MASS REARING OF THE RED PALM WEEVIL,
RHYNCHOPHORUS FERRUGINEUS OLIV., ON SUGARCANE AND
ARTIFICIAL DIETS FOR LABORATORY STUDIES:
ILLUSTRATION OF METHODOLOGY
Walid Kaakeh, Mahmoud M. Abou-Nour, and Ahmed A. Khamis
Department of Plant Production, Faculty of Agricultural Sciences,
UAE University, P. O. Box 17555, Al-Ain, U.A.E.
ABSTRACT
A method for laboratory mass rearing of the red palm weevil
(Rhynchophorus ferrugineus Oliv.) (RPW) were developed. Weevils,
initially obtained from the field, were maintained on the stems of sugarcane.
Prior to mass rearing, several artificial diets were formulated and preliminary
evaluated for development of the R. ferrugineus. Materials used for
preparations of various diets were: oats, coconut cake, coconut fruit pieces,
canned and/fresh pineapple, sucrose, molasses, egg yolk, salt, yeast,
vegetable oil, potatoes, soybean flours, date palms leaves and palm fiber
sheath, sugarcane fibers, bacto-agar, multi-vitamins, preservatives, and
water. Oat and white bean diets were preferred by 1st to 3rd larval instars,
while oats + fibers preferred by 4th to 5th larval instars. Larvae fully
developed on artificial diets and molted four times during their development
failed to construct cocoons because of the unavailability of fibers (palm or
sugarcane. Facilities, materials required, diet preparation and procedures,
and practical difficulties of rearing methods are discussed.
Additional Index Words: Rhynchophorus ferrugineus, pheromone,
trapping, palm trees
INTRODUCTION
The red palm weevil (RPW), Rhynchophorous ferrugineus Oliver
(Coleoptera: Curculionidae), is an economically important, tissue-boring
pest of date palm in many parts of the world. The insect was first described
in India as a serious pest of coconut palm (Lefroy, 1906) and later on date
palm (Lal, 1917; Buxton, 1918). The insect is a major pest of date palm in
some of the Arabian Gulf States including Saudi Arabia, United Arab
Emirates, Sultanate of Oman, and Egypt (Cox, 1993; Abraham et al. 1998).
34 5
The agroclimatic conditions prevalent in this region and the unique
morphology of the crop, coupled with intensive modern date palm farming,
have offered the pest an ideal ecological habitat (Abraham et al., 1998).
Red palm weevil is a concealed tissue borer and spends all of its life
stages inside the palm tree. Damage symptoms can be categorized by one or
more of the following (Abraham 1998): presence of the tunnels on the trunk
and base of leaf petiole made by the feeding grubs, oozing out of thick
yellow to brown fluid from the tree, appearance of chewed up plant tissue in
and around openings in the trunk, presence of a fermented odor from the
fluid inside infested tunnels in the trunk, presence of adults and cocoons in
the leaf axils, fallen empty pupal/chewed up frass on the ground around the
palm, breaking of the trunk or toppling of the crown when the palm is
severely infested.
Research on the biology and control of R. ferrugineus, for many
projects conducted in the last four years at UAE University by various
researchers, required large numbers of weevils of various stages. First
attempt to develop a method for mass rearing of this pest was made by the
authors during this period in the UAE. Rearing methods of this and several
related species were reported in other countries and with similar species
(Rahalkar et al., 1972, 1978, 1985; Rananavare et al., 1975; Giblin-Davis et
al., 1989; Weissling and Giblin-Davis, 1995). The objectives of this study
were to (1) illustrate the methodology of mass rearing R. ferrugineus on
sugarcane, and (2) develop artificial diets for rearing R. ferrugineus.
MATERIALS AND METHODS
Insects
Various stages of R. ferrugineus were collected from infested date palm
trees in Masafi area in Sharja Emirate in 1997. Each developmental stage
was placed individually in covered plastic container. Portable wood saw
was used to facilitate collecting weevils from heavily infested palm trees
(Figure 1).
34 6
Figure 1. Collection of various stages of R. ferrugineus from infested date
palm trees.
Rearing Room
Rearing of R. ferrugineus of various stages was carried out in a
controlled rearing room at the Plant Protection Laboratory of the Plant
Production Department, UAE University (Figure 2). The room was
maintained at 25 ± 2ºC and 60-70% RH. The photoperiod was approximately
12:12 L:D. The room contained three large working benches, electrical
outlets, stainless steel sink, side boards, autoclave, and a refrigerator. The
room was also used as a media room for handling and preparing materials of
artificial diets.
