natgillin/translations-raw huggingface dataset:
POKYNY K INTERPRETACI A PŘIJETÍ VÝSLEDKŮ
This test method describes an in vivo screening assay where sexually mature male and spawning female fish are held together and exposed to a chemical during a limited part of their life-cycle (21 days).
Tento oddíl obsahuje některé aspekty, které je třeba vzít v úvahu při interpretaci výsledků zkoušky pro různé měřené ukazatele.
At termination of the 21-day exposure period, two biomarker endpoints are measured in males and females as indicators of endocrine activity of the test chemical; these endpoints are vitellogenin and secondary sexual characteristics.
K výsledkům zkoušky je třeba přistupovat s opatrností, pokud se zdá, že zkoušená chemická látka způsobuje zjevnou toxicitu nebo ovlivňuje celkový stav zkušebního zvířete.
Vitellogenin is measured in fathead minnow, Japanese medaka and zebrafish, whereas secondary sex characteristics are measured in fathead minnow and Japanese medaka.
Při stanovování rozmezí zkušebních koncentrací by se mělo dbát na to, aby nebyla překročena maximální tolerovaná koncentrace a byla tak umožněna smysluplná interpretace údajů.
Additionally, quantitative fecundity is monitored daily throughout the test.
Je důležité, aby existovala alespoň jedna koncentrace, při které se neprojevují žádné příznaky toxických účinků.
Gonads are also preserved and histopathology may be evaluated to assess the reproductive fitness of the test animals and to add to the weight of evidence of other endpoints.
Příznaky onemocnění a příznaky toxických účinků se pečlivě posoudí a zaznamenají.
This bioassay serves as an in vivo reproductive screening assay and its application should be seen in the context of the “OECD Conceptual Framework for the Testing and Assessment of Endocrine Disrupting Chemicals” (30).
Je například možné, že tvorba VTG u samiček může být ovlivněna také obecnou toxicitou a toxickými účinky jiného než endokrinního charakteru, např. toxickými účinky na játra.
In this Conceptual Framework the Fish Short Term Reproduction Assay is proposed at Level 3 as an in vivo assay providing data about selected endocrine mechanism(s)/pathway(s).
Interpretace účinků však může být podpořena na jiných úrovních expozice, které nejsou zkresleny systémovou toxicitou.
Vitellogenin (VTG) is normally produced by the liver of female oviparous vertebrates in response to circulating endogenous oestrogen.
Existuje několik aspektů, které je třeba zvážit pro přijetí výsledků zkoušky.
It is a precursor of egg yolk proteins and, once produced in the liver, travels in the bloodstream to the ovary, where it is taken up and modified by developing eggs.
Obecně vzato by hladiny VTG v kontrolních skupinách samečků a samiček měly být odlišné – u jelečka velkohlavého a dania pruhovaného by se měly lišit přibližně o tři řády a u halančíka japonského přibližně o jeden řád.
Vitellogenin is almost undetectable in the plasma of immature female and male fish because they lack sufficient circulating oestrogen; however, the liver is capable of synthesising and secreting vitellogenin in response to exogenous oestrogen stimulation.
Příklady rozmezí hodnot zjištěných v kontrolních a exponovaných skupinách jsou dostupné ve validačních zprávách (1) (2) (3) (4).
The measurement of vitellogenin serves for the detection of chemicals with various oestrogenic modes of action.
Vysoké hodnoty VTG u samečků v kontrolních skupinách by mohly zpochybnit citlivost zkoušky a její schopnost odhalit slabé agonisty estrogenu.
The detection of oestrogenic chemicals is possible via the measurement of vitellogenin induction in male fish, and it has been abundantly documented in the scientific peer-reviewed literature (e.g. (7)).
Nízké hodnoty VTG u samiček v kontrolních skupinách by mohly zpochybnit citlivost zkoušky a její schopnost detekovat inhibitory aromatázy a antagonisty estrogenu.
Vitellogenin induction has also been demonstrated following exposure to aromatisable androgens (8, 9).
Při vypracování tohoto návodu byly použity validační studie.
A reduction in the circulating level of oestrogen in females, for instance through the inhibition of the aromatase converting the endogenous androgen to the natural oestrogen 17β-estradiol, causes a decrease in the VTG level which is used to detect chemicals having aromatase inhibiting properties (10, 11).
Pokud jde o kvantifikaci tvorby jiker, zde se projevují důležité rozdíly [variační koeficient (CV) se může pohybovat od 20 do 60 %], které mohou snížit schopnost zkoušky odhalit výrazný pokles tvorby jiker nižší než 70 %, jestliže se CV blíží 50 % nebo dosahuje ještě vyšší hodnoty.
