JOURNAL
OF INVERTEBRATE
PATHOLOGY
30, 155- 159 (1977)
3iology of Octomyomermis muspratti, a Parasite of Mosquitoes Relates to Mass Production1
as It
JAMES J. PETERSEN Gulf
Coast
Mosquito
Research
Laboratory, Agriculture,
Agricultural Lake Charles,
Research Louisiana
Service, 70601
United
States
Department
of
Received November 22, 1976 The first- and second-instar larvae of Culex p&ens quinquefasciatus were equally susceptible to parasitism by the mermithid nematode Octomyomermis muspratti; the third-instar hosts were signilicantly less susceptible. The yield of postparasitic nematodes was higher from hosts exposed as second instars then from hosts exposed as first or third instars at comparable levels of parasitism. Mortality of the host prior to emergence of the nematode occurred most frequently among firstand third-instar hosts. About 4% of the infected hosts exposed as first and second instars and 37% of the infected hosts exposed as third instars pupated prior to emergence of the postparasitic nematodes. The exposure of mosquitoes as second instars was the most efficient for mass rearing and resulted in the highest yields of postparasitic 0. muspratti.
INTRODUCTION
In recent years much attention has been focused on mermithid nematodes as biological control agents of mosquitoes, especially Romanomermis culicivorax (=Reesimermis nielseni) (Petersen, 1975). However, like any biological agent, a given mermithid species can only be effective under certain environmental conditions. It is therefore desirable to develop a number of these agents with differing environmental tolerances that would permit their use over a wide range of host habitats. The nematode, Octomyomermis muspratti (=Reesimermis muspratti), originally isolated from parasitized larvae of tree-hole Aedes and Culex mosquitoes in Zambia (Muspratt, 194.5), appears to have a greater tolerance to desiccation, salinity, and pollution (unpubl.) than does R. culicivorax, the only mermithid nematode now in mass production. 0. muspratti was the first reported mermithid nematode of mosquitoes to be maintained in the laboratory (MusPratt, 1947) and was later described and I In cooperation with McNeese State University. Lake Charles, Louisiana 70601.
studied in some detail by Obiamiwe and MacDonald ( 1973). A study was therefore conducted at the Gulf Coast Mosquito Research Laboratory, Lake Charles, Louisiana, to obtain specific data pertaining to the development of an in vivo mass-rearing system for 0. muspratti and, ultimately, to provide sufficient material for its safety and field testing. MATERIALS
AND METHODS
The stock cultures that produced the preparasitic (infective stage) nematodes were maintained and handled in a manner similar to that used for R. culicivorax but on a smaller scale (Petersen and Willis, 1972). Culex pipiens quinquefasciatus were used as hosts because they are easily reared in the laboratory and because members of the C. pipiens complex have been reported to be suitable hosts of 0. muspratti (Muspratt, 1965). A test consisted of exposing six groups of 100 first- or second-instar mosquito larvae to 0. muspratti at rates of 1, 2.5, 5, 7.5, 10, and 15 nematodes per larva, or of exposing seven groups of 100 third-instar hosts at ratios of 1, 2.5, 5, 7.5, 10, 15, and 20: 1. 155
Copyright 8 1977 by Academic Press. Inc. All rights of reprducfion in any form reserved.
