L. EISENHAUER, R. ESPINO, V. ROJAS, S. SPRINGER, G. GUZMÁN, C. PENA, AND E. RODRÍGUEZ
ABSTRACT
In the Caribbean and in Latin America, mosquitoes are irritating pests
that can oftentimes leave more than just an itch; they are also vectors
for widespread diseases such as malaria and dengue fever. For the purposes
of protecting both ecotourism and public health, it is necessary to
control mosquito populations. Research is being done to find natural
agents that can effectively serve as biological controls against larvae.
Since no prior research on mosquitoes has been done in Punta Cana, obtaining
a better understanding of the local mosquito populations was the goal
of this study. Focus was placed on determining which mosquito species
are present in the Punta Cana area, as well as the ecological characteristics
of their breeding habitats including water pH levels, surrounding vegetation,
and weather patterns. Over a two-month period, ten species were collected
from both salt and fresh water: Culex duplicator, Culex (Mel.) atratus,
Culex nigripalpus, Culex quinquefasciatus, Anopheles albimanus, Anopheles
crucians, Psorophora insularia, Psorophora pygmaea, Aedes aegypti, and
Aedes taeniorhynchus. Four different breeding sites were found in the
Punta Cana area; a transect was created at the largest site where individual
pools within the coral rock could be marked and identified for samples
and measurements. At all sites, it was discovered that precipitation
is directly correlated with life cycle patterns, and that many species
seem to prefer an average pH of 9. The average life cycle length for
A. aegypti was observed to be eight days, and P. insularia averaged
ten days. A. albimanus, the malaria vector, was the most frequently
caught species in the UV night traps. A. aegypti larvae was used to
perform preliminary work with bioassays to test for bioactive natural
products. Alpha-terthienyl proved to be a photoactive compound that
killed one hundred percent of the larvae within thirty minutes. Other
plant extracts found to be bioactive against mosquitoes were Zanthoxylum
martinicense root, Piper aduncum leaves and flowers, and Acacia macracantha
root.
Mosquitoes have been around for over one hundred million years. All
mosquitoes are not the same; in fact, there are over 3,500 species.
A mosquito is a small fly (Order Diptera) with a long and slender abdomen,
narrow wings, a long and firm proboscis, and long frail legs. The males
and sometimes the females feed on nectar. In the majority of species,
the female feeds on blood after mating in order to obtain protein for
her maturing eggs. Once mature, the eggs are laid in water and hatch
into aquatic larvae. After a few days they turn into pupa and then finally
hatch into the adult mosquito. (Pena 2001).
Seventy-four species of mosquitoes are known to exist in the Dominican
Republic (Belkin and Heineman 1973, Pena 1987). Some of these species
are important for matters of public health; for example, Anopheles albimanus
is a known vector of malaria, while Aedes aegypti carries dengue fever.
While these diseases are not necessarily serious problems in the immediate
Punta Cana area, they are large concerns in many other parts of the
Dominican Republic (Belkin and Heineman 1973, Pena 1987). Ecotourism
is a second reason why mosquito populations are being investigated.
Because Punta Cana’s major economic resource is tourism there is a need
for mosquito control. However, it is both difficult and expensive to
control adult mosquitoes using pesticides. Moreover, oftentimes these
chemicals are detrimental to the environment and surrounding biodiversity.
Our goal is to provide fundamental knowledge about the mosquitoes in
Punta Cana so that future research can be done on more environmentally
sound ways of controlling them. It is necessary to thoroughly understand
the details of each species’ life cycle, biology and habitats in order
to administer effective biological controls against the larvae.
Prior research has been conducted in other parts of the world on many
of the species that were also found to exist in the Punta Cana area.
Much research is devoted towards studying the species that are vectors
for disease. While many of these reports examine ways to control the
adult mosquito populations with insecticides, substantially fewer of
them discuss controlling the larvae with bioactive natural products.
MATERIALS AND METHODS
Ecological Study
Larvae
Breeding places were found either by word of mouth or by driving to
locations suspected to have water present. At each location, dippers
were used to examine the water for the presence of larvae. For each
pool with larvae present, the length, width depth and temperature of
the water was measured. Salinity, pH and dissolved oxygen levels were
also determined using an aquarium water testing kit. The amount and
type of vegetation in and around the pools was noted, along with the
color and turbidity of the water in each pool. Information from the
Punta Cana airport about the local weather patterns was obtained, and
a Global Positioning System (GPS) was used to mark the latitude and
longitude of each site.