34 7
Figure 2. Room for rearing R. ferrugineus at the Faculty of Agricultural
Sciences, United Arab Emirates University.
Equipments and Materials
Equipments and materials required for rearing R. ferrugineus on
sugarcane and artificial diets are given in Table 1.
Rearing on Sugarcane
Adults. Adults collected from the field (Figure 1) were cleaned and
kept (as group of at least 10 males and females but not sexed) in rectangular
plastic boxes with press-on tight-fitting lids, or kept as individual pairs of
males and females in 1 liter glass jars (Figure 3). All adults were provided
with at least 5 absorbent cotton wicks saturated with a 10% honey solution
for feeding and egg laying. Boxes and jars were staked side by side (or on
the top of each other) on working benches. Few holes were made on all lids
of boxes and jars for ventilation. Females lay their eggs on the cotton wicks
(i.e., oviposition site). Association of both sexes for 24 h ensures fertilization
of females and no further mating of females was necessary (Kaakeh, 1998).
Adults after emergence from cocoons were sexed and kept separately in
small jars for mating and egg laying. Sexing the adults was done according
to the presence of a series of black hairs on the dorsa, frontal part of snouts
of males and their absence in the females.
34 8
Table 1. Equipments and chemical materials used for rearing R. ferrugineus
on sugarcane and artificial diets.
• Blanace (up to 3 kg; triple beam balance or others)
• Electric blender/mixer
• Autoclave
• Refrigerator
• Portable wood cutting saw
• Rectangular plastic boxes
• Polyethylene collecting buckets
• Aluminum and plastic trays or boxes (various sizes) with tight-fitting
lids
• Glass and disposable plastic Petri dishes (50 and 100 mm In diameter)
• Camel-hair brushes (no. 2 preferred)
• Absorbent cotton or cotton wicks (sheets and bolls)
• Scissors
• Glass jars (1 liter)
• Measuring cylinders (50, 100, 200, and 500 ml)
• 40-mesh nylon netting
• Cork borer set
• Aluminum foil
• Transparent, polyethylene bottles, with tightly closing lid.
• Black fiber paper sheets
• Rubber bands
• Paper towels
• Absorbent cotton wicks
• Hand magnifiers
• Labpette digital micro-pipet, capacity 1-20 micro-liter
• Parafilm rolls (for sealing test tubes, flasks, Petri dishes, etc.)
• Eye wash bottles
• Disposable non-toxic particle masks
• Graduate and funnel brushes
• Glass shell and screw-cap vials
34 9
Figure 3. Rectangular plastic boxes and glass jars holding adults for mating
and oviposition. Plastic containers contained unsexed groups of
adults, while the glass jars hold pairs of sexed males and females.
Eggs. Cotton wicks holding eggs were removed from the oviposition sites
(i.e., large plastic boxes and small glass jars in figure 3) and placed in
separate boxes (Figure 4). Cotton wicks were wet with water to avoid
drying. Other eggs were transferred with the camel hair brush and placed on
wet filter papers inside the petri dishes for further studies. New cotton wicks,
saturated with a honey solution, were placed in all containers holding the
adult stages. After 2 to 3 days, larvae from hatched eggs were removed to
separate containers and provided with pieces of sugarcane stems for feeding.
Larvae. Newly hatched larvae on cotton wicks were transferred with a
camel’s hair brush to pieces of sugarcane stems (at least 15 mm in diameter)
(Figure 5). A small hole was made at the end of each piece of sugarcane
stem using a cork borer. One week after feeding, larger larvae were
transferred to larger fresh pieces of sugarcane stems (10 – 40 mm in
diameter; this was based on the size of larvae at different developmental
stages). Last larval instars made cocoons from the fibers inside the sugarcane
stems. When sugarcane was infested with Drosophilla flies, yellow sticky
traps were placed above the rearing containers as a control tool. Also, larvae
were reared individually (Figure 5) in sugarcane stems to avoid cannibalism.
35 0
Figure 4. Egg collection from cotton wicks and filter papers.
Figure 5. Handling and feeding of larvae in sugarcane stems.