ISSN 0022-201 I
156
JAMES
J. PETERSEN
The tests were replicated five times for each host instar. The mosquitoes were exposed to the nematodes in 100 ml of water in 150ml beakers for 24 hr, placed in trays, and allowed to develop by using standard rearing techniques. After 12 days, infected larvae were removed from the trays, isolated in individual cells of spot plates, and placed in a 100% RH environment (to prevent desiccation of the water in the cells) until counts were made of the emerging nematodes, Pupae were removed from the rearing trays as they developed and were dissected to determine the extent of parasitism. The preparasites were l- to 7-hr old at the beginning of the tests, and their numbers were determined by the procedure described by Petersen and Willis ( 1972). An unexposed control population was reared simultaneously with each of the 15 tests to permit a measurement of premature mortality caused by the parasites. RESULTS
First- and second-instar C. p. quinquefasciatus were not significantly different (P > 0.70) in their susceptibility to parasitism by
0. musprutti (Fig. 1). Parasitism of both instars was essentially linear when parasite dosages ranged from 1 to 7.5 per host and increased about 10% (b = 9.50 for first instars; b = 9.80 for second instars) for each unit increase in the ratio of parasites to hosts. Parasite to host ratios of 1: 1 and 7.5: 1 produced means of 23 and 85% parasitism, respectively. The susceptibility of third-instar C. p. quinquefusciatus was significantly lower (P < 0.01) than that for the younger instars, and the incidence of parasitism was linear when dosages ranged from 1 to 15 parasite per host. Parasitism increased about 5% (b = 4.49) with each unit increase in the ratio of parasites to host, and ratios of 1: 1 and 7.5: 1 produced means of 9 and 47% parasitism, respectively. Although infection levels at a given parasite to host ratio were the same for firstand second-instar larvae, hosts exposed as first instars produced fewer postparasitic nematodes than did those exposed as second instars, and hosts exposed as third instars produced the fewest postparasites at comparable levels of parasitism (Fig. 2). A linear relationship in numbers of postparasites produced existed for the first (b
loo-
. . . . . . . . 1st lnstor . . . . . . . . 2nd lnrlor w 3rd Inr1.r
0 100
FIG. 1. Relationship C&x
pipiens
250
500 NUMBER
OF
750 1000 PREPARAsITES
of parasite-host ratios to the incidence quinquefasciafus by Odomyomermis muspratti.
PER 100
1500 HOSTS
of parasitism
2000
of first-,
second-,
and third-instar
BIOLOGY
OF 0. MUSPRATTI
157
300’
250.
VI : 200. 0 2 z z 1508 f 2 z
loo-
50.
01 0
-
15
30
45 PERCENT
60 PARASITISM
75
, 100
90
FIG. 2. Relationship of the incidence of parasitism of first-, second-, and third-instar to the production of postparasitic Octomyomermis musprutti.
Culex p&ens
quinquefusciatrrs
= 1.76) and third instars (b = 1.56, and there was no evident upswing in the numbers of nematodes produced as the levels of parasitism approached 1%. However, postparasite production by hosts
exposed as second instars tended toward linearity (b = 2.43) only until the levels of parasitism reached about 90%. When levels of parasitism were above %, a noticeable increase was evident. The numbers of
0 0
15
30
75 PERCEtif
90
100
PARAS&A
FIG. 3. Relationship of premature mortality of first-, second-, and third-ins&r ,fasciarus to the incidence of parasitism by Octomyomermis musprutti.
C&x
pipiens
quinque-
158
JAMES J. PETERSEN TABLE
1
THE PERCENTAGE OF PUPATION OF FIRST-, SECOND-, AND THIRD-INSTAR CULEX PIPIENS QUINQUEFASCIATUS LARVAE INFECTED WITH OCTOMYOMERMIS
MUSPRATTI
48% (33-61%).
Pupation in indicated test (%) Host instar
1
2
3
4
5
x
1 2 3
1.2 8.5 22.2
8.0 2.5 8.2
4.6 1.3 63.8
5.7 3.9 2.1
1.2 3.8 89.9
4.1 4.0 37.2
postparasites per 100 exposed hosts averaged 293 (231-473). Premature mortality (mortality prior to emergence of the nematodes) of infected hosts was observed among all three instars (Fig. 3). The extent of this mortality was lowest in hosts exposed as second instars, but increased with the intensity of parasitism (b = 0.23). A mean of 25% (11-37%) mortality occurred when levels of parasitism ranged from 90 to 100%. Although the mortality rate was higher for hosts exposed as third instars (b = 0.29), the difference was not significant (P > 0.50)) and mortality usually occurred between 10 and 14 days after exposure while the hosts were in the process of molting to the pupal stage. In contrast, the mortality rate (b = 0.42) for hosts exposed as first instars was significantly higher (P = 0.05) than that for hosts exposed as second instars, and TABLE
2
PERCENTAGE OF PUPATION OF FIRST-, SECOND-, AND THIRD-INSTAR CULEX PIPIENS QUINQUEFASCIATUS LARVAE INFECTED WITH OCTOMYOMERMIS MUSPRATTI AT GWEN LEVELS OF PARASITISM IN THE HOST POPULATION Pupation at indicated level of parasitism (%I HO9
instar
O-20
21-40
41-60
61-80
81-100
,t’
SE
I
3.w 4.7 29.9
3.0 2.9 35.9
3.3 3.4 37.3
5.1 3.4 48.9
4.5 5.4 33.2
4.06 4.05 37.2
0.55 0.62 5.07
2 3
mortality usually occurred within 1- 3 days after exposure; mortality increased materially when the incidence of parasitism ranged between 90 and 100% and averaged
” Mean for all values within the given range for five tests.