The biological relevance of the vitellogenin response following oestrogenic/aromatase inhibition is established and has been broadly documented.
Omezí-li se CV na nižší hodnoty (kolem 20 až 30 %), bude mít zkouška přijatelnou průkaznost (80 %) pro detekci 40 až 50 % snížení tvorby jiker.
However, it is possible that production of VTG in females can also be affected by general toxicity and non-endocrine toxic modes of action, e.g. hepatotoxicity.
U jelečka velkohlavého by uspořádání zkoušky zahrnující čtyři opakování pro každou úroveň expozice mělo umožnit větší průkaznost ukazatele plodnosti v porovnání se zkušebním uspořádáním pouze se dvěma opakováními.
Several measurement methods have been successfully developed and standardised for routine use.
V praxi se doporučuje, aby každá laboratoř shromáždila své vlastní historické kontrolní údaje pro samečky a samičky v kontrolních skupinách a pro pozitivní kontrolu estrogenního působení chemické látky (např. 17-beta-estradiolu při koncentraci 100 ng/l nebo dobře známého slabého agonisty), jež vede ke zvýšené hodnotě VTG u samečků ryb, pozitivní kontrolu inhibice aromatázy chemické látky (např. fadrozolu nebo prochlorazu při koncentraci 300 μg/l), jež vede ke zvýšené hodnotě VTG u samiček ryb, a pozitivní kontrolu androgenního působení chemické látky (např. 17-beta-trenbolonu při koncentraci 5 μg/l), jež vede k indukci druhotných pohlavních znaků u samiček jelečka velkohlavého a halančíka japonského.
In medaka, there is a good correlation between VTG measured from blood and from liver (19).
Pro zajištění způsobilosti laboratoře lze všechny tyto údaje porovnat s dostupnými údaji z validačních studií (1) (2) (3).
Appendix 6 provides the recommended procedures for sample collection for VTG analysis.
Obecně lze říci, že měření VTG se považuje za pozitivní, dojde-li v porovnání s kontrolní skupinou ke statisticky významnému nárůstu VTG u samečků (p < 0,05) nebo ke statisticky významnému poklesu u samiček (p < 0,05) alespoň při nejvyšší koncentraci zkoušené látky a za nepřítomnosti příznaků obecné toxicity.
Kits for the measurement of VTG are widely available; such kits should be based on a validated species-specific ELISA method.
Pozitivní výsledek se dále podpoří prokázáním biologicky možné závislosti mezi dávkou a odezvou.
Secondary sex characteristics in male fish of certain species are externally visible, quantifiable and responsive to circulating levels of endogenous androgens; this is the case for the fathead minnow and the medaka — but not for zebrafish which does not possess quantifiable secondary sex characteristics.
Jak bylo uvedeno výše, pokles hodnoty VTG nemusí být zcela endokrinního původu; avšak pozitivní výsledek by měl být obecně interpretován jako důkaz endokrinního působení in vivo a měl by být obvykle podnětem ke krokům za účelem dalšího vyjasnění.
Females maintain the capacity to develop male secondary sex characteristics, when they are exposed to androgenic chemicals in water.
Regulační orgány mohou vyžadovat histopatologické hodnocení gonád s cílem stanovit reprodukční způsobilost zkušebních zvířat a umožnit posouzení výsledků zkoušky na základě závažnosti důkazů.
Several studies are available in the scientific literature to document this type of response in fathead minnow (20) and medaka (21).
Provedení histopatologie gonád nemusí být nezbytné v případech, kdy jsou buď VTG, anebo druhotné pohlavní znaky pozitivní (tj. zvýšení nebo pokles VTG, nebo indukce druhotných pohlavních znaků).
A decrease in secondary sex characteristics in males should be interpreted with caution because of low statistical power, and should be based on expert judgement and weight of evidence.
OECD (2006b).
There are limitations to the use of zebrafish in this assay, due to the absence of quantifiable secondary sex characteristics responsive to androgenic acting chemicals.
OECD (2007).
In the fathead minnow, the main indicator of exogenous androgenic exposure is the number of nuptial tubercles located on the snout of the female fish.
Owens JW (2007).
In the medaka, the number of papillary processes constitutes the main marker of exogenous exposure to androgenic chemicals in female fish.
US EPA (2007).
Appendix 5A and Appendix 5B indicate the recommended procedures to follow for the evaluation of sex characteristics in fathead minnow and in medaka, respectively.
OECD (2008).
