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Animal Care and Use Statement

Protocol Review Form June 2016

University of Santo Tomas

Institutional Animal Care and Use Committee
ANIMAL CARE AND USE STATEMENT
(Protocol Review Form)

UST-IACUC Code: ____________________ Application Fee Received by: _____________

(PRINT Name & Sign)
BAI Certificate No.: ____________________

(For UST-IACUC)
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Instructions. (a) Please complete the form by giving all the details asked for.
(b) Have the Protocol Review Form SIGNED and DATED by your respective ADVISER/S &
DEAN.
(c) Submit the accomplished Protocol Review Form to the IACUC Office at the Research
Center for the Natural Sciences (Rm. 202 TARC) for review and approval prior the conduct of
the scientific work involving animals.

RESPONSIBLE PERSON OR PRINCIPAL INVESTIGATOR [PRINT Complete Name,

Student Number & Contact Details (cell phone number and email address)]

Name: Diana Patricia D. Bala Cellphone number: 09175627781

Student Number: 2014 - 062605
Email Address: jrmissiona@gmail.com

Name: Allyssa Jiselle M. Cabalonga Cellphone number: 09178808647

Student Number: 2014 - 062663
Email Address: allyssajiselle@gmail.com

Name: Caitlene Joy C. Mejia Cellphone number: 09179701913

Student Number: 2014-065613
Email Address: caitlenemejia@yahoo.com

NAME OF ADVISER/S &/OR CO-ADVISER/S [PRINT Complete Name, College of

Affiliation & Contact Details (cell phone number and email address)]

Adviser: Asst. Prof. Jose Francis V. Abrantes, M.Sc.

Affiliation: Department of Biology
College of Science and
University of Santo Tomas
Email Address: fabrantes123@yahoo.com

Co-adviser: Prof. Gina R. Dedeles, PhD.

Affiliation: Department of Biology
College of Science and
Research Center for Natural and Applied Sciences
University of Santo Tomas
Email Address: grdedeles@yahoo.com
Cellphone number: 09185710803

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PROCEDURE(S) OR TITLE OF RESEARCH/STUDY:

Isolation and Identification of Marine yeasts from Mangrove leaves as feed supplement for
Oreochromis niloticus

PURPOSE/OBJECTIVES:
This study aims to isolate and identify marine yeasts from mangrove leaves, to screen protein
content of identified marine yeast, and to evaluate their efficacy as feed supplement to
Oreochromis niloticus

DURATION OR TIME FRAME:

8 weeks

BACKGROUND AND SIGNIFICANCE OF THE PROCEDURE OR RESEARCH:

Oreochromis niloticus, also known as the Nile Tilapia, is one of the useful species of
cichlids. Its fast growth, firm and tasty flesh, resistance against harsh conditions and ease
production of fingerlings under captivity makes it as an important species in aquaculture.
The characteristics of O. niloticus make it as an excellent model for studying
developmental in cichlids because of its high fecundity, a breeding capability which is 2-3
week spawning cycle, and a short generation period.
Marine microorganisms like marine yeasts live in extreme environments, providing
a unique potential for the synthesis of functional biomolecules (Connell et al., 2008). A
true marine yeast must be able to grow on or in a marine substrate (Kutty & Philip, 2008).
Marine yeasts are yeasts that are isolated from marine environments, can grow better on a
medium prepared using seawater rather than fresh water (Chi et al., 2010). Since almost all
yeasts can grow and tolerate high salinity concentrations, therefore, salinity tolerance does
not distinguish marine species from terrestrial species (Kutty & Philip, 2008). According
to reports, marine yeasts are able to produce many bioactive substances, such as amino
acids, glucans, glutathione, toxins, enzymes, phytase and vitamins with potential
application in the food, pharmaceutical, cosmetic and chemical industries as well as for
marine culture and environmental protection (Chi et al., 2009; Sarkar et al., 2010). The
factors affecting the distribution of marine yeasts include currents, migration of marine
organisms, and contamination from terrigenous sources (van Uden & Branco 1963, Fell
1967, Vogel et al. 2007, Kutty & Philip 2008). Marine yeasts are known to be agents of
biodegradation. Utilization of this potential in the biotransformation of cheap raw materials
or waste matter into value-added products will be a highly rewarding endeavor.
Biotransformation and the partial conversion of raw material into yeast biomass are very
significant in the nutritional composition of yeast and its potential use as animal or
aquaculture feed (Sarlin, P. J., 2005)
Marine yeasts are commonly found growing on mangrove leaves and on bodies of
water. These organisms are not well studied in the Philippines and thus this research could
contribute to knowing more about the organisms. The findings of the study can have a great
contribution to the fishing industry. This study can help achieve the optimum growth of
Nile tilapias and decrease its mortality rate. It could be cost-effective, helps lessen the use
of commercially produced feeds and use natural products as an alternative.

