The document summarizes key information about the zebrafish (Danio rerio) including its taxonomy, appearance, distribution, diet, breeding, development, husbandry, history of use as a model organism, and applications in various areas of research such as developmental biology, physiology, toxicology, genetics, and drug discovery. It also discusses zebrafish research centers globally and potential uses of zebrafish in evaluating natural products and toxicity of drugs from Unani medicine.

1. By:
AARIFA SABREEN SM
PG - I Year,
Dept. Of Ilmul Advia,
NIUM, Bengaluru
Under the guidance of:
Prof. Nasreen Jahan
Dept. Of Ilmul Advia
NIUM, Bengaluru.

2.  Introduction
 Taxonomy
 Appearance & Morphology
 Distribution
 Diet & Feeding
 Breeding
 Growth & Development
 Zebrafish Husbandry
 History
 Mapping of ZF research
-A global outlook
 Zebrafish as a ‘model’
organism
 Similarity
 Drug Discovery
 Toxicological studies
 Scope in Unani Medicine
 Conclusion
Tiny fish, big splash.

3. The Zebrafish is a small sized teleost fish inhabiting fresh water of
tropical regions.
The Zebrafish formerly called as Brachydanio rerio was changed as
Danio rerio in 1981.
The name Danio is derived from the Bengali word “dhani” which means
“of the rice fields”.
The Zebrafish was first described by Francis Hamilton, a surgeon with
the British East India Company, stationed principally in the West Bengal
at the beginning of 19th century.
He published “An Account of the Fishes found in the River Ganges &
its Branches” in 1822, which included 10 Danio species.

4. Kingdom Animalia
Phylum Chordata
Class Actinopterygii
Order Cypriniformes
Family Cyprinidae
Genus Danio
Species rerio

5.  The Zebrafish takes its name from the five uniform, pigmented horizontal
blue stripes on the side of the body, all of which extend to the end of the
caudal fin.
 Length usually less than 5cm.
 Body shape – fusiform, laterally compressed, with a terminal oblique mouth
directed upwards.
 The most reliable way to distinguish females from males is by the presence
of a small genital papilla in front of the anal fin origin (but this can only be
definitively determined after death).
 When alive, though similar in size and coloration, the sexes can be fairly
reliably distinguished by appearance. Reproductively mature females have a
fuller abdomen due to the developing eggs in the ovaries. Males are
generally more slender and darker in colour than females, and have more
yellow coloration in the anal fin.

7.  The zebrafish is originally found in Eastern India’s
Ganges River and native to the streams of the south-
eastern Himalayan regions.
 Also native to Pakistan, Bangladesh, Nepal and
Myanmar.
 It commonly inhabits streams, canals, ditches and ponds,
and slow-moving to stagnant water bodies including rice
fields.
 It has been introduced in Japan, Canada, US, Australia.
 It is common as an aquarium fish throughout the world.
 Zebrafish is a robust animal able to tolerate variations in
the water quality, often adapts to environmental changes
of temperature and pH.

8. Natural habitat of zebrafish

9.  The zebrafish is omnivorous.
 Its natural diet consists primarily of zooplankton and insects,
although phytoplankton, filamentous algae and vascular plant
material, spores and invertebrate eggs, fish scales, arachnids,
detritus, sand and mud have also been reported from gut
content analysis.
 Larvae are capable of independent feeding by 5 days – this is
necessary as yolk supplies are largely depleted by the end of
the first week.

10.  Zebrafish are broadcast spawners that release eggs and sperm in
a cloud over the substrate.
 They do not require a seasonal change in their day length to
bring them into a breeding state. When maintained under
laboratory conditions, they can be encouraged to breed
throughout the year, with females spawning every 2-3 days.
 A female generally produces around 100 transparent eggs in a
single spawning (can range between few eggs to over 1000 i.e.,
approx. 300-600). There is no parental care of the offspring
post-laying.
 Eggs have a diameter of about 1.0 - 1.5mm.

11. Life span:
 In the wild – 1 to 2 years.
 In the laboratory – 31/2 to 51/2 years.
Growth Stages:
0-72 hours post fertilization Embryos
72 hours – 13 days post fertilization Early larvae
14 days – 29 days post fertilization Mid larvae
30 days – 3 or 4 months Juveniles
When sexually mature Adults
Very rapid
development –
large no. of
experiments
possible in short
time period.

12. EMBRYO Zygote (0-0.75hr)
Cleavage (0.75-2.25hr)
Gastrula (5.25-10hr)
Juvenile (3-4 months)
Larva (72hr – 30 days
Hatching (48-72 hr)
Segmentation (10-24 hr)
Blastula (2.25-5.25hr)
Adult
Pharyngula (24-48 hr)
GROWTH STAGES
Embryo

13.  Possess sense of vision, olfaction, hearing,
balance.
 Robust circadian rhythm.
 Perceive threat.
 Shoaling behaviour.
 Similarity with humans
- Organ
- Genetic

14. ISSUE RECOMMENDATION
Housing systems All recirculating water system – water filter
system, water chemistry monitoring, germicidal
UV irradiation, light and temp. control units
Temperature Optimum range: 26 – 28.5 ˚C
Embryos : 28.5±0.5 ˚C to 120 hpf
Larvae and Adults: 24 – 29 ˚C
Dark-light cycle Standard static dark-light cycle
Environmental
enrichment
Zebrafish are a shoaling species and should be
kept in groups. Single ZF housing should be
limited to restricted periods.
Feeding 2-3 times a day with a combination of
dry and live feeds.

