(2) I + x N o^ , aré^shown in Table 6. These results indicated that
there was no significant difference in the percentage of females in- -s , seminated by either irradiated or untreated males. Any percentage less than 1007. was usually due to the females dying in the first 3 days of the experiment before the males began mating attempts.
i TABLE 6. Percentage insemination of Musca autumnalis females in the matings of: (1) normal females (N ¥ )_and ' irradiated males (I o^ ))»» and (2()) irradiated» females -.((I ,+.) and - normal males "(N cf* ) at all dose levels. ( - , • • a - — Females Insemi kated w o ^JDOBJB (krod) N + x I (2) X N o* (1) I + • 0.0 97 99 0.5 97 99 1.0 96. 99 'J ,;l.5 98 97 2.0 98 99 2.5 97 99 5.0 99 100 7.5 98 98 10.0 99 97
a • Mean of 4 replicates, 50 females.'per replicate. 30
' Examination of the spermathecae indicated that the irradiated
male transferred sperm as effectively as did the normal male. Also,,
the" irradiated female mated as readily as the normal female with normal
_.malejsï-—Motility-was observed-in-the sperm found in'the~8pennathecàé b£"~
females mated to irradiated and to normal males. Sperm motility is not •
usually affected until complete sterility is induced (LaChance e£ al.,
. 1967).' However, Terzain and Stah|er•(1958) reported that, 10.0 krad\
induced 1007» sterility without affecting sperm motility in Aedes
aegypti. These experiments showed that male sterility*was not a result
of aspermia, sperm immotility or inability of males to mate, but rather
to the lethal>effects induced in the sperm as a result of irradiation.
Dominant lethal mutations induced by irradiation produce
nuclear changes resulting in the death of the zygote (LaChance et al.,
1967). The nuclear changes induced by radiation and causing the death
of the zygote may be due to the direct, and indirect action of ionizing•
radiation on DNA^synthesis or the result df a base-change or base-
deletion altering the base sequence of a molecule (Pizzarello and
Witcofski, 1967). In formation of the zygote these genetic changes may
be enough to cause lethality. Most dominant lethal effects in'insects
are a failure of the embryo to develop. The frequency of 'dominant '
lethal mutation induced by radiation increases in proportion to the dose
(LaChance, 1967).' . ^
The sensitivity of immature pupaé to irradiation and the
gradual increase in radio-resistance with age has already been des-
cribed for the face fly and is similar to that described for other
dipterous pupae *(Nairs_1962; Bushland and Hopkins, 1953). By the tins
the pharate adult is visible in the puparium (3 to 4 days in the face 31
fly) cell division and differentiation has' been replaced by cell en-
largement and'decreased radiosensitivity. Cell division and replace-
ment still occurs'in the gonads and these are easily damaged by
radiation. Consequently the sterilizing dose and 'maximum tolerated "J
dose differ so enormously. This difference has been demonstrated for
other diptera as well (Bushland and Streeter, 1971; and Davis et _al.,
1959).. Auerbach and Slizynski (1956) found that spermatids at the
point of transformation into spermatozoa were the most sensitive germ
cells to the mûtagenic action of: X-rays in Drosophi-la and mice. tSperma-
-i ^—*
togonia were the least sensitive. Conversely primary spermatocytes and
«permatogonia are more* sensitive to the lethal action of radiation in '
the screw-worm fly Cochliomyia hominivorax (Ct/querel) , (Riemann, 1967)
and house flies1(Riemann and Thorson, 1969).
These experimental results alsduindicated that female ste-
rility was mainly,a result of loss of egg production and to some extent
to the lethal effects of radiation on the egg at doses of radiation
less than 1.0 krad. Inability on the part of the female to mate was
not a factor. Egg. production by insects is largely dependent upon the ' /
•differentiation of oocytes from oogonia and the proper function of the
nutritive cells. Severe damage to the oogonia, and at certain
the nutritive cells, can result in permanent loss of egg product i/bn.
JLOSS of egg production has been observed repeatedly aftW
treatment of female insects with ionizing radiation (Bushland and
Hopkins, 1951 and 1953; Annan, 1955; Henneberry, 1963; Gregory, 1969). Loss of egg production in the female7facJ e 'fly was thought to be due to injury to the oogonia but this can only be ((hown histologically. The
appearance of-nurse cells in the first egg chamber of the screw-worm 1 v "* 32
fly pupa has been observed 1 day prior to eclosion (LaChance and Bruns,
1963) and if this is the same for the face fly, then irradiation would
damage the oogonia and not the nurse cells. The ovary of the female
— face-fly is more-sens it ive-toTadiation-itljury-than- the teatis of the ";
male. A similar sensitivity was reported for thevstabie fly (Offori,
1970) but the reverse was true with the screw-worm fly (Bushland and
Hopkins, 1953). / *
(e) Adult Survival >
Longevity of the irradiated insect is as important to the
sterilè-male technique as the 'actual sterilizing dose. The sterilized,
insect must surv.ive long enough to compete against native insects for
mates to play its part in the release program. Therefore, daily sur- i vlval of the irradiated face flies was recorded as part of the ste-
. rility experiment/, and tabulated as the percentage survivals at 7, 14, / - and 21 days. Percentage survival of irradiated males and females are
shown in Tables 7 and 8 respectively.
/ Treatment of the data for the percentage male survival
(Table 7) "by analysis of variance produced a high degree of signific-
ance for the treatment and time factors. Treatment-time interaction
was.not significant. At 7 and 14 days, there was little difference,
between the survivals of irradiated and untreated males, however there
was greater difference at 21 days.
I 33
TABLE 7. Percentage survival of male -Musca 'autumnalts adults at 7, 14, and 21 days for different radiation doses.
• a 'Mâle survival (%) at; Dose (krad) 7 days 14 days 21 days
0.0 98 93 84
0.5 98 96 82
1.0 95 "86 61
1.5* 93 76 38
2.0 95 77' 37'
2.5 95 74 45
5.0 94 71 40
7.5 97 . 70 38
10.0 92 " 74 53
Mean of 4 replicates, 50 males each replicate.
Statistical significance for: Treatment f ^0.01
Time „ P <0.01
Treatment-Time Interaction P ^>0.01
Variance analysis of the data for the percentage female / "V survival (Table- 8) showed no significant difference for the treatment 0 I effect (a small degree of significance at P = 0.05), but a high degree of significance for the time effect^ Treatment-time interaction was not significant. 34
TABLE 8. Percentage survival of female Musca autumnsills adults at 7, 14, and 21 days for different radiatlron doses. • \ a Female survival (%) at: Dose (krad) 7 days 14 days 21 days.
0.0 92 81 58
0.5 98 82 61
1.0 V 96 75 42
1.5 93 73 39
2.0 94 * 77/ 44.' 1 - 2.5 93 72 39
5.0 " 93 77 •*•- 46
7.5 95 80 48 -
10.0 93 71 • 41
•/ a Mean of 4 replicates, 50 females, each..
Statistical significance for: ,
, . . Treatment P ^ 0.01
. Time P < 0.01
Treatment-Time Interaction P ^0.01
The results indicated that males survive significantly better than females between 14 and 21 days. This was probably due to the added stress of oviposition in the females. Decreased longevity has been reported in screw-worm flies (Bushland and Hopkins, 1953) at the sterilizing dose of irradiation whereas an increase in longevity was reported for male and female codling moths (White et al., 1972). No difference in adult longevity' at the sterilising dose was recorded for 35 the stable fly (pffori, 1970) or horn fly, Haematobia irritana (L.)
