fertilizers and phosphorous-based chemicals [1-4]. About 95% of the phosphate produced around the world is consumed in fertilizer industry [5, 6]. Phosphate deposits can be divided into three groups (і) sea sediments, (іі) igneous and (ііі) biogenetic deposits [7]. The distribution of phosphate resources in the world is approximated as follows: 75% from sedimentary marine deposits, 15-20% from igneous, metamorphic and weathered deposits, and 2-3% from biogenic sources [5-8]. A large proportion of the world's phosphate reserves are sedimentary deposits containing a considerable amount of carbonate minerals. In terms of quality and P2O5 grade, phosphate can be divided into three groups: low - grade ores (12-16% P2O5, intermediate-grade ores (17-25% P2O5) and high-grade ores (26-35% P2O5). Deposits containing of 28-38% P2O5 are considered commercial grade deposits and have high economic value to mine and process [9].