15. Sanitation of
equipment &
Hygiene
SOPs, Disinfection of equipments after
each use, Use of hand disinfection to
avoid cross contamination & zoonosis.
Health monitoring
and Diseases
Isolation/Quarantine, sanitation, hygiene,
sick should be discarded soon, water
quality, equipment maintenance,
vertenarians, euthanasia?
Water quality Monitoring and documentation of the
parameters typically done on a daily basis
and for some on a weekly basis
Breeding,
Identification
Avoid repeated inbreeding.
Date of fertilization, number of fish & key
genetic info needs to be traceable.

16. Zebrafish
Developmental
biology
Physiology
Toxicity
Genetics
Drug Discovery
Neuroscience
Cardiovascular
Disease Model
Little fish with mighty aspects

17.  The use of zebrafish as a model organism was pioneered at
the University of Oregon, USA, by George Streisinger in
1970s. He is the “Founder Father” of Zebrafish
Developmental and Genetic Research.
 Although zebrafish research began in early 1970s, it is
only since the 1990s/2000s that it has shot to prominence.
 In 1990s, the first large scale mutagenesis of zebrafish was
conducted by Christiane Nusslein Volhard, Oxford
University, UK., to identify developmental mutations.
 In 1995, Thomas Look of Dana-Faber Cancer Institute,
Boston used the translucent zebrafish in cancer studies.

18.  On the basis of analysis of 17,151 records on zebrafish
from ZFIN – the research performance on this model
organism has been evaluated. The earlist research work
on ZF as reflected in it goes back to 1951, after rather
slow growth till the 1980s, research on ZF gained
momentum in the 1990s. Analysis shows a rapid &
consistent increase in the publication output with 226
publications in the year 1996, to 1929 publications in
the year 2012.

20. Zebrafish research
centres & locations in
India

21.  What is a model organism?
 Why use zebrafish?
 Zebrafish as a Replacement
alternative
 Genetic similarity to humans.
 Ethical consideration?

22. Clinical
research
Preclinical Research

23. eye
heart
swim
bladder
muscle block
segments
ear
brain
notochord
~3.5 mm
Zebrafish embryo – 5dpf
Genetic

24.  Mostly used strains:
 According to ZFIN website,
>800 biological laboratories
around the world conduct basic
and applied research with ZF.
 These labs. use ZF to study
human diseases (neural disorders,
cancer, infectious diseases, CVD,
kidney diseases, diabetes, blindness, deafness, digestive
diseases, hematopoiesis and muscle disorders).
Mostly used Strains
AB
Casper
Ekkwill
Nadia
Wild Indian
Karyotype
Wild Caught
Tubingen
Emerging Model System:

25. Zebrafish in Toxicity
studies
•Developmental toxicology
(Teratogenecity)
•General toxicology
•Forensic toxicology
Fish Embryo Acute Toxicity Test (FET)
by OECD – Test no.236:
• Duration - 96hpf

26. Compound
name
Activity Dose Effect/Toxicity
observed
Gentamicin Antibiotic 5µm
Cisplatin Anticancer drug 50µm
Doxorubicin Anticancer drug 30.3mg/l Teratogen, kidney,
CVS, liver
Dexamethasone Corticosteroid 324mg/l Liver, GIT, kidney
Caffeine Methylxantine drug 108.4mg/l Behavioral: musle
contraction or
spasticity
Methotrexate Anticancer drug 454mg/l Teratogen, GIT, liver,
kidney
Vinblastine Chemotherapeutic
drug
100µm
Quinine Antimalarial drug 200µm
Neomycin Antibiotic 10µm

27. Zebrafish in Drug Discovery

28. Disease ZF model Purpose of the study
Obesity •Diet-induced
•Transgenic model
•To modulate the process of obesity
•Drug Screening
•Treatment
Dyslipidaemia &
Atherosclerosis
•Diet-induced •To r/o Histopathological changes
•To describe lipid & lipoprotein
metabolism
NAFLD •Diet-induced
•Transgenic
•To deduce the mechanism
•To develop a therapy
•Developmental biology
Type 2 DM •Immersed in High
glucose soln.
•Diet induced
•Transgenic
HUMAN DISEASE MODELS IN
ZEBRAFISH

29. •Small & robust
•Vertebrate
•Simple nature of their natural environment
•Year round spawning
•High fecundity (~300-600 at a time)
•Fertilization is external
•Optical transparency
•Very rapid development
•Early embryo permeable to small compounds
•Similarities &
•Dissimilarities with humans
•Easy maintenance
•Genetic similarity to humans
•Low housing costs
•Research can run continuously
•Low cost per assay
•Embryos accessible noninvasively
•Live imaging
•Large no. of exp. in short time
•Suitable for drug testing
•Serves as biomedical models
•With some limitations.

30.  Bioassay of natural products
 Toxicological evaluation of drugs
 Drug discovery

31.  In the last few years, the use of zebrafish in
scientific research is growing very rapidly.
 Developmental biology research on zebrafish has
been the key area of research for all period.
 It is expected that in the coming years,
biochemistry and molecular biology and
neurosciences with regard to zebrafish will
emerge as leading areas of research.
 Application of Zebrafish model to evaluate the
bioassay, toxicity of herbal drugs particularly of
Unani system can be initiated in the future.