35 1
Pupae. After 10-14 days of feeding of last larval instars, the stem pieces of
sugarcane were split open and cocoons were collected. Cocoons were placed
in a plastic containers or metal trays, wet with water as needed, and closed
with lids. Two weeks after collecting the cocoons, they were checked daily
for adult emergence. Adults were collected by hand and placed in plastic
containers (as unsexed groups of adults) or placed individually in glass jars
(as sexed, paired males and females).
Figure 6. Collection of pupae of R. ferrugineus from pieces of sugarcane
stems (a) and plastic containers holding cocoons (b).
Rearing on Artificial Diets
Preparation of Diets
Artificial diets were prepared for mass rearing of R. ferrugineus
because of the unavailability of sugarcane in UAE and this was a limiting
factor in culturing this insect. Diets were also developed to avoid the use of
expensive palm tissues for culture of weevils.
Table 2 lists materials used for the preparation of several artificial
diets. Different diets were evaluated in preliminary tests for larval and adult
biomass gain, survival, and the rate of development Three diets were
selected (Figure 7) and percentage of dry ingredients is listed in Table 3.
Diet A is an mainly an oat diet, Diet B is an oat diet plus palm and coconut
tissue, while diet C is mainly a white bean diet. Ingredients and water (1 - 2
liter of water for diets weighing 500 - 1000 g) were blended for
approximately 5 minutes. All diets included bacto-agar, multi-vitamins,
chemical preservatives. Bacto-agar was dissolved in water and added to
other ingredients. The mixtures of all diets were then autoclaved for 20 min
at 120ºC. Diets were poured in diet stainless-steel round trays or cups while
still warm. All trays and cups were stored at room temperature until
a b
35 2
required. Larvae were placed on diets after total coolness. As with rearing
R. ferrugineus on sugarcane, when artificial diets were infested with
drsophilla flies, yellow sticky traps were placed above the rearing containers
as a control tool. Also, fungal and bacterial contamination may develop if
chemical preservatives were not incorporated in the diets. This may also
occur if high quantity of water was mixed with the dry ingredients of diets.
Table 2. Materials used for the preparation of artificial diet for rearing R.
ferrugineus in the laboratory.
Food Materials
Oats Date palm leaves and fiber sheath
White beans Canned or fresh pineapple
Sugarcane bagasse Soybean flours
Fresh sugarcane stems Coconut fruit pieces
Fresh coconut cake Coconut fibers
Potatoes Vegetable oil
Egg yolk Wesson salt
Vitamin tablets* Brewer's yeast or other brands
Honey
Chemical preservatives
Methyl para-hydroxybenzoate **
Auromycin
4M Potassium hydroxide***
Sorbic acid****
Bacto-agar
___________________________________________________________
* vitamin tablets contained vitamins A, B1, B2, B6, B12, D2, E, K1,
Riboflavin, nictotinamide, and others.
** 14% solution In 95% ethyl alcohol. 5 ml water was added to 95 ml
absolute ethyl alcohol (95% solution) and then 140 g methyl parahydroxybenzoate
was dissolved in 95% ethyl alcohol (amount of 15
ml solution per 1 kg diet was used)
*** 56 g potassium hydroxide in 250 ml distilled water (the amount of use
5 ml solution per 1 /kg of diet was used).
**** 12.5% stock solution In 95% ethyl alcohol. About 125 g sorbic acid
was dissolved in 1 liter of 95% ethyl alcohol (the amount of 15 ml
solution per 1 kg of diet was used) (Rahalkar et al. 1985).
35 3
Table 3. Percentage of dry ingredients of three artificial diets used for
rearing R. ferrugineus.
% Dry Ingredient in
________________________________________
Dry Ingredient Diet A Diet B Diet C
Oats 57 48 15
White Beans - - 65
Palm and coconut tissues - 15 -
Sugar 22 19 10
Molasses 11 10 -
Brewers yeast 9 7 9
Salt 1 1 1
Figure 7. Artificial diets in small plastic trays or cups or in stainless steel
containers for feeding of larvae of various sizes.
Larval Feeding on Artificial Diets
Prior to mass rearing, several artificial diets were formulated and
preliminary evaluated for development of R. ferrugineus. Second instar
larvae were washed in tap water, weighed, and placed into holes made on
artificial diets to facilitate feeding. Larvae were transferred one per diet cup
with a fine camel hair brush (Figure 8). The weight for each larva (n = 8)
was recorded weekly for 6 weeks, and was then compared with its starting
35 4
weight. When larvae reached the end of the last larval instar, they were
placed in the stems of sugarcane for constructing cocoons. Diets were
checked daily for any dead larvae in which they were replaced.
