Blood-feeding Frequency and the Number of Eggs of Aedes aegypti

Anh Nguyen
March 2017
Academic Advisor: Brandy Fleming Academic Advisor, Undergraduate Programs
Plant Pathology, Entomology & Nematology
Global Disease Biology & Entomology Majors
Research Mentor: Christopher Barker - Department of Pathology,
Microbiology, & Immunology
School of Veterinary Medicine

Abstract
Mosquitoes are vectors that transmit several diseases in human history such as plague,
malaria and Zika. Adult female mosquitoes need blood to develop their eggs. Studying
the basic needs of bloodsucking arthropods is a means toward understanding their
behavior in relation to the vertebrate host. [introduce Aedes aegypti here and why use this
breed instead of other breed]The results reported in this study extend the information on
the relation between the numbers of eggs that are developed by Aedes aegypti and the
frequency of their blood meals. In this study, it was investigated whether the frequency of
the blood meals affected the number of eggs of adult female Aedes aegypti.[similar to the
previous sentence] Mosquitoes were separated into two groups with different blood-
feeding frequencies for 25 days. One group was bloodfed everyday. The other group was
bloodfed every three day. Their eggs were collected every seven daysand counted after
each collection. The results showed that the moquitoes bloodfed more frequently laid
more eggs than the mosquitoes bloodfed less frequently.

Introduction

Mosquitoes are members of the family Culicidae. There are about 3500 species
worldwide (CDC, n.d.). Mosquitoes are found on every continent except Antarctica. They
can be found everywhere on earth, from deserts to tropical forests, from high mountains
to ocean tidal zones. Male mosquitoes feed on nectar while female mosquitoes can blood-
feed on a wide range of vertebrates including mammals, birds, reptiles and amphibians
(Kay et al., 2007). Therefore, mosquitoes are considered one of the most dangerous
animals, since they kill millions annually, sicken hundreds of millions and cost nations
billions. [how do they kill animals and people? They transmit disease or something?]

Aedes aegypti is a member of the family Culicidae, genus Aedes. It is the vector for
transmitting several tropical diseases, such as dengue, yellow fever, chikungunya and
Zika. The mosquito can be recognized by the white markings on its legs and on the upper
surface of its thorax. Aedes aegypti originated in Africa, but now it can be found in
tropical and subtropical regions throughout the world (Womack, 1993). In order to
survive and reproduce, female Aedes aegypti mosquitoes use two different food sources.
They need sugar for energy, which can be taken from sources such as nectar. They also
need blood as a source of protein for egg development. After taking a blood meal, it takes
the mosquitoes two to three days to digest the blood. Blood is stored in the stomach and
synthesized by the the midgut into free amino acids. These are used as building blocks for
the synthesis of egg yolk proteins. Blood restriction can affects the number and the
quality of eggs. [why is it used as an experiment target?]

In this experiment, the data provided shows the effect of blood availability in the number
of eggs. These data not only form the basis for understanding of mosquito habit and
reproduction but are also directly relevant for public health due to mosquitos role as a
vector of several infectious diseases.
Materials and Methods

Experimental system
The Aedes aegypti used in the experiment was 1-week-old adult females obtained from
Davis Arbovirus Research and Training, University of California, Davis). Aedes aegypti
occurs throughout the tropics and subtropics. During the adult stage, mosquitoes can
experience periods of blood restriction and competition.

Experimental design

The experiment was run in a climate chamber set to 25C, 80% relative humidity and at
12-hour-light and 12-hour-dark regime. 60 adult female mosquitoes were caged and
separated into 2 groups of 30 naming Group A and B. Moistened cotton balls with 10%
sugar solution were set up in the mosquito cages and replaced every 48 hours. These
cotton balls were removed 4 hours before the mosquitoes were bloodfed. Female
mosquitoes were bloodfed with 3ml of sheep blood each time using the Hemotek
membrane feeding system. Group A was bloodfed daily while group B was done every 3
days. Seed germination paper were used to collect eggs. Each cage had a plastic cup
containing seed germination paper and was one third filled with deionized water. The
papers were collected every 7 days and replaced with new ones. The experiment was run
in 28 days.