The 21-day fish assay includes the evaluation of quantitative egg production and preservation of gonads for optional histopathology examination.
Androgenic and estrogenic effects of the synthetic androgen 17alpha-methyltestosterone on sexual development and reproductive performance in the fathead minnow (Pimephales promelas) determined using the gonadal recrudescence assay.
Some regulatory authorities may require this additional endpoint for a more complete evaluation of the reproductive fitness of the test animals, or in cases where vitellogenin and secondary sex characteristics did not respond to the chemical exposure.
Ankley G.T. et al.
Although some endpoints may be highly diagnostic (e.g. VTG induction in males and tubercle formation in females), not all endpoints (e.g. fecundity and gonad histopathology) in the assay are intended to unequivocally identify specific cellular mechanisms of action.
Application of an ELISA to quantify vitellogenin concentrations in fathead minnows (Pimephales promelas) exposed to endocrine disrupting chemicals.
Rather, the suite of endpoints, collectively, allows inferences to be made with regard to possible endocrine disturbances and thus provide guidance for further testing.
CEFIC-EMSG research report reference AQ001.
Although not endocrine specific, fecundity, due to its demonstrated sensitivity across known endocrine active chemicals (5), is an important endpoint to include because when it and other endpoints are unaffected one is more confident that a compound is not likely endocrine active.
CEFIC, Brussels, Belgium.
However, when fecundity is affected it will contribute heavily in weight of evidence inferences.
Fathead minnow (Pimephales promelas) vitellogenin: purification, characterization and quantitative immunoassay for the detection of estrogenic compounds.
Guidance on data interpretation and acceptance of test results is provided further in this test method.
Comparative Biochemistry and Physiology.
Definitions used in this test method are given in Appendix 1.
Panter G.H. et al.
In the assay, male and female fish in a reproductive status are exposed together in test vessels.
Fenske M. et al.
Their adult and reproductive status enables a clear differentiation of each sex, and thus a sex-related analysis of each endpoint, and ensures their sensitivity towards exogenous chemicals.
Development and validation of a homologous zebrafish (Danio rerio Hamilton- Buchanan) vitellogenin enzyme-linked immunosorbent assay (ELISA) and its application for studies on estrogenic chemicals.
At test termination, sex is confirmed by macroscopic examination of the gonads following ventral opening of the abdomen with scissors.
Comp.
An overview of the relevant bioassay conditions are provided in Appendix 2.
Biochem.
The assay is normally initiated with fish sampled from a population that is in spawning condition; senescent animals should not be used.
Phys.
Guidance on the age of fish and on the reproductive status is provided in the section on Selection of fish.
C 129 (3): 217-232.
The assay is conducted using three chemical exposure concentrations as well as a water control, and a solvent control if necessary.
Holbech H. et al.
Two vessels or replicates per treatment are used for zebrafish (each vessel containing 5 males and 5 females).
Development of an ELISA for vitellogenin in whole body homogenate of zebrafish (Danio rerio).
Four vessels or replicates per treatment are used for fathead minnow (each vessel containing 2 males and 4 females).
Comparative Biochemistry and Physiology.
This is to accommodate the territorial behaviour of male fathead minnow while maintaining sufficient power of the assay.
Part C Pharmacology, toxicology and endocrinology; 130: 119-131.
Four vessels or replicates per treatment are used for medaka (each vessel containing 3 males and 3 females).
Rose J. et al.
The exposure is conducted for 21-days and sampling of fish is performed at day 21 of exposure.
Vitellogenin induction by 17b-estradiol and 17a-ethinylestradiol in male zebrafish (Danio rerio).
Quantitative fecundity is monitored daily.
C.
On sampling at day 21, all animals are killed humanely.
131: 531-539.
Secondary sex characteristics are measured in fathead minnow and medaka (see Appendix 5A and Appendix 5B); blood samples are collected for determination of VTG in zebrafish and fathead minnow, alternatively head/tail can be collected for the determination of VTG in zebrafish (Appendix 6); liver is collected for VTG analysis in medaka (Appendix 6); gonads are fixed either in whole or dissected for potential histopathological evaluation (22).
(16) Brion F. et al.
TEST ACCEPTANCE CRITERIA
Development and validation of an enzyme-linked immunosorbent assay to measure vitellogenin in the zebrafish (Danio rerio).