Figure 8. Feeding of larvae on artificial diets.
RESULTS AND DISCUSSION
Previous attempts that were made by several researchers followed
several steps: Sugarcane was a good substitute for coconut for rearing R.
ferrugineus (Rahalkar et al., 1972); sugarcane was later incorporated in
nutrient agar for feeding young larvae and sugarcane stem pieces for feeding
of older larvae (Rananavare et al. 1975). Rahalkar et al. (1978, 1985)
improved the culture of R. ferrugineus by developing an artificial diet
containing sugarcane bagasse (fiber), coconut cake, yeast, sucrose, minerals,
vitamins, and preservatives. Giblin-Davis et al. (1989) cultured R. cruntatus
and R. palmarum on decomposing pineapple. In our study, there were
variations in the quality of developed diets made using different ingredients.
Diet A was mainly made by oats, diet B was made using oats and fibers,
while diet C was made using white beans. Diets A and B were preferred by
young larvae (1st to 3rd larval instars) while diet B was preferred by older
larvae (4th and 5th larval instars).
35 5
The average biomass of larvae at various stages, survival, percentage
of larvae that went into cocoon, and biomass and percentage of emergence of
adults varied greatly between the diets tested: a range of larval biomass after
three weeks of feeding on artificial diets was 0.25 – 2.5 g and after 5 weeks
was 0.5–4.5 g. The weight gained from feeding increased considerably in a
three week period (i.e., larvae with weight ranged from 0.5 to 0.11g
increased 40 to 80X, that is 4.18 to 4.54 g). The weight gained varied with
the type of diet provided to the larvae. The percentage of larval survival
ranged from 10 to 90%. The biomass of emerged adults ranged from 0.8 to 2
g. The percentage of adult emergence ranged from 10 to 80%.
Larvae were fully developed on artificial diets and molted four times
during their development (Figure 9). These larvae failed to construct
cocoons because of the unavailability of fibers (palm or sugarcane) or there
was not enough quantities of high fiber substrates in diet B. All larvae fed by
either of the three artificial diets were transferred to sugarcane stems for
making cocoons. Additional studies are required to search for alternative
food sources to develop diets that R. ferrugineus can fully develop on it
without transferring last larval instar to sugarcane stems for cocoon
construction.
Figure 9. Development of first instar larvae on artificial diet (a), and the
molting of fourth instar larva to the fifth larval stage.
The success of our three artificial diets (Table 3) in providing the necessary
nutritional requirement for molting and development was based on data
recorded ing various life parameters of R. ferrugineus on sugarcane and
artificial diets (a study conducted by the authors, see the next manuscript in
this proceedings). The study on life parameters of R. ferrugineus included
the following: fecundity or the number of eggs per female, egg viability or
the percentage of hatch, larval developmental period, larval biomass, pupal
35 6
period, percentage of larval survival, percentage of adult emergence,
fecundity, fertility, and female:male ratio.
LITERATURE CITED
Abraham, V. A., M. A. Al-Shuaib, J. R. Falleiro, R. A. Abozuhairah, and P.
S. P. V. Vidyasagar. 1998. An integrated management approach for
the red palm weevil Rhynchophorus ferrugineus Oliv. A key pest of
date palm in the Middle East. 3: 77-83.
Buxton, P. A. 1918. Report on the failure of date crops in Mesopotamia in
1918. Agric. Directorate, M. E. F. Bassarah Bull. No. 6.
Cox, M. L. 1993. Red palm weevil, Rhynchophorus ferrugineus, in Egypt.
FAO-Plant-Protection-Bulletin 41: 1, 30-31.
Giblin-Davis, R. M., T. J. Weissling, A. C. Oehlschlager, and L. M.
Gonzalez. 1994. Field response of Rhynchophorus cruntatus
(Coleoptera: Curculionidae) to its aggregation pheromone and
fermenting plant volatiles. Florida Entomol. 77: 164-177.
Lal, Madan Mohan. 1917. Rept. Asst. Prof. Entomol; Rept. Dept. Sagr.
Punjab, for the year ended 30th June, 1917.
Lefroy, H. M. 1906. The more important insects injurious to Indian
Agriculture. Govt. Press, Calcutta, India.
Kaakeh, W. 1998. The mating behavior of the red palm weevil,
Rhynchophorus ferrugineus Oliv. (Coleoptera: Curculionidae). Emir.