'(Lewis and Eddy, 1964). .
4. Effects of Irradiation on a Native Colony of Face FJLies.
An insect colony which_is reared under laboratory conditions^ different to those found in nature, may be a process of selection, result in a strain very different from-the parental stock. This selection fox laboratory conditions'^ may^.resul t in behavioral changes which potentiall' ^ y- affec• t ,the "abilitk- \y to survive and compete satis- factorily with the wild flies in the^hatural environment. A colony of
/ '• •- . field-qollected face flies, referred to as the native colony, had been / " ' ' ' S '<.n rearedI under laboratory conditions through eight generations. At this time no\rEfferencV e was noticed between the duration of this colony's life cycle and that of the* USDA laboratory colony.
Experiments were conducted with this native colony (eighth generation) to determine the effects of radiafion on the adult emer- gence, fertility, and male survival'in order 'to be able to .compare the results with those obtained for the USDA colony. Pupae, 5' days old, were Irradiated at 2.5 and 5.0 krad and the emerging adults placed in small cages in the following combinations of 50 irradiated males (I /) and 50 normal males (N o*' ) with 50 normal females (N % ) each:- „
2.5 krad • 5.0 tarad 50 1^ + 50 N % 50 I o^ + 50 N °-
Control 5ONo^ + 50 N/£ 50 N
Daily survival for males was recorded until the completion of the
• • < test at 21 days. Eggs were ,4-qJLlected and counted on days 5 to 21, /and
_ ' j the numbers' of the resulting pupae and adults were tabulated. Three
replicates were completed for- each dose level. , , • ;
t #*• >n , ,
(a) " Male^Fertility • • ' \ , -
.The mean number of eggs laid, by normal females mated to
i ' . ) • irradiated males and the resulting pupae and adults are shown in Table
' ' '..'•, ,,;% ' 9. Adult emergence from irradiated pupae are also shown. '_
TABLE 9. Eggs laid, pupae formed, and emergence of Muaca autumnalis^-r <* is» aduLts from the mating of normal, native colony females to native colony males, irradiated as 5-day-old pupae at different radiation doses. „$
Y Mean ' tJ Dose Adult emergence from . no. of a b, Adult b (krad) irradiated pupae (%) eggs laid Pupation (%) emergence (7.)
0. 0* 87. 3 2 ,775 67. 3 + 2. 5 52.6
2. 5 89. 2 3 ,502 > .0 0
5. 0 86. 7 2 ,474 0
Mean of 3 replicates, 50 females per replicate. A 1.0% difference
represents 90 eggs approx. b \. - \ • % determined fronrno. eggs laid.
The results indicated that a dose of 2.5 krad sterilized
males of the native colony completely. There was no significant
difference between the mean percentage adult emergence from irradiated
and untreated native colony pupae. The. niéan percentage emergence of
at •'•/. L
37
adults from irradiated pupae of the native colony was .similar to that
fronl irradiated pupae of the USDA colony. No comparison was drawn be-
tween the number 'of eggs laid by.native colony females and USDA colony
-feinaleB~since~the~èxpëtfiinent8"we"r^c^n*dûcltëd""at" different times of the
year. However, native colony females di,d'not oviposit as readily
through cheesecloth as did USDA "females. * ' VL
(b) Male Survival *
Data for the percentage survival'of irradiated and untreated
male native colony flies are shown in Table 10. Treatment, of the data
by analysis of variance showed a/significance for, the treatment and
time effects, but, no significance for the treatment-time interaction. ! These results were similar to"those obtained for'USDA'males (Table 7);/
TABLE 10. Percentage* survival of native «colony male . 7-', Muaca autumnal is adults, at 7, 14, and 21 , /<• days for different.radiation doses.
i , - ' - à m Male survival (%) at • Dose (krad) 7 days 14 days ,'[ 21 days
1 i 0.0 92 84 76 ! 2.5 89 • 55 35
• 5.0 . 87 61 -> . 37
Mean of 3 replicates, 50 females per replicate.
Statistical significance for: ' --'- \ / Treatment. p <£ o.oi Time. ! ' ' P<[p.01y
Treatment-Time Interaction P ^-0.01 • » ^ i
1 .> 1 38
5. Degree of Permanence of Male Sterility.
Previous experiments demonstrated that male face flies were
sterilized completely by a dose of radiation of 2.5 krad administered
• ' « / to the 5-day-dld pupae. However, these_ results_did__not__shaw_whether
the induced sterility _was permanent; or whether there was a return of
fertility after a period of time. Therefore, experiments were con- %
ducted to determine whether or not there was any return'of fertility in
males sterilized at' 2.5 krad' and subjected to multiple .matings over a
period of 22 days. . ' • ' ,. , * ,
t ' -.V Pupae were irradiated at 2'. 5 krad and the emerging adults r i * , . / i placed in two small edges in the following combinations of 50 irra-, • "-":-'
diated males (I o^ ) and 50 normal males (N o^ ), each with 50 normal
females (N % ). . - f
Control 50N/ + 50 N % Cage 1
2.5 krad . 50 I / + 50 N ? Cage 2
Eggs were collected and counted on days 5, 6,- and 7, and the
resulting pupae recorded. After oviposition on day 7, the female flies
in both cages were replaced by virgin female flies 4 to 5 days old
equal to the number of /male flies'remaining alive in each cage. Eggs
were collected on days/8, 9, and 10, counted, and the resulting pupae
-recorded. After oviposition on day 10, female flies were replaced
'with virgin flies as before. This procedure was repeated'for each
successive 3-day period until males had the opportunity to mate with
6 separate groups of virgin females. All female flies were dissected
to determine whether insemination had occurred at each mating. Male r 39
survival was recorded at 7, 14, and 21 days for each of 3' replicate
completed.
.(a) —Percentage-Pupation" -•*>—
• The results of these tests are shown in Table 11. The mean
number of eggs/ deposited per 3-day period was 457/ for females mated to \
^untreated males and 368 for females mated to irradiated males. The
difference was probably due to a better suryival of the untreated males
compared to irradiated males at the end of each 3-day period. This'
v^ould result in a greater number of virgin females being introduced to
the normal males for each 3-day period.. The difference in the number
of eggs produced was most evident at the fourth, fifth, and sixth .
mating. The smaller number of eggs laid by both groups in the first
period was probably due to experimental error since peak- oviposition
usually occurred on dayV5^ dur ing/previous experiments in agreement with
the findings of Miller and Treece (1968). l . . ?