There was a significant difference between tests in the percentage of infected hosts pupating for a given instar, which indicated that factors other than host age at the time of exposure affected pupation of infected hosts (Table 1). When the percentage of infected pupae was compared with the incidence of parasitism for each instar (an indirect measurement of the effect of multiple parasitism), no correlation was obtained (regressions were positive for all three instars but none were significant; Table 2). The incidence of infected pupae was the same for hosts exposed as first and second instars and averaged about 4%. However, pupation was significantly higher (P < 0.01) for third-instar hosts, averaging 37%. DISCUSSION Octomyomermis muspratti differs biologically from the more intensely studied species R. culicivorax in the following ways: 0. muspratti required 1 l- 14 days to emerge from its host at 26-28°C (ambient temperature) compared with 7-10 days for R. culicivorax. The ability of hosts infected with 0. muspratti to pupate increased with age; hosts infected with R. culicivorax failed to pupate (Petersen and Willis, 1970). Males rarely made up more than 30% of 0. muspratti populations, even when infection levels in the host population approached 100%. This contrasts with 75% males for R. cuiicivorax under similar conditions (Petersen, 1972). More importantly and as previously mentioned, preliminary observations indicate that 0. muspratti is more tolerant of pollution and salinity and, thus, may be useful in certain environments that are inhibitory to R. culicivorax. Since the biology of these two species differs significantly, modified rearing techniques will be necessary to mass produce 0.
BIOLOGY
The data show that first- and second-instar C. p. quinquefasciatus are equal& susceptible to 0. m~s~ratti, and that both are about twice as susceptible as thirdinstar hosts. However, when premature mortality is considered, first-instar hosts appear less favorable than do second-instar hosts because of their much higher mortality rate, especially at high levels of parasitism. Third-&tar hosts appear undesirable because of their higher mortality and their ability to pupate while infected. The numbers of nematodes produced substantiate these observations and show that hosts exposed as second instars produced substantially more nematodes than the other instars, especially at the higher levels of infection. Other factors such as feeding regimens and their influence on male-female ratios that may influence in vivo mass production of 0. muspratti remain to be studied (Obiamiwe and ~ac~n~d, 1973; Petersen, 1972). Also, Muspratt (1965) reported that high temperature and food rich in vitamin content may result in accelerated pupation of larvae infected by the nematode. It is possible that with the proper restrictive diet, third-instar hosts may be the best host stage for mass rearing of this parasite. 0. muspratti is presently maintained in the laboratory with little difficulty, but more research is needed before the parasite can
musprutti.
1.59
OF 0. MUSPRATTI
be produced in sufficient testing.
numbers for field
ACKNOWLEDGMENT The author thanks J. Muspratt, Entomologist, South African Institute for Medical Research, Johannesburg, South Africa, for generously supplying the initial culture of 0. mrrsprutfi.