For the test results to be acceptable the following conditions apply:
Environmental Toxicology and Chemistry; vol 21: 1699-1708.
the mortality in the water (or solvent) controls should not exceed 10 per cent at the end of the exposure period;
Yokota H. et al.
the dissolved oxygen concentration should be at least 60 per cent of the air saturation value (ASV) throughout the exposure period;
Development of an ELISA for determination of the hepatic vitellogenin in Medaka (Oryzias latipes).
the water temperature should not differ by more than ± 1,5 °C between test vessels at any one time during the exposure period and be maintained within a range of 2 °C within the temperature ranges specified for the test species (Appendix 2);
Jpn J Environ Toxicol 4:87–98.
evidence should be available to demonstrate that the concentrations of the test chemical in solution have been satisfactorily maintained within ±20 % of the mean measured values;
Tatarazako N. et al.
evidence that fish are actively spawning in all replicates prior to initiating chemical exposure and in control replicates during the test.
Guidance Document on Fish Gonadal Histopathology.
oxygen and pH meters;
OECD Environmental Health and Safety Publications Series on Testing and Assessment No.
equipment for determination of water hardness and alkalinity;
123, Paris.
adequate apparatus for temperature control and preferably continuous monitoring;
OECD (2000) Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures.
tanks made of chemically inert material and of a suitable capacity in relation to the recommended loading and stocking density (see Appendix 2);
Series on Testing and Assessment No.
spawning substrate for fathead minnow and zebrafish, Appendix 4 gives the necessary details.
Review.
suitably accurate balance (i.e. accurate to ± 0,5 mg).
Hutchinson T.H. et al.
Any water in which the test species shows suitable long-term survival and growth may be used as test water.
(2006b).
The pH of the water should be within the range 6,5 to 8,5, but during a given test it should be within a range of ± 0,5 pH units.
Screening and testing for endocrine disruption in fish-biomarkers as „signposts,“ not „traffic lights,“ in risk assessment.
In order to ensure that the dilution water will not unduly influence the test result (for example by complexion of test chemical); samples should be taken at intervals for analysis.
Miles-Richardson S.R. et al.
Measurements of heavy metals (e.g. Cu, Pb, Zn, Hg, Cd, and Ni), major anions and cations (e.g. Ca2+, Mg2+, Na+, K+, Cl–, and SO42–), pesticides (e.g. total organophosphorus and total organochlorine pesticides), total organic carbon and suspended solids should be made, for example, every three months where dilution water is known to be relatively constant in quality.
Effects of waterborne exposure to 17b-estradiol on secondary sex characteristics and gonads of the fathead minnow (Pimephales promelas).
If water quality has been demonstrated to be constant over at least one year, determinations can be less frequent and intervals extended (e.g. every six months).
Aquat.
Some chemical characteristics of acceptable dilution water are listed in Appendix 3.
Toxicol.
The stock solution should preferably be prepared by simply mixing or agitating the test chemical in dilution water by using mechanical means (e.g. stirring or ultrasonication).
47, 129-145.
Saturation columns (solubility columns) can be used for achieving a suitable concentrated stock solution.
Martinovic D. et al.
However, in case a solvent is necessary, a solvent control should be run in parallel, at the same solvent concentration as the chemical treatments.
Characterization of reproductive toxicity of vinclozolin in the fathead minnow and co-treatment with an androgen to confirm an anti-androgenic mode of action.
For difficult to test chemicals, a solvent may be technically the best solution; the OECD guidance document on aquatic toxicity testing of difficult substances and mixtures should be consulted (23).
Environ.
The choice of solvent will be determined by the chemical properties of the substance or mixture.
Toxicol.
The OECD guidance document recommends a maximum of 100 μl/l, which should be observed.
Chem.
However a recent review (24) highlighted additional concerns when using solvents for endocrine activity testing.
27, 478-488.
Therefore it is recommended that the solvent concentration, if necessary, is minimised wherever technically feasible (dependent on the physical-chemical properties of the test chemical).
OECD (2006c), Current Approaches in the Statistical Analysis of Ecotoxicity Data: A Guidance to Application, OECD Environmental Health and Safety Publications, Series on Testing and Assessment No.54, OECD, Paris.
A flow-through test system will be used.
US EPA (2008), Peer-Review Results for the Fish Short-Term Reproduction Assay, dated 30 January 2008, US Environmental Protection Agency, Washington DC.
Such a system continually dispenses and dilutes a stock solution of the test chemical (e.g. metering pump, proportional diluter, saturator system) in order to deliver a series of concentrations to the test chambers.
OECD (2012), OECD Conceptual Framework for Testing and Assessment of Endocrine Disrupters, OECD Environmental Health and Safety Publications, Series on Testing and Assessment No.
The flow rates of stock solutions and dilution water should be checked at intervals, preferably daily, during the test and should not vary by more than 10 % throughout the test.