TABLE 11. Eggs.^aid and pupae formed from the mating of normal, virgin , J Musca autumnal is females (N ,$ ) with untreated maples (N
Mean No. of Eggs from; Pupation Resulting (%)• (1) (2) Period Matings (days) N + x N N + x N- N . 1 5-7 228 219 72.9 + 5.8 0 2 8-10 592 .565 49.3 +7.0 0 • 3 '*'. 11-13 567 547 66.4 + 3.3 0.9
4 14-16 562 481 69.0 + 3.2 0 •**> 5 s 17-19 385 251 71.5 +4.7 0, 20-22 407 149 42.9 + 9.7 0
Mean of 3 replicates. 7T
40
The'mean pupation for untreated flies for six matings was
60.9% and for the irradiated flies 0.07». ' There was a suddert drop in'
percentage pupation (Tablé 11) between days 8 and 10 (Becond mating) •^ - _£or_the__females _mated_to- untreated • males.—Thi8~was~followed by a
gradual increase in the percentage pupation'through days 17 to,19 '
when another sudden drop occurred. These decreases in percentage .*»' pupation can not be explained satisfactorily. Since the number of
females inseminated (Table 12) during the,, same period did not indicate
a_similar trend, sperm depletion in the males can be ruled out. Ex-
perimental error was not? thought to be a factor since results from
all 3 replicates indicated, a drop at both these points. ,
Only a very small percentage pupation (0/9%) was found with
females mated to irradiated males (Table 11). This occurred in 2 of- \
3 replicates'between days 11 and 13. Since one untreated male had
been found with the virgin females before their addition to the ex-
perimental males, this,.apparent return to fertility between days 11 •
and 13 was probably due to this"male matirig with a few females. These
results indicated that the sterility produced by irradiation of 5-day- 13 old pupae with 2.5 krad was permanent and that no recovery of fertility c occurred in males.through 22 days.
(b) Percentage of Females Inseminated
^ Henneberry and McGovérri (1963) using Droa'ophila melanomas ter
and Tsiropolous and Tzanakakis (1970) using the olive fly Dacus oleae
(Gmelin) reported that the store of sperm of irradiated males was ex-
hausted after several matings. Single mating tests completed on the
face fly had not indicated how long the supply of sperm from irradiated * /
41
males, would last. This was determined by examination' of all females '
from multiple,matings of the male permanence test for insemination.
Insemination depended on finding sperm in one or
; spermathecae.. Data on the number of females inseminated by irradiated
and normal males during the six/ separate 3-day periods are shown in J
Table 12. V,
TABLE 12. Percentage insemination of normal, virgin Musca autumnalis females by: (1) untreated males (N1 cf% ), and (2) irra- diated males•(I d* ) for' six separate 3-day periods.
Females inseminated (?<•) by males during days: Dose (krad) 5-7 8-10 11-13 14-16 17-19 ' 20-22
0.0 (N/nN'?) 83 82 77 82 78 75
2.5 (I d* x N % ) 81 71 45 20 0 15
a • Mean of 3 replicates, 50 males each.
' No difference was recorded in the number of females ih-
seminated by untreated males during^ the experiment. Usually all three
spermathecae contained sperm.'.. This indicated,, that the males' supply of. , • ' f sperm was- never depleted. However-, fewer females were inseminated by
irradiated males (Table 12). By days 14 to 16, usually only one sperma- fi . \ '
theca contained.sperm. ' No sperm was found in the spermathecae of
females for the period 17. to 19 days. The presence of sperm in the
spermathecae of a' few females (1 out of 5, 1 out of 15, and 5 out of
20) during the period 20.to 22 days was thought to be due to, mating by
flightless males which were unable to mate before the death of the more
aggressive males. ,
A 42
•»•>. The permanence test indicated that irradiated males were able to transfer sperm through four matings, whereas normal males trans- ferred sperm through six matings. A few sperm could be found in the spermathecae of females mated-by-irradiated males'up to" "thefourth r " & mating. By the fifth mating the spermathecae contained flebris of dead
'-.ri ^l ,,%.'"$* , cells, probably primary spermatogonia and spermatocytes. As mentioned '
•previously, primary spermatocytes and primary spermatogonia are easily killed by
1967) and house flies (Riemann and Thorson, 1969). Therefore,, after a few matings the male fly becomes aspermic. This indicated that re- covery had not occurred at thé sterilizing dose (2.5 krad). This was also shown histologically (Ritcey, personal communication).
(c) Male Survival . * ,1 ,.
Data for the percentage survival of irradiated and untreated males during, the sterility permanence test are shown in Table 13. No, significant difference was found between the percentage survival of irradiated males and untreated males at 7 and 14 days. However, a significant difference was apparent at 21 days. The results indicate that the males.survived significantly better under the conditions of single matings (Table 7) than the condition of multiple matings (Table
13) which is probably the, situation found in nature.
Longevity is inaccurate as an indicator of sexual vigour as some inactive males'•lived as long as sexually aggressive males
(Çaumhoyer, 1965). There was some suggestion of.an inaccuracy using longevity as an indicator of sexual vigour with the face fly.v1 During"' the permanence test, females dissected after mating with males for the
t 'i if
v
• ••! '
' "•% ' ' ' ' -'r v *:> ï . V , ,,/j,,<;< fifth time showed no sperm in the spermathecae. However, at the sixth „,/»',;'s.t"•'*'' " mating, some spermathecae contained sperm and it was thought that some •
'v^"-'-* ' less aggressive males had'jma_ted__with_these_females,—being-given-the — _ .——
«•J/;*;)}chance after the more aggressive males had died after the fifth mating. * ft
• -- • -t. TABLE 13. Percentage survival of untreated*and irradiated * male Musca autumnalis adults at 7, 14,.and 21 days during Che male sterility permanence testl J o. a ' • •* Survival of males (%) at: j Dose (krad) 7 days . 14 days 21 days f 0.0 84 . 76 55 / V-dT" 2"5 89 64 17'
a / <^> • ' Mean of.3 replicates, 50 males each replicate. fy
-
. 6. Mating Habits of the Female Face Fly. x\ i?
The female face fly has been reported to mate only once _ (Teskey, 1969; Killough and McClennan, 1969). Those attempting-to mate •
again had not received a -fuLl complement' of sperm at the first mating
(Killough and McClennan, 1969). Wang (1964) and Lodha jet al., (L970)
- reported repeated matings by female face flies. However, Lodha et al.,
(1970) also stated that one mating was sufficient to sustain normal,
' egg production through three gonadotrophic cycles. If the sperm
•< transferred by sterilized male flies was not-as motile as that pro-
duced by-normal males It would be important'to determine whether or v i - - not the females mated more than once. In females that mate more than
.) > 44
once, sperm immotility would permit the production ôf fertile eggs'With
subsequent matings. 'Also, preferential usage of stored sperm has been
reported for^emale Glossina austeni Newst. (Curtis, 1968). It was
-decided,- •therefore-,~to™determine~if~the"femaré"fàcë™~f ly"mâ~të~d once or-
several times^r^Th^r^T e sterile-male technique provided a good' method fô*K - ' *'%£*£. . ' . ' •• ' ) V this determination,, Two experiments were conducted using males stery-
lized by 5.0 krad-of radiation. ' - In the first test, 25 untreated males (N d* ) and 25 irra-
diated males (I df ) were placed with 25 normal females (N + ) each, in
small cages, in the following combinations:-
Cage 1 25 I d* + 25 N ?
Cage 2 25 I (/ + 25 I ? •• , . ,
Cage 3 25 iV + 25 N ?
Cage 4 25 N o^ + , 25 N Î •
Cages 2, 3, and 4 served as controls. On day 10 of the 'test, the irra-
'diated males 'in Cage 1 were replaced with normal 5-day-old males equal z to "the number- of normal females left alive. Eggs were collected daily
from allNcages beginning on day 5 until termination of the experiment
on day 21. Eggs were counted and the resulting percentage pupation re-
corded as before. Three replicates of this test were conducted.