J. Agric. Sci. 10: 24-46.
Rahalaker, G. W., M. R. Harwalkar, and H. D. Rananavare. 1972.
Development of red palm weevil, Rhynchophorus ferrugineus Oliv.
Indian J. Entomol. 34: 213-215.
Rahalaker, G. W., A. J. Tamhankar, and K. Shanthram. 1978. An artificial
diet for rearing red palm weevil, Rhynchophorus ferrugineus Oliv. J.
Plantation Crops 6: 61-64.
Rahalaker, G. W., M. R. Harwalkar, H. D. Rananavare, A. J. Tamhankar,
and K. Shanthram. 1985. Rhynchophorus ferrugineus, pp. 279-286. in
35 7
P. Singh and R. F. Moor [eds.]. Handbook of insect rearing. Elsevier,
New York, NY. Vol. 1.
Rananavare, A. J. K. Shanthram, M. R. Harwalkar, and G. W. Rahalkar.
1975. Method for the laboratory rearing of red palm weevil,
Rhynchophorus ferrugineus Oliv. J. Plantation Crops 3: 65-67.
Weissling, T. J. and R. Giblin-Davis. 1995. Oligidic diets for culture of
Rhynchophorus cruntatus (Coleoptera: Curculionidae). Frlorida
Entomologist 78: 225-234.
RHYNCHOPHORUS FERRUGINEUS OLIV., ON SUGARCANE AND
ARTIFICIAL DIETS FOR LABORATORY STUDIES:
ILLUSTRATION OF METHODOLOGY
Walid Kaakeh, Mahmoud M. Abou-Nour, and Ahmed A. Khamis
Department of Plant Production, Faculty of Agricultural Sciences,
UAE University, P. O. Box 17555, Al-Ain, U.A.E.
ABSTRACT
A method for laboratory mass rearing of the red palm weevil
(Rhynchophorus ferrugineus Oliv.) (RPW) were developed. Weevils,
initially obtained from the field, were maintained on the stems of sugarcane.
Prior to mass rearing, several artificial diets were formulated and preliminary
evaluated for development of the R. ferrugineus. Materials used for
preparations of various diets were: oats, coconut cake, coconut fruit pieces,
canned and/fresh pineapple, sucrose, molasses, egg yolk, salt, yeast,
vegetable oil, potatoes, soybean flours, date palms leaves and palm fiber
sheath, sugarcane fibers, bacto-agar, multi-vitamins, preservatives, and
water. Oat and white bean diets were preferred by 1st to 3rd larval instars,
while oats + fibers preferred by 4th to 5th larval instars. Larvae fully
developed on artificial diets and molted four times during their development
failed to construct cocoons because of the unavailability of fibers (palm or
sugarcane. Facilities, materials required, diet preparation and procedures,
and practical difficulties of rearing methods are discussed.
Additional Index Words: Rhynchophorus ferrugineus, pheromone,
trapping, palm trees
INTRODUCTION
The red palm weevil (RPW), Rhynchophorous ferrugineus Oliver
(Coleoptera: Curculionidae), is an economically important, tissue-boring
pest of date palm in many parts of the world. The insect was first described
in India as a serious pest of coconut palm (Lefroy, 1906) and later on date
palm (Lal, 1917; Buxton, 1918). The insect is a major pest of date palm in
some of the Arabian Gulf States including Saudi Arabia, United Arab
Emirates, Sultanate of Oman, and Egypt (Cox, 1993; Abraham et al. 1998).
34 5
The agroclimatic conditions prevalent in this region and the unique
morphology of the crop, coupled with intensive modern date palm farming,
have offered the pest an ideal ecological habitat (Abraham et al., 1998).
Red palm weevil is a concealed tissue borer and spends all of its life
stages inside the palm tree. Damage symptoms can be categorized by one or
more of the following (Abraham 1998): presence of the tunnels on the trunk
and base of leaf petiole made by the feeding grubs, oozing out of thick
yellow to brown fluid from the tree, appearance of chewed up plant tissue in
and around openings in the trunk, presence of a fermented odor from the
fluid inside infested tunnels in the trunk, presence of adults and cocoons in
the leaf axils, fallen empty pupal/chewed up frass on the ground around the
palm, breaking of the trunk or toppling of the crown when the palm is
severely infested.