150, OECD, Paris.
Care should be taken to avoid the use of low-grade plastic tubing or other materials that may contain biologically active chemicals.
ZKRATKY A DEFINICE
When selecting the material for the flow-through system, possible adsorption of the test chemical to this material should be considered.
látka nebo směs.
Test fish should be selected from a laboratory population, preferably from a single stock, which has been acclimated for at least two weeks prior to the test under conditions of water quality and illumination similar to those used in the test.
variační koeficient.
It is important that the loading rate and stocking density (for definitions, see Appendix 1) be appropriate for the test species used (see Appendix 2).
enzymová imunosorpční analýza.
mortalities of greater than 10 % of population in seven days: reject the entire batch;
Osa HPG
mortalities of between 5 % and 10 % of population: acclimation for seven additional days; if more than 5 % mortality during second seven days, reject the entire batch;
hypothalamo-hypofýzo-gonadální osa.
mortalities of less than 5 % of population in seven days: accept the batch.
Velikost násady
Fish should not receive treatment for disease during the acclimation period, in the pre-exposure period, or during the exposure period.
živá hmotnost ryb na jednotku objemu vody.
Pre-exposure and selection of fish
maximální tolerovaná koncentrace, představuje přibližně 10 % mediánu letální koncentrace (LC50).
The one to two-week pre-exposure period is recommended with animals placed in vessels similar to the actual test.
Hustota obsádky
Fish should be fed ad libitum throughout the holding period and during the exposure phase.
počet ryb na jednotku objemu vody.
The exposure phase is started with sexually dimorphic adult fish from a laboratory supply of reproductively mature animals (e.g. with clear secondary sexual characteristics visible as far as fathead minnow and medaka are concerned), and actively spawning.
vitelogenin je fosfolipoglykoprotein, prekurzor proteinů vaječného žloutku, který se obvykle vyskytuje u sexuálně aktivních samiček všech vejcorodých druhů.
For general guidance only (and not to be considered in isolation from observing the actual reproductive status of a given batch of fish), fathead minnows should be approximately 20 (± 2) weeks of age, assuming they have been cultured at 25 ± 2 °C throughout their lifespan.
EXPERIMENTÁLNÍ PODMÍNKY PRO SCREENINGOVOU ZKOUŠKU ENDOKRINNÍHO PŮSOBENÍ NA RYBÁCH
Japanese medaka should be approximately 16 (± 2) weeks of age, assuming they have been cultured at 25 ± 2 °C throughout their lifespan.
Typ zkoušky
Zebrafish should be approximately 16 (± 2) weeks of age, assuming they have been cultured at 26 ± 2 °C throughout their lifespan.
průtoková
Egg production should be assessed daily during the pre-exposure phase.
Teplota vody
It is recommended that spawning be observed in all replicate tanks prior to inclusion in the exposure phase of the assay.
Kvalita osvětlení
Quantitative guidance on desirable daily egg production cannot be provided at this stage, but it is relatively common to observe average spawns of > 10 eggs/female/day for each species.
Fluorescenční zářivky (širokospektrální)
A randomised block design according to egg production output should be used to allocate replicates to the various experimental levels to ensure balanced distribution of replicates.
Intenzita osvětlení
TEST DESIGN
10–20 μE/m2/s, 540–1000 luxů nebo 50–100 ft-c (okolní laboratorní hodnoty)
Three concentrations of the test chemical, one control (water) and, if needed, one solvent control are used.
12–16 h světlo, 12–8 h tma
The data may be analysed in order to determine statistically significant differences between treatment and control responses.
< 5 g na l
These analyses will inform whether further longer term testing for adverse effects (namely, survival, development, growth and reproduction) is required for the chemical, rather than for use in risk assessment (25).
Velikost zkušební komory
For zebrafish, on day 21 of the experiment, males and females from each treatment level (5 males and 5 females in each of the two replicates) and from the control(s) are sampled for the measurement of vitellogenin.
10 l (minimálně)
For medaka, on day 21 of the experiment, males and females from each treatment level (3 males and 3 females in each of the four replicates) and from the control(s) are sampled for the measurement of vitellogenin and secondary sex characteristics.
2 l (minimálně)
For fathead minnow, on day 21 of exposure, males and females (2 males and 4 females in each of the four replicates) and from the control(s) are sampled for the measurement of vitellogenin and secondary sex characteristics.
5 l (minimálně)
Quantitative assessment of fecundity is required, and gonadal tissues should be fixed in whole or dissected for potential histopathological evaluation, if required.
Objem zkušebního roztoku
Selection of test concentrations