The primary study site was a 60ft. X 100ft. area adjacent to the sea
and divided into 15 20ft. X 20ft. quadrants. A total of 88 pools of
water in coral rock holes were dispersed throughout the grid. Each pool
was then examined for the presence or absence of mosquito larvae within
it. Once a new life cycle began, ten pools noted as having the most
mosquitoes present were then selected as our sample and marked. Water
measurements were then made every few days on each of these ten pools.
To measure the amount of larvae present, a total of ten random dips
were taken. After each dip, the larvae were counted and placed in a
separate container. After counting the larvae, they were returned to
the sampled pool.
In addition, ten very small pools of water containing larvae were found
in front of a house in Juanillo (a small town in the Punta Cana area)
and were also used in this study. Other dispersed sites were marked
and studied as well.
Adults
To gather information about the most active species of adult mosquitoes
at night in the Punta Cana area, night traps were used. Adult mosquitoes
were captured in the field using battery-powered UV light night traps
suspended from trees in unlit areas. Captured mosquitoes were classified
using a key and then placed in the cage for observation or mounted (Belkin
1970). A stereoscope was used for observation of mounted mosquitoes.
Biology
We were interested in the pattern of the life cycles of both P. insularia
and A. aegypti. A. aegypti was chosen for study because this species
is often used in laboratory experiments to conduct bioassays and because
its eggs can be easily obtained. P. insularia was selected because it
is a very common mosquito in the area: a large breeding site was found
which allowed for daily observations to be made. A. aegypti eggs were
collected from the Centro Nacional de Control de Enfermedades Tropicales
(CENCET) in Santo Domingo (although they can also be found in Punta
Cana) and raised in artificial containers both inside and outside of
the lab. Comparisons were made between the influences of the differing
environmental factors of the two containers. Brewer’s yeast was fed
to the larvae every two days. The P.insularia eggs are difficult to
obtain and hatch in a laboratory; therefore, the larvae were observed
in their natural environment at the coral rock holes.
Estimations were made about the timing of each stage of the life cycle
for both species. Once the larvae transformed into pupa, the containers
were placed into screened metal cages. Once the pupa metamorphosed into
adults, they were fed sugar. Blood was also provided for the females
by having a participant place his arm into the cage each night. Having
this blood meal allowed for the females to then lay their eggs in a
container of water that was placed in the cage. A layer of napkins lined
the inside of the container and served as a surface on which the females
could to deposit their eggs. For any eggs that hatched, observations
and procedures were repeated.
Bioassays
Bioactive natural products may possibly be used in the future as a means
of controlling mosquito larvae without damaging the environment. Preliminary
testing was done on A. aegypti larvae to search for potentially bioactive
compounds in organic extracts. Organisms tested were Zanthxylum martinicense
(root), Acacia macracantha (root), Piper aduncum (flower and leaf),
Opuntia ficus-indica (stem), Ravovlfia tetraphylla (root), the sponge
Pseudoceratina crassa, the paratoid gland of the frog Bufo marinus,
and a compound derived from the Asteraceae family, alpha-terthienyl.
For each bioassay, 25mL of water was deposited in a petri dish containing
mosquito larvae. Four drops of each extract were dropped into the dish
using a pipette. Four drops of ethanol were added to the control group.
Larvae were checked every fifteen minutes for the first hour and then
at the end of the twenty-four hours.
RESULTS
A total of ten species of mosquitoes were identified in four breeding
sites in the Punta Cana area: Culex duplicator, Culex (Mel.) atratus,
Culex nigripalpus, Culex quinquefasciatus, Anopheles albimanus, Anopheles
crucians, Psorophora insularia, Psorophora pygmaea, Aedes aegypti, and
Aedes taeniorhynchus. Species were identified using guides by Belkin
(1970) and Bram (1967). See Table 2 for a complete list of the species
found and their individual breeding places.
Coral Rock Holes
The only species observed in the coral rock holes by the seaside was
Psorophora insularia. It is known as a salt water mosquito; however,
at no point in the life cycle were the larvae found in the plot closest
to the shoreline. During the peak of the life cycle, the larvae counted
in the ten sampled pools ranged from 68 to 522, the average being 248.
The average values for pH, salinity, dissolved oxygen and temperature
were 9, 11.8??, and 9.8??, 36.8C respectively. The average area of the
pools was 88.5 inches X 46.8 inches. The minimum and maximum water depths
were 1.64 inches and 6.2 inches, with an average of 3.25 inches.
The length of the life cycle for P. insularia in its natural habitat
was observed to be an average of ten days. The time span from egg to
pupa was approximately eight days, and from pupa to adult was within
48 hours. In the artificial containers at the lab, the life cycle length
averaged 18 days, with a time span of 16 days from egg to pupa, and
within 48 hours from pupa to adulthood.