Research on the biology and control of R. ferrugineus, for many
projects conducted in the last four years at UAE University by various
researchers, required large numbers of weevils of various stages. First
attempt to develop a method for mass rearing of this pest was made by the
authors during this period in the UAE. Rearing methods of this and several
related species were reported in other countries and with similar species
(Rahalkar et al., 1972, 1978, 1985; Rananavare et al., 1975; Giblin-Davis et
al., 1989; Weissling and Giblin-Davis, 1995). The objectives of this study
were to (1) illustrate the methodology of mass rearing R. ferrugineus on
sugarcane, and (2) develop artificial diets for rearing R. ferrugineus.
MATERIALS AND METHODS
Insects
Various stages of R. ferrugineus were collected from infested date palm
trees in Masafi area in Sharja Emirate in 1997. Each developmental stage
was placed individually in covered plastic container. Portable wood saw
was used to facilitate collecting weevils from heavily infested palm trees
(Figure 1).
34 6
Figure 1. Collection of various stages of R. ferrugineus from infested date
palm trees.
Rearing Room
Rearing of R. ferrugineus of various stages was carried out in a
controlled rearing room at the Plant Protection Laboratory of the Plant
Production Department, UAE University (Figure 2). The room was
maintained at 25 ± 2ºC and 60-70% RH. The photoperiod was approximately
12:12 L:D. The room contained three large working benches, electrical
outlets, stainless steel sink, side boards, autoclave, and a refrigerator. The
room was also used as a media room for handling and preparing materials of
artificial diets.
34 7
Figure 2. Room for rearing R. ferrugineus at the Faculty of Agricultural
Sciences, United Arab Emirates University.
Equipments and Materials
Equipments and materials required for rearing R. ferrugineus on
sugarcane and artificial diets are given in Table 1.
Rearing on Sugarcane
Adults. Adults collected from the field (Figure 1) were cleaned and
kept (as group of at least 10 males and females but not sexed) in rectangular
plastic boxes with press-on tight-fitting lids, or kept as individual pairs of
males and females in 1 liter glass jars (Figure 3). All adults were provided
with at least 5 absorbent cotton wicks saturated with a 10% honey solution
for feeding and egg laying. Boxes and jars were staked side by side (or on
the top of each other) on working benches. Few holes were made on all lids
of boxes and jars for ventilation. Females lay their eggs on the cotton wicks
(i.e., oviposition site). Association of both sexes for 24 h ensures fertilization
of females and no further mating of females was necessary (Kaakeh, 1998).
Adults after emergence from cocoons were sexed and kept separately in
small jars for mating and egg laying. Sexing the adults was done according
to the presence of a series of black hairs on the dorsa, frontal part of snouts
of males and their absence in the females.
34 8
Table 1. Equipments and chemical materials used for rearing R. ferrugineus
on sugarcane and artificial diets.
• Blanace (up to 3 kg; triple beam balance or others)
• Electric blender/mixer
• Autoclave
• Refrigerator
• Portable wood cutting saw
• Rectangular plastic boxes
• Polyethylene collecting buckets
• Aluminum and plastic trays or boxes (various sizes) with tight-fitting
lids
• Glass and disposable plastic Petri dishes (50 and 100 mm In diameter)
• Camel-hair brushes (no. 2 preferred)
• Absorbent cotton or cotton wicks (sheets and bolls)
• Scissors
• Glass jars (1 liter)
• Measuring cylinders (50, 100, 200, and 500 ml)
• 40-mesh nylon netting
• Cork borer set
• Aluminum foil
• Transparent, polyethylene bottles, with tightly closing lid.
• Black fiber paper sheets
• Rubber bands
• Paper towels
• Absorbent cotton wicks
• Hand magnifiers
• Labpette digital micro-pipet, capacity 1-20 micro-liter
• Parafilm rolls (for sealing test tubes, flasks, Petri dishes, etc.)
• Eye wash bottles
• Disposable non-toxic particle masks
• Graduate and funnel brushes
• Glass shell and screw-cap vials
34 9
Figure 3. Rectangular plastic boxes and glass jars holding adults for mating
and oviposition. Plastic containers contained unsexed groups of
adults, while the glass jars hold pairs of sexed males and females.