REFERENCES MUSPRA~T, J. 1945. Observations on the larvae of tree-hole breeding culicini (Diptera:Culicidae) and two of their parasites. J. Entomol. Sot. Afv.. 8, 1320. MUSPRATT, J. 1947. The laboratory culture of a nematode parasite of mosquito larvae. J. ~~ro~o~. Sot. Afr.. 10, 131-132. MUSPRATT, J. 1965. Techniques for infecting larvae of the C&x pipiens complex with a mermithid nematode and for culturing the latter in the taboratory. Bull. WHO, 33, 140- 144. OBIAMIWE, B. A., AND MACDONALD, W. W. 1973. A new parasite of mosquitoes, Reesimermis musprat?i sp. nov. (Nematoda:Mermithidae), with notes on its life-cycle. Ann. Trap. Med. Parasitol.. 67, 439-444. PETERSEN, J. J. 1972. Factors affecting sex ratios of a mermithid parasite of mosquitoes. J. Nematol., 4, 83-87. PETERSEN. J. J. 197.5.Status of nematodes as mosquito control agents in North America. In Proc. Mosq. Abat. Sym., Feb. 25-27, pp. 181-190. Edmonton, Alberta, Canada. PETERSEN, J. J., AND WILLIS, 0. R. 1970. Some factors affecting parasitism by mermithid nematodes in southern house mosquito iarvae. J. Econ. Entomol., 63, 175-178. PETERSEN. J. J.. AND WILLIS, 0. R. 1972. Procedures for the mass rearing of a mermithid parasite of mosquitoes. ~5squ~~oNews, 32, 226-230.
OF INVERTEBRATE
PATHOLOGY
30, 155- 159 (1977)
3iology of Octomyomermis muspratti, a Parasite of Mosquitoes Relates to Mass Production1
as It
JAMES J. PETERSEN Gulf
Coast
Mosquito
Research
Laboratory, Agriculture,
Agricultural Lake Charles,
Research Louisiana
Service, 70601
United
States
Department
of
Received November 22, 1976 The first- and second-instar larvae of Culex p&ens quinquefasciatus were equally susceptible to parasitism by the mermithid nematode Octomyomermis muspratti; the third-instar hosts were signilicantly less susceptible. The yield of postparasitic nematodes was higher from hosts exposed as second instars then from hosts exposed as first or third instars at comparable levels of parasitism. Mortality of the host prior to emergence of the nematode occurred most frequently among firstand third-instar hosts. About 4% of the infected hosts exposed as first and second instars and 37% of the infected hosts exposed as third instars pupated prior to emergence of the postparasitic nematodes. The exposure of mosquitoes as second instars was the most efficient for mass rearing and resulted in the highest yields of postparasitic 0. muspratti.
INTRODUCTION
In recent years much attention has been focused on mermithid nematodes as biological control agents of mosquitoes, especially Romanomermis culicivorax (=Reesimermis nielseni) (Petersen, 1975). However, like any biological agent, a given mermithid species can only be effective under certain environmental conditions. It is therefore desirable to develop a number of these agents with differing environmental tolerances that would permit their use over a wide range of host habitats. The nematode, Octomyomermis muspratti (=Reesimermis muspratti), originally isolated from parasitized larvae of tree-hole Aedes and Culex mosquitoes in Zambia (Muspratt, 194.5), appears to have a greater tolerance to desiccation, salinity, and pollution (unpubl.) than does R. culicivorax, the only mermithid nematode now in mass production. 0. muspratti was the first reported mermithid nematode of mosquitoes to be maintained in the laboratory (MusPratt, 1947) and was later described and I In cooperation with McNeese State University. Lake Charles, Louisiana 70601.
studied in some detail by Obiamiwe and MacDonald ( 1973). A study was therefore conducted at the Gulf Coast Mosquito Research Laboratory, Lake Charles, Louisiana, to obtain specific data pertaining to the development of an in vivo mass-rearing system for 0. muspratti and, ultimately, to provide sufficient material for its safety and field testing. MATERIALS
AND METHODS
The stock cultures that produced the preparasitic (infective stage) nematodes were maintained and handled in a manner similar to that used for R. culicivorax but on a smaller scale (Petersen and Willis, 1972). Culex pipiens quinquefasciatus were used as hosts because they are easily reared in the laboratory and because members of the C. pipiens complex have been reported to be suitable hosts of 0. muspratti (Muspratt, 1965). A test consisted of exposing six groups of 100 first- or second-instar mosquito larvae to 0. muspratti at rates of 1, 2.5, 5, 7.5, 10, and 15 nematodes per larva, or of exposing seven groups of 100 third-instar hosts at ratios of 1, 2.5, 5, 7.5, 10, 15, and 20: 1. 155
Copyright 8 1977 by Academic Press. Inc. All rights of reprducfion in any form reserved.