. ;In this experiment, the test females (Cage 1) produced eggs
resulting irTlfpercentage pupation of 0.0% and 0.3% before and after
introduction of normal males, respectively. The females of Cage 2
produced no eggs, as would be expected, and; no pupation«resulted from
the eggs oviposited by the females of Cage 3 (Tables 4 and 5). Eggs
-produced by the normal females[of Cage 4 resulted in a 70.3% d
i \ 45"
pupation. • , . ,
In a second test, sexes were kept separate for 5 days, then
placed together in small_dages.. as..,£ollpws: —~ -T —"; —
Cage 1 50 N ? + 50 1 / V:
' '* ' fl'"1 '* Cage 2 50^ N + + ,0' 50 N o< fl -
Eggs were collected on the next day (day 6), at which time the male
flies were replaced with male flies of the complementary test. "**
Example :
Cage 1 N + + 50 I eK before 6th day
/ 50 -N, °- + \50 N o? after 6th day /
• ' •' Eggs-were then collected for a further 5 days and all per-
centage pupations recorded. Three replicates of this test were com-
•>• pleted. _ . ' •
In this test, females mated with untreated males and then
caged with irradiated males produced eggs resulting in 63.6 ±- 4.1%
and 69.7 + 2.9% pupation whereas females mated with irradiated males
and then caged with untreated males resulted in 0.07. and 0.3 + 0^2%
pupation. • v
These results showed that the female face fly received a full
complement of sperm in a 24-hour period and probably mates only once if
x complète sperm.transfer occurs in a single mating. This is "in agree-
t i ment with the .findings of Killough and McClehnan (1969). Multiple
mating attempts probably do occur if the female has not been success- "' fully inseminated at the first copulation, which would account for the
reports by Wang (1964) and Lodha et al., (1970) of several matings in 46
the female face fly. ' '»•<*» •
Studies by Riemànn £t 111. , (1967) on the house fly indicate
that sperm transfer" is not necessary to initiate a single mating res-
™ponse"~ift~femalesT These"7mthôFs"~ândRÏëmanïr and^Thoraon (1969b) re-
ported that stimulation to oviposit and loss of. receptivity to males
by. female house flies, Musca domestica (L.) are caused by the presence
of the accessory material produced in the male ejaculatory ducts. The
male face .fly, like the male house fly has no.accessory glands. Since
the irradiated male face fly can transfer sperm for only four to five
matings, the transfer, of ejaculat'ory duct fluid preventing loss of f e- "
male Veceptivity/toould be an advantage after the fifth mazing in a
stenile-male release. However, the presence of accessory"
from the ejaculatory ducts and it» effect on female face fly recept-
ivity has, not been investigated.
7. Effect of Irradiating Pupae Under Anoxic Conditions.
Ionizing radiation produces mutations in the cell nucleus by • ' c \
both direct and indirect effects. - The formation of free radicals by
the indirect effect and their action on the genes is.probably as im-
portant as the direct effect on the genes. Since a large part of the
cell is made up of water, most of the indirect effect will be the
action of radiation on the water molecule with production of free
radicals from which chain reactions may start (Pizzarello and Witcofski,
1967). Oxygen enhances the effects of radiation by Interacting with'
radiation-produced free radicals to produce auto-oxidatlve chain re-
actions and promote the formation of hydrogen peroxide. Changing the"
composition of a gas mixture will effect a change in the oxygen tension .47 at the tissue or cellular level. This will then af^ct the interaction of oxygen and the free radicals and produce a protective effect during irradiation. The fact that reducedJ3xy_gejti__tension_decreases...damage frcfm ionizing radia,tion has been known for some time (Thoday and Read,
1947). Both nitrogen and carbon-dioxide have been shown to have a 1 protective action on pupae during irradiation (Baumhover, 1963;
Smittle, 1967"; and Hooper, 1971a). Therefore, face fly pupae were i irradiated in nitrogen to determine the effects of anoxic irradiation.
/ Pupae, 5 days old, were irradiated at 2.5 and 5.0 krad in a nitrogen atmosphere.* Control pupae were not treated,with-nitrogen.
Pure nitrogen (less than 2.0% oxygen) was circulated through the lid of.the vial containing the pupae for 15.'minutes. The lid was then sealed with tape, the pupae irradiated and immediately removed to a clean vial. Upon emergence, adults were placed in small cages in ' combinations of 50 irradiated males (I é1 ) and 50 normal males (N (? ] each with 50 normal female,s (N °- ):-
.^2.5 krad 5 .0 krad o 50 I / + 50 N 50 I o* N i o Control 50N/ + 50 N 50 N c? +* 50 .N
Male "survival was recorded daily unt^l the termination of the experiment at 21 days. Eggs were collected and counted each day from day 5 to 21 and the resulting percentage pupation and.adult emergence recorded for
3 replicates. ' .
(a) Adult Emergence and Male Fertility
The adult emergence from irradiated pupae, the mean number of 48
eggs laid and their resulting percentage pupation and adult emergence
are shown in Table.14. The results indicated that there was no sig- \ ntficant difference between the mean percentage emergence from un- ,
-treated pupae* and pùpaë'irradlatedàC 2.5 krlîd and 5.Ô kràd under
anoxic cqnditions. Comparison of the adult "emergence from pupae irra-
diated at 2.5 and 5.0 krad in air and anoxia showed no significant
difference between them. Under the anoxic conditions used,, the irra-
diation dose had to be doubled to achieve the same degree of sterility
as pupae irradiated in air. This is in agreement with the results
reported .by Hooper (1971a).
Dose ' Emergence from Mean no. Adult (krad) irradiated pupae (%) eggs'laid Pupation (%) emergence (%)
0.0 92.4 3,544 • 68.8 + 1.6 58.7 + 2.5
2.5 92.9 3,999 |1.9 + 0.9 1.5 + 0.8
5.0 93.4 3,087 0.2 + 0.1 0.1
Mean of 3 replicates, 50 females per replicate.
% determined from no. eggs laid.
The protective action of decreased oxygen tension could be
important in any sterile-male release program. ' Storage of large
numbers of pupae prior to irradiation could reoult in anoxic conditions
produced by carbon dioxide released, particularly aa pupal sstaboliom
is increasing at the tints of irradiation of the face fly (Guerrâ and
1 < ' Cochran, 1970). Irradiation of the"pupae under these conditions might
result in only partially sterilized adults on release.. Therefore,
irradiation pf large numbers' of pupae should be_cjarr_ied__out_min_,well=——
aerated containers to eliminate this hazard.*
' ' t
' (b) Male Survival. - ' ';
Data for the percentage survival of untreated males and males
irradiated at 2.5 krad' and 5.0 krad in nitrogen are shown in Table 15.
'There was a significant difference,in the treatment and time effects,
bût no significance for the treatment-time interaction. A comparison
. between mean percentage survival for males irradiated in nitrogen and
jnales irradiated, in air showed no significant difference, at 7, 14, and
21 days, at the 2.5 krad dose or the 5.0 krad dosé, v
i5. Percentage survival of untreated male Kusca autumnalis adults and males irradiated under anosic conditions at 7, 14, and 21 days. -
Ï a • Male survival (%) at: Dose (krad) 7 days 14 days 21 days
0.0 96 90, - 79 2.5 •95 75 38 • 5.0 89 57 29 • 'a. 1 /a ' * ' ' **. / Mean of 3 replicates, 50 males per replicacfe.
l Statistical/Qignificonce for: Treatxcant , P 0.01 V Tims ~^ P 0.01 J i Treatment-Time Interaction P 0.01 i .-
50 ' ! 8. Competitive Studies on the Sterilized Male Face Fly.