Eggs. Cotton wicks holding eggs were removed from the oviposition sites
(i.e., large plastic boxes and small glass jars in figure 3) and placed in
separate boxes (Figure 4). Cotton wicks were wet with water to avoid
drying. Other eggs were transferred with the camel hair brush and placed on
wet filter papers inside the petri dishes for further studies. New cotton wicks,
saturated with a honey solution, were placed in all containers holding the
adult stages. After 2 to 3 days, larvae from hatched eggs were removed to
separate containers and provided with pieces of sugarcane stems for feeding.
Larvae. Newly hatched larvae on cotton wicks were transferred with a
camel’s hair brush to pieces of sugarcane stems (at least 15 mm in diameter)
(Figure 5). A small hole was made at the end of each piece of sugarcane
stem using a cork borer. One week after feeding, larger larvae were
transferred to larger fresh pieces of sugarcane stems (10 – 40 mm in
diameter; this was based on the size of larvae at different developmental
stages). Last larval instars made cocoons from the fibers inside the sugarcane
stems. When sugarcane was infested with Drosophilla flies, yellow sticky
traps were placed above the rearing containers as a control tool. Also, larvae
were reared individually (Figure 5) in sugarcane stems to avoid cannibalism.
35 0
Figure 4. Egg collection from cotton wicks and filter papers.
Figure 5. Handling and feeding of larvae in sugarcane stems.
35 1
Pupae. After 10-14 days of feeding of last larval instars, the stem pieces of
sugarcane were split open and cocoons were collected. Cocoons were placed
in a plastic containers or metal trays, wet with water as needed, and closed
with lids. Two weeks after collecting the cocoons, they were checked daily
for adult emergence. Adults were collected by hand and placed in plastic
containers (as unsexed groups of adults) or placed individually in glass jars
(as sexed, paired males and females).
Figure 6. Collection of pupae of R. ferrugineus from pieces of sugarcane
stems (a) and plastic containers holding cocoons (b).
Rearing on Artificial Diets
Preparation of Diets
Artificial diets were prepared for mass rearing of R. ferrugineus
because of the unavailability of sugarcane in UAE and this was a limiting
factor in culturing this insect. Diets were also developed to avoid the use of
expensive palm tissues for culture of weevils.
Table 2 lists materials used for the preparation of several artificial
diets. Different diets were evaluated in preliminary tests for larval and adult
biomass gain, survival, and the rate of development Three diets were
selected (Figure 7) and percentage of dry ingredients is listed in Table 3.
Diet A is an mainly an oat diet, Diet B is an oat diet plus palm and coconut
tissue, while diet C is mainly a white bean diet. Ingredients and water (1 - 2
liter of water for diets weighing 500 - 1000 g) were blended for
approximately 5 minutes. All diets included bacto-agar, multi-vitamins,
chemical preservatives. Bacto-agar was dissolved in water and added to
other ingredients. The mixtures of all diets were then autoclaved for 20 min
at 120ºC. Diets were poured in diet stainless-steel round trays or cups while
still warm. All trays and cups were stored at room temperature until
a b
35 2
required. Larvae were placed on diets after total coolness. As with rearing
R. ferrugineus on sugarcane, when artificial diets were infested with
drsophilla flies, yellow sticky traps were placed above the rearing containers
as a control tool. Also, fungal and bacterial contamination may develop if
chemical preservatives were not incorporated in the diets. This may also
occur if high quantity of water was mixed with the dry ingredients of diets.
Table 2. Materials used for the preparation of artificial diet for rearing R.
ferrugineus in the laboratory.
Food Materials
Oats Date palm leaves and fiber sheath
White beans Canned or fresh pineapple
Sugarcane bagasse Soybean flours
Fresh sugarcane stems Coconut fruit pieces
Fresh coconut cake Coconut fibers
Potatoes Vegetable oil
Egg yolk Wesson salt
Vitamin tablets* Brewer's yeast or other brands
Honey
Chemical preservatives
Methyl para-hydroxybenzoate **
Auromycin
4M Potassium hydroxide***
Sorbic acid****
Bacto-agar
___________________________________________________________
* vitamin tablets contained vitamins A, B1, B2, B6, B12, D2, E, K1,
Riboflavin, nictotinamide, and others.
** 14% solution In 95% ethyl alcohol. 5 ml water was added to 95 ml
absolute ethyl alcohol (95% solution) and then 140 g methyl parahydroxybenzoate
was dissolved in 95% ethyl alcohol (amount of 15
ml solution per 1 kg diet was used)
*** 56 g potassium hydroxide in 250 ml distilled water (the amount of use
5 ml solution per 1 /kg of diet was used).