ISSN 0022-201 I
156
JAMES
J. PETERSEN
The tests were replicated five times for each host instar. The mosquitoes were exposed to the nematodes in 100 ml of water in 150ml beakers for 24 hr, placed in trays, and allowed to develop by using standard rearing techniques. After 12 days, infected larvae were removed from the trays, isolated in individual cells of spot plates, and placed in a 100% RH environment (to prevent desiccation of the water in the cells) until counts were made of the emerging nematodes, Pupae were removed from the rearing trays as they developed and were dissected to determine the extent of parasitism. The preparasites were l- to 7-hr old at the beginning of the tests, and their numbers were determined by the procedure described by Petersen and Willis ( 1972). An unexposed control population was reared simultaneously with each of the 15 tests to permit a measurement of premature mortality caused by the parasites. RESULTS
First- and second-instar C. p. quinquefasciatus were not significantly different (P > 0.70) in their susceptibility to parasitism by
0. musprutti (Fig. 1). Parasitism of both instars was essentially linear when parasite dosages ranged from 1 to 7.5 per host and increased about 10% (b = 9.50 for first instars; b = 9.80 for second instars) for each unit increase in the ratio of parasites to hosts. Parasite to host ratios of 1: 1 and 7.5: 1 produced means of 23 and 85% parasitism, respectively. The susceptibility of third-instar C. p. quinquefusciatus was significantly lower (P < 0.01) than that for the younger instars, and the incidence of parasitism was linear when dosages ranged from 1 to 15 parasite per host. Parasitism increased about 5% (b = 4.49) with each unit increase in the ratio of parasites to host, and ratios of 1: 1 and 7.5: 1 produced means of 9 and 47% parasitism, respectively. Although infection levels at a given parasite to host ratio were the same for firstand second-instar larvae, hosts exposed as first instars produced fewer postparasitic nematodes than did those exposed as second instars, and hosts exposed as third instars produced the fewest postparasites at comparable levels of parasitism (Fig. 2). A linear relationship in numbers of postparasites produced existed for the first (b
loo-
. . . . . . . . 1st lnstor . . . . . . . . 2nd lnrlor w 3rd Inr1.r
0 100
FIG. 1. Relationship C&x
pipiens
250
500 NUMBER
OF
750 1000 PREPARAsITES
of parasite-host ratios to the incidence quinquefasciafus by Odomyomermis muspratti.
PER 100
1500 HOSTS
of parasitism
2000
of first-,
second-,
and third-instar
BIOLOGY
OF 0. MUSPRATTI
157
300’
250.
VI : 200. 0 2 z z 1508 f 2 z
loo-
50.
01 0
-
15
30
45 PERCENT
60 PARASITISM
75
, 100
90
FIG. 2. Relationship of the incidence of parasitism of first-, second-, and third-instar to the production of postparasitic Octomyomermis musprutti.
Culex p&ens
quinquefusciatrrs
= 1.76) and third instars (b = 1.56, and there was no evident upswing in the numbers of nematodes produced as the levels of parasitism approached 1%. However, postparasite production by hosts
exposed as second instars tended toward linearity (b = 2.43) only until the levels of parasitism reached about 90%. When levels of parasitism were above %, a noticeable increase was evident. The numbers of
0 0
15
30
75 PERCEtif
90
100
PARAS&A
FIG. 3. Relationship of premature mortality of first-, second-, and third-ins&r ,fasciarus to the incidence of parasitism by Octomyomermis musprutti.
C&x
pipiens
quinque-
158
JAMES J. PETERSEN TABLE
1
THE PERCENTAGE OF PUPATION OF FIRST-, SECOND-, AND THIRD-INSTAR CULEX PIPIENS QUINQUEFASCIATUS LARVAE INFECTED WITH OCTOMYOMERMIS
MUSPRATTI
48% (33-61%).