Mating competitiveness refers to the ability of the sterile
males to compete with the fertile males for mates. In nature, it, is
. probably-tihe-sum- of— dispersal— ability" mat ing"ag^ë88"lvenësi^",attract- f ion to sex pheromorie and the' number of potential matings. Apart from 0 •J» . : the induction of sterility, it is important that the irradiated male - in a sterile-male release, program be as sexually competitive as the'
, ii wild maiei Competitiveness has ( been evaluated in three ways: (1) by,
comparing the number of observed matings; (2)'comparing the number of
females mated or inseminated, and (3) by ratio tests. The first
method has not been employed in this study.
.(a) Number of Females Inseminated '
An experiment was designed to determine the number of normal
les -which could be inseminated by one normal male, or one irra- .. ^ • s i '«' ,. male within a 24-hour period. Males and females were kept
.' separate until day 4 after emergence when one male was placed with ten ':;, • . . * -,
• normal.females in a 1-quart cardboard container, with a screen top.
* e * Females were dissected 24 "hours later to determine if they had been IS. 'inseminated. ' '. Fifty replicateV s 'of this desig- 'n were. , completed •>fo r each — of four treatment levels — (1) untreated, (2) 2.5 krad, (3) 5.0 krad
in air, and ' (4) 5\0 krad in nitrogen. I» '
A frequency distribution of females inseminated by each male --
for these four treatment levels is shown in Figure 7. The mean number '-v
of females inseminated, the range, and the percentage of males mating • the males irradiated at 5.0 krad in air inseminated significantly; fewer with five or more females is shoun in Table 16. Results indicated that '• -'".' •J'K>'(V\''I[,{
/ />,'
/ , ;/ ,
Figure 7. Frequency distribution of the number of Musca autumnalis
females, inseminated by male face flies, untreated and irradiated at,2.5 and 5.0~krad in. air and 5.0 kra'd in/ nitrogen.
i'
.' £;>•
y • x • .. 51 I /
l' y
_ o • ° <. a 1 i - o' - o u C- o i DDDC DC - . • DDDC --1. • DDC • DDQCOODDC: DDDDDDE L- ^ • DDC ODDDDDDDDC|^ * s w DPDDDDC D'DDDC k • ODDDC ODDC / / ""/ Q D ODDC lad :rl '• / • -o/ Lo - / / - A J» 7, o a» rL
/ / .
ESd / / IS. ta. O c - o t a a . Ê2 . , DC a tsa ^ / a oc o DDDDDD£>^ 1 ; OD •° I 2 ' - eDDDDDD[-e> •" DDDDC » CD & * 1 DC /S DCIDDDtDDDDC i f' / .•ODDDDi ''• ÏDDDD- « •r DDdDDDILea oDDq-- ; • , • • - one J, o & k r •
i
i
i /• I, 52 \ • females than either the normal males or males irradiated at 2.5 krad or
5.0* krad under anoxic conditions. There was no difference in the numbers of females inseminatedJby_normal.males, or-males- irradiated~at~~~
2.5 krad. . ' '
TABLE 16. Mean number,of Musca autumnalis females inseminated, the range, and the percentage of males inseminating 5 or more females for the dose levels: (1) untreated, (2)1 2.5* krad in air, (3) 5.0 krad in air, and / (4) 5.0 krad in anoxia.
Dose \ Mean no. a Males inseminating (krad) females inseminated Range 5 or more females (7<.)
0.0 0-9 60.0
2.5 1-8 '52.0
5.0 0-7 24.0
5.0 (Anoxia) 1-8 34.0
Mean of 50 replicates, 10 females per replicate. ' b, c • ,' \ ' ' Means characterized by same letter are not significantly dtf-£erent at P= 0.05 level of probability.
' -The results (Table 16) indicated that the male face fly irra- diated at 2.5 krad was as virile as, the normal male.. Males irradiated at 5.0 krad were significantly less vy.irile.' Lodha el: ail., (1970) re- \ ported that the normal male face fly may mate with as many as eight females, .with an average Of four, in agreement with the results reported here. .Normal male screw-worm-flies mated with as many as six females
(Bushland and Hopkins, 1951). ,No difference in the percentage of females mated by sterile or. untreated males was recorded for stable 53
flies' (Offori, 19(70)" but irradiated males of Anopheles pharoensis were
reported to mate with,more females ,than did normal males (Tantawy v
.et al, , 1967) .
(b) Ratio Tests
Egg-hatch data, obtained/from the mating of normal females
with various ratios of irradiated males to normal males, are probably
the best estimate of the confpetitiveness of the irradiated male. • Egg-
hatch will be the net result of•the interplay of factor» such as
copulatory efficiency, sperm transfer, and sperm competitiveness.
' Basically, any factor affecting competitiveness will be refle'cted in
the egg hatch or pupal formation which was the index used to test j
competitiveness in.the face fly.
Ratio tests were used to determine the competitiveness of
male flies irradiated at 2.5 and 5.0 krad in air, and 5.0 krad under
anoxic conditions. Following irradiation of the 5-day-old pupae, the
resulting adults.were placed in three small cages, so that each con-' o ' '
tained 50 normal females (N + ) with different ratios of irradiated
males (I cr ) to normal (N 1:1:1, the numbers of flies in the three cages would be:-
* , H* * Cage 1 50 :: 50 : 50
Cage 2 50 •: 50 t , 50
Cage 3 Control 0 : 100 : 50 With th,i8 experimental design, the effect of the total male population1 ' ' ' ' / . !>on the survival, oviposit ion,,and fertility of the caged flies could be
— determined.-- ~ ~~ "~ , , '
, • -\ •
P% At 2,5'krad, the following ratios of irradiated males :
normal males'':' normal females were tested1 1:0:1, 0:1:1, 1:1:1, 2:1:L,
5:1:1, and 1:2:1. In addition, flies in a 5:1:1 ratio were tested in large cages. At 5.0 krad, ratios of 1:1:1, and 2:1:1, and 5:1:1
' • " *
were tested, the latter in square cages. At 5.0-1 krad-under anoxia,
only the ratios 1:1:1 and,2:1:1 were completed. Each experiment was replicated 3 times at each ratio used. Eggs were collected and counted Y on.days 5, 7, 9, and 11, and the. resulting pupae recorded» as the
observed percentage pupation. The experiment''terminated on day 11.
- Competitiveness8 was estimated by a comparison between the
observed percentage pupation and the expected percentage pupation
determined from the controls for each ratio test. .The results obtained
from 3 replicates at each ratio for the 2.5 krad dose in air are shown,
in Table 17 and the results obtained for the 5.0 krad dose in air and
under anoxia are shown in Table 18.