Egg counting

The number of eggs were counted by scanning the egg papers and measuring with
IMAGEJ.

Statistical analysis

All data were analyzed using R 3.2.2 software. The number of bloodfed mosquitoes and
the number of eggs were examined by t-test. The relationship between the number of
bloodfed mosquitoes and the number of eggs was examined by linear regression.

Results

The total number of daily bloodfed mosquitoes were more (mean = 3.12 moquitoes,
standard error = 2.18) than the total number of every-3-day bloodfed mosquitoes (mean =
1.32, standard error = 2.21), (p < 0.05) (Firgure 1)
 100

Number of bloodfed
80
Firgure

mosquitoes
1. 60
Number
of 40

20

0
Everyday Every 3 days
bloodfed mosquitoes in 25 days.

The total number of eggs of daily bloodfed mosquitoes were more (mean = 286.75 eggs,
standard error = 40.01) than the number of eggs of every-3-day bloodfed mosquitoes
(mean = 172.00 eggs, standard error = 48.81) (p < 0.05) (Figure 2).

1500

1200
Figure
Number of eggs

2.
900

600

300

0
Everyday Every 3 days
Number of eggs collected in 28 days.
The effect of the frequency of blood meals on the number of eggs produced was
significant (Figure 3)

1500

1200
Number of eggs

900

600

300

0
33 78
Number of bloodfed mosquitoes

Figure 3. The relationship between the number of bloodfed mosquitoes and the number of
eggs produced.

Discussion

The experiment shows that the frequency of blood meals has effects on the mosquito
reproduction success, particularly the number of eggs.

In the everyday group, there were total 78 times of blood meals taken while in the every-
3-day group, there were 33 blood meals taken. 78 was the number of blood meals taken
by the daily fed group as 33 was counted for the 3-day-fed group. There were 61 less
blood meals taken in the every-3-day mosquito group than in the everyday group. The
blood meal difference between two groups was 61. Moreover, there were 11477 eggs
produced in the everyday group, which was more than the every-3-day group (688 eggs)
1187 eggs. 1477 eggs were produced by the daily fed group and about 2 times more than
the 3-day-fed group (688 eggs).

Female mosquitoes requires blood to develop eggs. After fed, up to 3 days are needed for
mosquitoes to digest blood. In the daily fed group, female mosquitoes are fed and rested
without a concern of shortage of blood source. This procedure increased the number of
blood meals taken. In the other group, female mosquitoes were bloodfed every 3 days.
There was rest time for them to digest the blood and lay eggs. While in rest time each
mosquito competed with each other to be fed. The hungry mosquitoes had to fight for
survival in the limited blood source. Mosquitoes in this group had less blood meals than
the other group. More well-fed mosquitoes resulted in more eggs produced and vice
versa.

In this experiment, 30 mosquitoes were caged together as a group. The number of

mosquitoes being actual fed were not counted precisely due to the crowded population.
Some mosquitoes were not bloodfed at all whereas some had more than one meal. Some
factors impacted the total number of eggs. Some eggs were autogenous and hatched in
the egg cup before being collected. Moreover, different number of dead mosquitoes were
found in the two groups. Therefore, the correct number of eggs being produced could not
be counted.

Conclusion

The experiment proved that frequency of blood meals has strong effect on the number of
mosquito eggs produced in the same other conditions. This result could have a significant
role in the evolution of mosquito reproduction. Other interesting experiments should be
considered are the co-evolvement of mosquitoes with their host and the reproduction of
mosquitoes without the limited source of sugar.

References
M. Womack (1993). "The yellow fever mosquito, Aedes aegypti". Wing Beats.