DESCRIPTION OF METHODOLOGIES/EXPERIMENTAL DESIGN:

A. Type of animal to be used:
Juvenile Tilapias (Oreochromis niloticus)
B. Source of the animals:
University of the Philippines National Institute of Health Building,
623 Pedro Gil Street,

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Ermita, Manila, 1000 Metro Manila

C. Reason/Basis for selecting the animal species:
Model animal for polyclonal antibody production (Hanly et al., 1995)

D. Sex and number of animals:

Sex/Age: Male or Female/1-3 days old
Number of animals per treatment: 10
Total number of animals: 50

E. Quarantine and/or acclimation or conditioning process

Indicate where: Animal house, UST Thomas Aquinas Research Complex (5th floor)
Duration: 60 days

Conditions
F. Animal care procedures
1. Cage Type & Size: 30 X 36 X 42 steel cages (elevated)
Type of Bedding: none used
2. Number of animals per cage: 10
3. Cage cleaning method:
Aquarium is equipped with air pump and filter. The water will be changed every
week.
4. Room temperature: 25.5 1.4 C
pH: 7.9 0.2
Humidity: 50% humidity
Ventilation: A clean, dirt and smoke free environment with fans for the
rabbits
Lighting: 12 hours light and 12 hours dark
5. Animal diet and feeding
Diet concentrations: 100% feeds
80% feeds 20% marine yeasts
70% feeds 30% marine yeasts
60% feeds 40% marine yeasts
50% feeds 50% marine yeasts
Feed: Muesli Rabbit Feeds (BioResearch)
Water: Purified Drinking Water (Absolute Distilled Drinking Water, Asia Brewery,
Inc.)

G. Experimental or animal manipulation methods

The protocol that will be used will be based on Thermo Fisher Scientifics Custom Rabbit
Polyclonal Antibody Production Protocols (Thermo Fisher Scientific, n.d.)
1. General description of animal manipulation methods
Nile Tilapias will be used to measure the efficacy of marine yeasts as a feed
supplement. Different diets will be fed to the tilapias, and their body weight and length
will be measured after 60 days

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2. Dosing method:
Antigen shot: Two (2) ml of the synthetic peptide admixed with 2 mL of
complete/incomplete adjuvant.
The initial antigen shot (2nd week) will make use of a complete adjuvant preparation
while the booster doses (4th and 6th week) will use the incomplete adjuvant preparation
(refer to table below).
Two hundred l of the antigen will be subcutaneously injected in 10 sites (dorsolumbar
to the ventrothoracic and abdominal regions; refer to figure below).

3. Specimen or biological agent:

Blood will be collected through the auricular marginal vein using a 5 ml 21-gauge
needle syringe. Three (3) ml of blood will be collected once every week except for the
final (8th) week where 5-10 ml of blood will be collected.

Week 1 2 3 4 5 6 7 8
Antigen Shot
Blood Collection

Diagram of a rabbit showing the injection sites [red spots; on both left and right sides (total of 10)].

4. Animal examination procedures and frequency of examinations: animals will be

examined daily by the researcher.
1/2 cup of pellets per 6 lbs. of body weight daily
Drinking water must be constantly supplied
Check for cages and lock stability
Regular check of humidity and temperature of the animal house

5. Use of anesthetics: none.

i. Where will this be bought? NA
ii. Who will administer? NA
iii. Licensed veterinarian: Name & License no. NA

6. Surgical procedures: none.

7. Euthanasia: none.

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8. Animal Disposal:
Veterinarian will be informed of any dead animal that has been used in the experiment.
Dead animals will be properly identified, placed in disposable properly labeled yellow
plastic bags, and maintained at refrigeration temperature until disposal in accordance with
the institutional and government regulations. Dead animals will be given to the technician
for proper disposal. Should the animals be alive after the experiment, they would be put up
for adoption by the researchers.

H. Is there a non-animal model applicable for the procedure/study? If so, please provide the reasons
for not using it. None.

I. Indicate the names and qualifications of all personnel who will be responsible for conducting the
procedures.

Name: Diana Patricia D. Bala

Student Number: 2014 - 062605
Email Address: jrmissiona@gmail.com
Cellphone number: 09175627781

Name: Allyssa Jiselle M. Cabalonga

Student Number: 2014 - 062663
Email Address: allyssajiselle@gmail.com
Cellphone number: 09178808647

Name: Caitlene Joy C. Mejia

Student Number: 2014-065613
Email Address: caitlenemejia@yahoo.com
Cellphone number: 09179701913

J. References

K. DECLARATION BY THE RESPONSIBLE PERSON:

I ACCEPT RESPONSIBILITY FOR ASSURING THAT THE PROCEDURES/ STUDY WILL BE

CONDUCTED IN ACCORDANCE WITH THE APPROVED PROTOCOL.

I ASSURE THAT ALL PERSONNEL WHO USE THIS PROTOCOL AND WORK WITH
ANIMALS, HAVE RECEIVED APPROPRIATE TRAINING/ INSTRUCTIONS IN
PROCEDURAL AND HANDLING TECHNIQUES, AND ON ANIMAL WELFARE
CONSIDERATIONS.

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I AGREE TO OBTAIN WRITTEN APPROVAL FROM THE INSTITUTIONAL ANIMAL CARE

AND USE COMMITTEE PRIOR TO MAKING ANY CHANGES AFFECTING MY PROTOCOL.
I ALSO AGREE TO PROMPTLY NOTIFY THE IACUC IN WRITING OF ANY EMERGENT
PROBLEMS THAT MAY ARISE IN THE COURSE OF THIS STUDY, INCLUDING THE
OCCURRENCE OF ADVERSE SIDE EFFECT.

Signature over Printed Name of the Responsible Person:

Date:______________________
Diana Patricia D. Bala

Date:______________________
Allyssa Jiselle M. Cabalonga

Date:______________________
Caitlene Joy C. Mejia

Date:______________________
Asst. Prof. Jose Francis V. Abrantes, M.Sc.,

Date:______________________
Prof. Gina R. Dedeles, Ph.D.

Prof. John Donnie A. Ramos, PhD Date:______________________

Dean, UST College of Science

Daria L. Manalo, DVM Date:______________________

Institutional Veterinarian

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