The results (Table 17) indicated that the irradiated males
were as competitive as normal males, at* the ratios 1:1:1, 2:1:1,^5:l:'l,
and 1:2:1 but less competitive at the 5:1:1 ratio in large cages. In-
creasing male populations (2:1:1 and 5:1:1 ratios) affected,the number
of eggs laid by females but not their fertility, as indicated'by the
percentage pupation.for the controls at each ratio. There was a
significant difference between the percentage survival of irradiated
male populations and the untreated male populations (control) at 2.5
kfad using the ratios 2:1:1 and 5:1:1 in the large cage tests. There , 55 was no, sigrfif icant 'difference at the other ratios.
TABLE 17. » Mean number of eggs laid and the resulting pupation _i _ (ob^erye^d jind_.expected)._ f rog-the-mat ings -of - normal — „.__ r female Musca autumnal is (N + ) with different ratios of normal males (N dr, ) and males irradiated at 2.5 krad (I d* ). .
Ratio: Mean no. * e888 Per Observed Expected b, Id* : N d* o replicate pupation (?<,) pupation *(7.) * \ 1:0:1 1,573 0.0
0:1:1C - 1,550 65.3 +' 8.8 ,
1:1:1 1,036 22.9 + 1.5 27.9 V 0:2:1° - 929 , 55.7 + 2.9
2:1:1 878 24.9 + 2.5 20.2
0:3:1 1,109 60.8 + 3.3
X 412 8.1 ± 2-5 0:6:1° 355 67.0 ± 2-5
• 1:2*1 1,041 43.4 + 2,2 41.6 e 0:3:1 , 970 62.4 + '2.2 -
5sl:l 718 25.5 +' 1.4 11.7 0:6:lC 705 70.4 + 3.3
Mean of 3 replicates, 50 females per replicate.
Determined' from controls.
C t • Controls for each ratio test. d Large cage test.
r rx
56
The results (Table 18) indicated that males irradiated at
5.0 krad were less competitive than normal males'at all ratios tested.
-There wa8-a"8ignificant"^if£ërencë" in ^percentage survival between •
males'irradiated at 5.0 krad and untreated males at the ratios 1:1:1 V
and 2:1:1, but no difference at the 5;1:1 ratio in'thëcvfikrge cage test.
TABLE 18.' Mean number of eggs laid and the resulting pupation (observed and expected) from matings of- normal \ Musca- autumnalia females (N ? ) with normal males (N o*7 ) and males irradiated (I o^ ) at 5.0 krad/in air and 5.0 krad under anoxic conditions. v Ratio: Mean no. eggs per a Observed Expected : N d* : N replicate puypation (%) pupation (7«)
1:1:1 ' 1,666 42.4 + 1 | 7.1 32.6 r 0:2:lC , 1,552 2.9 t * 65.1
2:1:1 1,383 37.2 + 6.3 26.4 " 0:3:lC 1,2.09 79.2 + 2.3
5:1:1 1,072 23.5 + •6.5 12.1 0:6:1° 702 72.4 + 6.3
In Nitrogen: • 1:1:1 1,234 37.4 37.1 0:2:1 1,230 74.1 + 5.2 —
2:1:1 1,230 21.9 + 2.4 18.0
0:3:1° •1,013 54.1 + 6.8 Mean of 3 replicates, 50 females per replicate. 7 Determined from controls.
Controls for each ratio test. '
Large cage, te'st.
I 57
Males " Irradiated at 5.0 krad under anoxia were as. normal males at both ratios tested (Table 18). Untreated males sur-
vived significantly better than males irradiated under anoxia at both
the 1:1:1 and 2:1:1 ratios. The difference between the percentage
pupation obtained for the irradiated flies and their'controls for each '
ratio was hughly significant, (Chi-square P = 0.01) at al^ levels of
radiation.. . , • i ' -
During the competitive tests at 5.0 krad in air using the
5:1x1 ratio in large cages, some of the ma^Le flies became infected with
Entomophthora muscae (Cohn) Fresenius. Typical infections of male and
female flies are shown in Figure 8, "This adversely6 affected the sur-
vival rates of the irradiated males in the second and third replicates.
It is«almost certain^Chat only irradiated flies were 'infected as, no
.deaths, either male or female were recorded in untreated flies. It is
not known to what extent the infection affected the competitiveness,
but the results indicated that the irradiated flies were .not as com-
petitive as normal males,"a result perhaps influenced by the' infection.
The ratio tests showed that competitiveness' decreased with
increasing dose (from'2.5 to 5.0 krad) in the face fly, a result
similar to that noted for Ceratitis capitata (Hooper, 1971b and 1972).
As has been mentioned, irradiation of face flies in nitrogen de- creased the competitiveness less at the higher dose (5.0 krad), a^e-
sult similar to that reported by Hooper (l'971a). However, the male, face fly, irradiated at 2.5 krad was as competitive as the untreated males in small cages. If significant differences in competitiveness values of males are to "be demonstrated, extensive replication Is > necessary. This is particularly important in larger, cage tests which fi '
Figure^. Infection of (a) male (Mag. x 25) and (b) female (Mag. x 12)
,Musca autuinnalls adultly Entonujphthora speciea (Em).
\
^ 4 58
-
* r
i '
•
_,, _ — -
• < i'
•
a
4
• - • ... ' '
•
- s y l •• b) . •
t
0 * •
•
» •••••,1:.
i 7
• . y •
indicated the sterilized face fly to be less competitive, than ,the nor-
mal male at both radiation doses. Tests in which irradiated males
—competed"With"wild male "flies for females would give an indication as
to how the laboratory reared males might perform in a release.
Males of Ceratitis capitata (Wiedemann) irradiated in
nitrogen were more competitive males than those irradiated in air
(Hooper, 1971a). This was important, since the sterilizing dose of
radiation reduced the sexual competitiveness of the males (Hooper and
Katiyar, 1971). However, this is probably not very significant in
* the face fly unless a 5.0 krad level of irradiation is to be used,
since the male fly irradiated at 2.5 krad was as competitive as the
untreated male at most ratios tested.
9.- The Effect of Irradiation on the Nematode Parasité,
Heterotylenchus autumnalis, of the Face Fly.
The nematode Heterotylenchus autumnalis, Nickle was first
described by Stoffolano and Nickle (1966) and has since been found in
face flies throughout North America. Detailed accounts of the life
cycle of this nemato&e^have been presented by Nickle (1967), Treece and
Miller (1968), and Stoffolano (1970b). The parasite is found in male
and female fy.es but in the male fly the nematode is unable to com-
plete its life cycle. Parasitized female face flies axe "steVi^lized" ,
when thousands of infective nematodes invade the flies* ovaries. The
nematodes complete their development in the ovaries and inhibit -all r normal egg production. "They are transmitted by "mock" oviposition to- fresh cattle droppings where ttfey mate and the infective female in-
•" vades newly hatched face^fly larvae. / , ' V- 60
t,v,), This i-nematode parasite has not lived up,\to, its potential as a
b i oc ont rpl< l'agent against the, face fly." ' An, attempt, was .made to determine
whether this/parasite, cou Id be integrated witTv.the"' sterile'males and '
...females ,i- since ""it"was tealized'that! "the sterile-male,technique for con-
trol of face flies could not succeed alonel'-^ExpetimentslWere conducted
pn face'^fly|'pupae parasitized with the nematbde parasité Heterotylenchus \
autumnal is using small doses of radiation to determine:- Cl) whether * '"•'
or not the nematodes would survive the irradiation dose, and (2) whether^
or not they would be sterilized by that dose.