It is also interesting to note that there is a direct correlation between
the amount of rainfall and the mosquito population. As the month of
July progressed the rainfall decreased and the number of mosquito also
decreased. [See Figure 2 for the comparison between rainfall and number
of mosquito larvae].
Juanillo
In Juanillo, a roadside branching off the Punta Cana Marina, two adjacent
breeding sites were found; one site was a large ground pool, and the
other site was a series of small rock holes and crab holes. Four species
of mosquito larvae were found residing together in the ground pool:
Anapholes albimanus, Culex duplicator, C. nigripalpus, and C. qinquefasciatus.
The pH, salinity and oxygen levels were 10, 0?? and 8??, respectively.
The depth of the pool varied with rainfall and at times dried up completely.
Noted characteristics of the water included a reddish tint, high turbidity
and a stagnant smell. In the coral rock and crab holes, Aedes taeniorhynchus,
Psorophora pygmaea, Anopholes albimanus, A. crucians, Culex nigripalpus
were found. [See Table 2 for the distribution of species in each pool].
The depths ranged from 2 to 17.5 inches, with the majority of the pools
averaging three inches. The average area of the pools was 18.78 inches
X 14.1 inches. The average pH, temperature and oxygen levels were 8,
32C and 3??, respectively. The coloration, smell and vegetation in the
pools varied.
Ecological Reserve
There are several lagoons within the ecological reserve. Mosquito larvae
were found in small areas of water along the trail or adjacent to the
lagoons. No larvae were found in the actual lagoons. Adults of Anopheles
crucians and Culex (Mel.) atratus were all found near Laguna Baygua,
one of the lagoons. The area of the water was ten feet by two feet.
Very high vegetation levels were noted throughout the water. The pH,
salinity and oxygen levels were 8, 22?? and 7??, respectively.
Bioassays
The life cycle of Aedes aegypti in the artificial containers averaged
eight days. The time span from egg to pupa was six days and pupa to
adult averaged two days. Alpha-terthienyl proved photoactive, causing
100% mortality of the larvae within thirty minutes when exposed to UV
light. Within 24 hours, both flower and leaf extracts of Piper aduncum
also had significant activity (% mortality not available). Two roots
also showed bioactivity: the Zantoxylum martiniscense root induced 80%
mortality and the Acacia macracantha root induced 20% mortality. Dead
larvae were found in pools where an unidentified fungus was growing,
but the fungus was not collected and tested.
Night traps
Anopheles albimanus was the only adult mosquito species captured in
night traps at any of the four sites.
DISCUSSION
Life cycle lengths vary among mosquito species. It is important to be
able to predict the life cycle length of a target species so that they
can be controlled in an efficient and effective way. Results showed
that precipitation patterns determine the start of the life cycles.
Therefore, we can use this information to target a species at the peak
of its cycle. A second option is to attempt to control mosquitoes during
drier periods. When water supplies are limited, it is believed that
species form symbiotic relationships within the water pools. The species
would be concentrated in smaller areas, allowing bioactive compounds
to be easily administered to many types of species at once. It may not
be possible to kill all species with one type of bioactive component
due to biological differences among the mosquitoes. For example, a bioassay
using a photoactive chemical from the Asteracae family immediately killed
Aedes aegypti larvae, yet did not have any effect on P. insularia larvae.
No bioassays have been done comparing larvae of different species sharing
the same pools.
Variations in pH, oxygen levels, salinity, sunlight and individual characteristics
of species will determine what bioactive agent prove to be effective
in each case. For example, Bacillus thuringiensis (BT), is a commonly
used organic pesticide that is known to be effective against mosquito
larvae. BT would not be effective against Psorophora insularia, however,
for they are found in rock holes with a consistent pH of nine; BT requires
a pH less than seven to survive (Fields, Paul et al., 1991).
The results of the bioassays on Aedes aegypti larvae revealed promising
results. More tests need to be done to verify results and also to test
other types of larvae. Also, adverse environmental effects must be foreseen
before any implementation of biological control. The goals of using
natural agents to control mosquito populations are to be environmentally
friendly and to preserve as much biodiversity as possible. The untested
fungi found in pools of dead would be the most ideal component to use
for this species since it naturally grows in the Psorophora insularia
habitat.
More research on the mosquito populations in Punta Cana must be done
to better understand each of the species. Lab results are often unreliable
when at tempting to accurately predict what will occur in natural habitats.
Therefore, on-site observational studies and tests are highly recommended.
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Clark, G. 2000. Mosquito vector control and biology in Latin
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Fields, Paul G., John Arnason, et al. 1991. Phototoxins as insecticides
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