**** 12.5% stock solution In 95% ethyl alcohol. About 125 g sorbic acid
was dissolved in 1 liter of 95% ethyl alcohol (the amount of 15 ml
solution per 1 kg of diet was used) (Rahalkar et al. 1985).
35 3
Table 3. Percentage of dry ingredients of three artificial diets used for
rearing R. ferrugineus.
% Dry Ingredient in
________________________________________
Dry Ingredient Diet A Diet B Diet C
Oats 57 48 15
White Beans - - 65
Palm and coconut tissues - 15 -
Sugar 22 19 10
Molasses 11 10 -
Brewers yeast 9 7 9
Salt 1 1 1
Figure 7. Artificial diets in small plastic trays or cups or in stainless steel
containers for feeding of larvae of various sizes.
Larval Feeding on Artificial Diets
Prior to mass rearing, several artificial diets were formulated and
preliminary evaluated for development of R. ferrugineus. Second instar
larvae were washed in tap water, weighed, and placed into holes made on
artificial diets to facilitate feeding. Larvae were transferred one per diet cup
with a fine camel hair brush (Figure 8). The weight for each larva (n = 8)
was recorded weekly for 6 weeks, and was then compared with its starting
35 4
weight. When larvae reached the end of the last larval instar, they were
placed in the stems of sugarcane for constructing cocoons. Diets were
checked daily for any dead larvae in which they were replaced.
Figure 8. Feeding of larvae on artificial diets.
RESULTS AND DISCUSSION
Previous attempts that were made by several researchers followed
several steps: Sugarcane was a good substitute for coconut for rearing R.
ferrugineus (Rahalkar et al., 1972); sugarcane was later incorporated in
nutrient agar for feeding young larvae and sugarcane stem pieces for feeding
of older larvae (Rananavare et al. 1975). Rahalkar et al. (1978, 1985)
improved the culture of R. ferrugineus by developing an artificial diet
containing sugarcane bagasse (fiber), coconut cake, yeast, sucrose, minerals,
vitamins, and preservatives. Giblin-Davis et al. (1989) cultured R. cruntatus
and R. palmarum on decomposing pineapple. In our study, there were
variations in the quality of developed diets made using different ingredients.
Diet A was mainly made by oats, diet B was made using oats and fibers,
while diet C was made using white beans. Diets A and B were preferred by
young larvae (1st to 3rd larval instars) while diet B was preferred by older
larvae (4th and 5th larval instars).
35 5
The average biomass of larvae at various stages, survival, percentage
of larvae that went into cocoon, and biomass and percentage of emergence of
adults varied greatly between the diets tested: a range of larval biomass after
three weeks of feeding on artificial diets was 0.25 – 2.5 g and after 5 weeks
was 0.5–4.5 g. The weight gained from feeding increased considerably in a
three week period (i.e., larvae with weight ranged from 0.5 to 0.11g
increased 40 to 80X, that is 4.18 to 4.54 g). The weight gained varied with
the type of diet provided to the larvae. The percentage of larval survival
ranged from 10 to 90%. The biomass of emerged adults ranged from 0.8 to 2
g. The percentage of adult emergence ranged from 10 to 80%.
Larvae were fully developed on artificial diets and molted four times
during their development (Figure 9). These larvae failed to construct
cocoons because of the unavailability of fibers (palm or sugarcane) or there
was not enough quantities of high fiber substrates in diet B. All larvae fed by
either of the three artificial diets were transferred to sugarcane stems for
making cocoons. Additional studies are required to search for alternative
food sources to develop diets that R. ferrugineus can fully develop on it
without transferring last larval instar to sugarcane stems for cocoon
construction.
Figure 9. Development of first instar larvae on artificial diet (a), and the
molting of fourth instar larva to the fifth larval stage.
The success of our three artificial diets (Table 3) in providing the necessary
nutritional requirement for molting and development was based on data
recorded ing various life parameters of R. ferrugineus on sugarcane and
artificial diets (a study conducted by the authors, see the next manuscript in
this proceedings). The study on life parameters of R. ferrugineus included
the following: fecundity or the number of eggs per female, egg viability or
the percentage of hatch, larval developmental period, larval biomass, pupal
35 6
period, percentage of larval survival, percentage of adult emergence,
fecundity, fertility, and female:male ratio.
LITERATURE CITED
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35 7
P. Singh and R. F. Moor [eds.]. Handbook of insect rearing. Elsevier,
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