Five-day-old pupae from a nematode parasitized colony-were
used for irradiation. After irradiation, pupae were handled in the
same manner as normal irradiated pupae. Two dosages were employed. • < * fc - • • . ' In the first three tests, 1.0 krad was used. Previous results (Tables r
3 and 4) had shown that this dose produced a 987. sterility in males.
Female/é so irradiated produced few eggs. • It was thought that the
abirljLty to produce a few eggs might be important to the successful de-
velopment of the parasitic nematode'. In a fourth test, a higher dose
(2.5 krad) was employed. Following this exposure, males were ste-
rilized permanently and females produced no eggs (Tables 3 and 4).
In the.first 2 experiments, untreated "and irradiated flies
from a nematode parasitized colony (parental colony P) and flies from a '
USDA colony were combined* in two separate cages (Cage 1 and Cage 2).
Cage 1 contained 200 untreated, first generation females from para-
sitized, pupae, whereas Cage 2 contained 200 irradiated, first
generation females from parasitized pupae. To ensure adequate egg
production, 100 normal USDA females and 100 normal USDA males were
added to each cage. The addition of the normal USDA males prevented • " ny possible parasitization arising from matings using parasitized '
males. Oviposition medium was offered for 1 hour on'the 14th day after
...emergence. - -After -oviposit iony~aH~" females"were dissected (in 0.9% a. 'Rfnger's solution) to determine the level of parasitism of the first
generation adults. The eggs produced by the first- generation adults
were handled according to normal rearing procedures.* The.resulting . \t >f r
second generation adults were kept 14 days, at which time they were
dissected to determine the level of 'parasitism by the nematode. Re- •
sul^Cof these first 2 experiments are shown in, Table 19. *
TABLE 19. Levels of parasitism for Musca autumnalis females with Che nematode, Heterotylenchus autumnalis, through two generations after irradiation of 5-day-old parasitized pupae at 1.0 and' 2.5 krad.-
Nematode Infection (%) of: Irradiated females, Untreated females, 1 Parental generations - generations/ Experiment colony (P) 1 ; 2 1 2
6.3 6.0 ; 41.2 8.5 21.4
4.0 5.0 75.0 • > ,4.0 . 28.0
.28.7 49.0 c 25.0 0 0 0 0
300 females dissected for first generation level'of parasitism. All
second generation females resulting from first generation adults
dissected. • - • ' ', b 450 females dissected for first generation'level of parasitism. AljL second generation females resulting from first generation adults /*.
dissected. t * c No data as a result of infection by Entomophthora species.
4 . V-A 62 .1
The level of parasitism increased from 6.37. and 4.0% in the
parental stock (.in the untreated females. The increase Was more dramatic with irra-
..diatedrfemales j - increasing -to 41r2Z~afid 75.07." in™"thê~"secônd generation
adults.a *The'se results-indicated that either there was a'more com- 4
•\ of egg-laying females "in Cage 1 at the first generation was Influencing
. the, level of parasitism of untreated second generation females. If we
- presume a 10% level of parasitism in the first generation adults
t • . • (actually 6.0'dnd 8.5% in the first experiment) for Irradiated and un- i .treated females, Cage. 1 would contain 280 egg-laying females. Cage 2
would have the same number of,, females but the 180 females from the para- ./ s itized pupae , were irradiated'and produced only a few eggs, therefore, ,
Cage 2 contained only 100 egg-laying USDA females and this was thought
.- to influence the result's*9of the first 2 experiments. .* '. '_ ' Tcv investigaçe^che' possibility of a more,competitive nematode, I ' a third experiment usiixg^l^-O ij£rad and a fourth experiment .at 2.5 krad
' •* werfe conducted using a 'slrghtly.^different experimental design'.- The, •
difference was the addition of 150''irradiated USDA females to Cage 1
' and 150 normal USDA females to .Cage'jL. j This design provided the same , r number of flies in both cages and approximately the dame number of .egg-
producing females, depending on the, level .pf parasitism of the female
face flies.. As in the- previous-,test8y females from the first and
' ' ,' $ ' • " . second generations were dissected at 14 days of age to determine the •; ' ' f>' •' •*'"**'.; -•:'{" \ ^levels of paraa.itisn and" the results tabulated (Table 19.).' \ \ • -, ; • '•• In the third fexper Leant, levels of paras it 1 a =v.tor the
parental varïd' :Vrst gt'eeratten adiflta Vsre =£ich higher than these f du
4 \ •
. $ V
63 y ri1* , , in the first 2 experiments. Consequently, only a few pupae survived * v ' ' y and these were preserved to maintain W:he stock colony. These results » . • • • •_ -f- »« indicated that_the greater .number—of~egg-laying females-in"Câgé"l""at~ " * " /
the first generation were influencing the levels of parasitism of the
second generation and were not the res\ilt of a. more competitive
nematode. , ' • . .
Infection by 'tan Entomophthora species affected the results of
the fourth»experiment. This fungus -attacked only irradiated'flies
(Fig. 8). Survival of first generation adults was poor, so that at
day 14, only one\ parasitized female had survived and.no p"arasitization
of second generation adults was achieved. Untreated f irst*gè-neration
adults>were not carried through to the second generation. Dissection
of first generation females showed invasion of the ovaries by the
•' • , *
nematodes in both the irradiated and untreated females. This indicated
that parasitization of second generation 'adults Would«probably have
occurred had, not the Entomophthora infection been present. The fungus
infection was so complete that it made determination of parasitism in
' females almost impossible. > • , ** ,
Although-no critical.estimate was made of the number, of ih- . i ; " 'a
fective nematodes present in the haemocoel or\oyaries of either un- • ,
treated or irradiated females at each level of ^irradiation, there was
no obvious difference, between the two groups of females'. This' in- ^ *
'dicatéd that' the'nemacodes were not being ste-rilize,d( at the levels .of
Irradiation used. .••/", . ' ' • •
Gasjogenetlc fçnale nenatodes were t i3und tror di§e.d
•r^il- t'liea In an e-ifwr:renu As^iLSfet^'-^s. da, . art-r "•« .«xperiaer.ir it.t
\ '. 64
Infective nematodes were never found invading ovarian tissue before day
9. Infective nematodes are shown invading the ovaries of irradiated and untreated females in Figure 9. The large numbers of infective _neraatode8 supported-by-a'^elttà'le-fly^is 'shown in Figure 10. So many r /' ' ' • • I • • nematodes are present in the haemocoel that ,the abdominal sclerites
have separated releasing some of the nematodes. „ "/ 'jr/H '•-'? Percentages for adult eclosion from irradiated and untreated
'"''• / '' • ' ' parasitized pupae were similar. At 1.0 krad, 90.47» of adults emerged* / i whereas at 2. 5 krad 92.37o of adults emerged. The-corresponding eclosion8 for untreated adults was 89.4% and 91.4% respectively. Some difficulty was encountered in maintaining a stock of .
'/flutes with a high level of parasitism. Part of this'difficulty may
have been to larval mortality caused by the•nematode. Branch and Nicholas (1970JJO) reported that as few as two nematode larvae of the Heterotylenchus species may kill larvae of the Australia\ n bush fly,
Musca vetustissima Walker. However, by. careful .mixing of paras it iz'ed '
face flies with non-parasitized flies, it was -possible to produce con-
sistently a generation of>flies with a 25-30% level'of parasitism.\
Further refining of this technique to ensure adequate infusion of non-'
• * i f i
parasitized flies or by addition of non-parasitized eggs to eggs and
nematodes collected from a 'nematode-parasitized colony, should maintain
production of flies with a level of parasitism of 757. or higher. * A
level of parasitism this high used in a sterile-male release would
«*, certainly be more advantageous than the release of sferile insects '
alone v .• .'••••' I
Figure 9. Infective nematodes \(N) invading the ovarian tissue of
14-dky-old (a) irradiated and (b) untreated Musca • * autumnalia females.and also present in the oviduct.(OV)
' of the, flies. (M*.g. (a) x 25; (b) x 50). 65 I
ii
1 1
• / * \
, r
•
f
t s-
r ! , fi
t *
) Figure*10. Infedtive nematodès (N) present in the haemocoel of an
'untreated Mtisca autumnal is female protruding through v,
the intersegmental membrane. ('Mag. x 12).
\
i u
66 r
^ < U
DISCUSSION AND CONCLUSIONS ' . t ' !
Discussion, pertinent to the results for each experiment, -has •
__been?preBented -in -the-previous-section"t"d"mainTrâin~clïntinuity. ^Thi's
section will present a discussion of the general e-ffiêcts of,:
irradiation on the, face fly and suggest a potehtJ/al role of «|
, male technique in an integrated control 'program for this pestt'1
Relï'e'ase of sterileS^male' insects is usually considered for
three types of circumstances: (1) for control of important pesés»that
are normally present in small numbers; (2)- control of newly estab-,
lished insect populations before the population density becomes high;
or' (3), as an adjunct to other control measures,when the target species
is abundant and widelf&^&tributed (Kniplihg1, 1955). The third 'sit- uation applies to the^sfaac'e if$Y fly with insecticides .and' bioc^ntrol agents has been inadequate. Ob-'
viously. a ccraibfnati'ortVoftfconM^l techniques must be employed for *,Hi? ~
successful control oif/thiJs'[(^efc1^ * To evaluate the potential of sterile
males in such an in^.eg^ra^t|ed concrol program,, research was conducted to
examine the effects oï.irradiation on the face fly.
An integrated control program through the use of insecti-
cides, biôcontrol agents, or cultural .practices might be employed,to
reduce face" fly'populations to a low level", at which- time, releases of
stBerile^male.8 would be economically- f,easib,le . The theoretical ad- / •>a vantages of integrating two or more control sfocedures to achieve
eradication by sterile-male releases has been di.scussed by Knipling J ' ^ • tages of integrating .insedti^
cidal cûntrol with "the s^cer 1 le-=^lea appro-ach have been discussed •. ™
ieai'.is v\ î !» p-r p^21 a t -. c-r. s -: V.e t \ z a l'l . rc--'rrar: ~s- 's .s_c- a.' 68 \ •
Musca dome8tica (L.) (LaJJrecque and Weidhaas, 1970).' The integration
of techniques.such as: (1) application of insecticides, , (2) chemo-
sterilant baits, and (3) sterile-male releases would'redûce^Jthe_number^
of treatments required with insecticides or chemosteril'ants, reduce the
number of insects needed for the releases, and reduce the tine required
to reach theoretical eradication. These techniques combined with other
control techniques: (1) us.e of pheromones (Chaudury ££ a^. , 1972);
(2) release of irradiated, flies parasitized with H. autumnalis;
(3) treatment with larVicides; and (4) releases of parasites and pre-
* •••'.;
dators, could'possibly be used in, an integrated control' program against
the face fly. Success would depend on the timing of these integrated
control-measures to obtain the optimum effect from each.
~N-"*-v Recently completed research .has diiown that irradiation of the
5-day-old. pupae with 2.5 krad^induced permanent sterility in the male
face'fly and resulted in a total loss of egg production in the female
without adversely affecting their longevity. Competitiveness, as ,
i • < i
measured by mating tests and ratio tests, indicated that the irradiated
male was as competitive as the normal male. These- results indicated
that the face, fly can be sterilized without harmful side.-effects;. the
sterile-male technique shows some promise in an integrated control program1. However, further competitive tests are necessary with large X cages, since extensive replication is necessary to determine differ-
ences in competitiveness. Furthermore, these tests should be extended
to include competitive tests with wild flies, for if wild females pre-t
f erred wild males, or if the-females wojild not mate at all with the*
introduced naLee, Chen the release pr,ograa would t>^ '•nanrrf*««ful.
2xy«.r : -*r t s wur. a native colony of face flies. Indicated no" dit f ercs.cc' 69
/ from the USDA colony in the dose necessary to induce sterility or '
longevity in the male. No comparison was made between the native
ny and USDA colony using competitive tests. However, in "a com- / par ison of two laboratory strains and one wild strain of'house flies, ,\ / Fye and LaBrecqu'e {1966) found that females of all strains mated more
readily with males of their own strain whether or not the males were '
sterilized. ' . - • .
It « apparent that the sterile-male approach alone cannot
be used practically to control or eradicate populations of such great
density and dispersal as the Jsace fly. However, when integrated with
other control methods, it would have a much better chance of con-
tributing to a successful control program. Initially, face fly pop-
ulations would' have to be reduced with control, measures such as timed p
aduXricide and larvicide applications. This could possibly be com-
bined with, chemosterilant baits and releases of promising biocontrol
agents.k Once the face fly population had been reduced to sufficiently '
low levels; the release of sterile males and females could begin. It
was shown that the nematode parasite, Heterotylenchus autumnalis sur-
vived i'r radiât ion in the face fly pupae without itself being sterilized. "'
The competitiveness of the' irradiated, parasitized adult flies is un- ,
known. However, if they are as competitive as normal flies,, then the," X release of these parasitized flies'might prove, more beneficial than the/ release crçy'j^Jie sterile males alone. Not only would the sterilized v
males compete for mates', but the "steri le'1, females would increase- the
level of H.' Is in the natural habitat during false) ovipostçion
and also cocpete for =-ites Releases should be tired :oAbdel-Malek .et al. , 1969) .
l| The possibility of the irradiated fly being more' susceptible
„ —-.-. f —<—1~ to fungus infection might complicate a release program. Jaffi (1967) reported an increased susceptibility to infection by Triboliutn castaneum (Herbst.) and T. confuaum Duv. to Bacillus thuringiens°is
Berliner following exposure to X-rays. Infection of dipteran genera
•/- • " , by Entomophthora species in'Ontario seems 'to reach a peak in the fall.
(MapLeod', 1956; Miller and McClanahan, 1959). However, no infection by IS. muscae has been recorded in field-collected face flies. A re- lease program for sterile face flies, timed for the spring would ( probably minimize losses due to thiJ"i3 fungus.
•s» I
"3\ ,< 1 i' V I 7
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.BAUMHOVER, A.H. 1963. Influence of aeration during'gamma irradiation of screw-worm pupae. J. Ecoft. Entomol. 56(5): 628-631.
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I— ! BURTON,' R.P. , and E.C. TURNER. 1968. Laboratory propagation of Mu8cidifurax raptor on face fly pupae. J. Econ. Entomol. 61(5): 1380- 1383. BUSHLAND, R.C.,*and D.E. HOPKINS.^ 1951. Experiments with screw-worm flies sterilized by-X-rays. J. Econ.'Entomol; 44(5): 725-731.
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