Pupation in indicated test (%) Host instar
1
2
3
4
5
x
1 2 3
1.2 8.5 22.2
8.0 2.5 8.2
4.6 1.3 63.8
5.7 3.9 2.1
1.2 3.8 89.9
4.1 4.0 37.2
postparasites per 100 exposed hosts averaged 293 (231-473). Premature mortality (mortality prior to emergence of the nematodes) of infected hosts was observed among all three instars (Fig. 3). The extent of this mortality was lowest in hosts exposed as second instars, but increased with the intensity of parasitism (b = 0.23). A mean of 25% (11-37%) mortality occurred when levels of parasitism ranged from 90 to 100%. Although the mortality rate was higher for hosts exposed as third instars (b = 0.29), the difference was not significant (P > 0.50)) and mortality usually occurred between 10 and 14 days after exposure while the hosts were in the process of molting to the pupal stage. In contrast, the mortality rate (b = 0.42) for hosts exposed as first instars was significantly higher (P = 0.05) than that for hosts exposed as second instars, and TABLE
2
PERCENTAGE OF PUPATION OF FIRST-, SECOND-, AND THIRD-INSTAR CULEX PIPIENS QUINQUEFASCIATUS LARVAE INFECTED WITH OCTOMYOMERMIS MUSPRATTI AT GWEN LEVELS OF PARASITISM IN THE HOST POPULATION Pupation at indicated level of parasitism (%I HO9
instar
O-20
21-40
41-60
61-80
81-100
,t’
SE
I
3.w 4.7 29.9
3.0 2.9 35.9
3.3 3.4 37.3
5.1 3.4 48.9
4.5 5.4 33.2
4.06 4.05 37.2
0.55 0.62 5.07
2 3
mortality usually occurred within 1- 3 days after exposure; mortality increased materially when the incidence of parasitism ranged between 90 and 100% and averaged
” Mean for all values within the given range for five tests.
There was a significant difference between tests in the percentage of infected hosts pupating for a given instar, which indicated that factors other than host age at the time of exposure affected pupation of infected hosts (Table 1). When the percentage of infected pupae was compared with the incidence of parasitism for each instar (an indirect measurement of the effect of multiple parasitism), no correlation was obtained (regressions were positive for all three instars but none were significant; Table 2). The incidence of infected pupae was the same for hosts exposed as first and second instars and averaged about 4%. However, pupation was significantly higher (P < 0.01) for third-instar hosts, averaging 37%. DISCUSSION Octomyomermis muspratti differs biologically from the more intensely studied species R. culicivorax in the following ways: 0. muspratti required 1 l- 14 days to emerge from its host at 26-28°C (ambient temperature) compared with 7-10 days for R. culicivorax. The ability of hosts infected with 0. muspratti to pupate increased with age; hosts infected with R. culicivorax failed to pupate (Petersen and Willis, 1970). Males rarely made up more than 30% of 0. muspratti populations, even when infection levels in the host population approached 100%. This contrasts with 75% males for R. cuiicivorax under similar conditions (Petersen, 1972). More importantly and as previously mentioned, preliminary observations indicate that 0. muspratti is more tolerant of pollution and salinity and, thus, may be useful in certain environments that are inhibitory to R. culicivorax. Since the biology of these two species differs significantly, modified rearing techniques will be necessary to mass produce 0.
BIOLOGY
The data show that first- and second-instar C. p. quinquefasciatus are equal& susceptible to 0. m~s~ratti, and that both are about twice as susceptible as thirdinstar hosts. However, when premature mortality is considered, first-instar hosts appear less favorable than do second-instar hosts because of their much higher mortality rate, especially at high levels of parasitism. Third-&tar hosts appear undesirable because of their higher mortality and their ability to pupate while infected. The numbers of nematodes produced substantiate these observations and show that hosts exposed as second instars produced substantially more nematodes than the other instars, especially at the higher levels of infection. Other factors such as feeding regimens and their influence on male-female ratios that may influence in vivo mass production of 0. muspratti remain to be studied (Obiamiwe and ~ac~n~d, 1973; Petersen, 1972). Also, Muspratt (1965) reported that high temperature and food rich in vitamin content may result in accelerated pupation of larvae infected by the nematode. It is possible that with the proper restrictive diet, third-instar hosts may be the best host stage for mass rearing of this parasite. 0. muspratti is presently maintained in the laboratory with little difficulty, but more research is needed before the parasite can
musprutti.
1.59
OF 0. MUSPRATTI
be produced in sufficient testing.
numbers for field
ACKNOWLEDGMENT The author thanks J. Muspratt, Entomologist, South African Institute for Medical Research, Johannesburg, South Africa, for generously supplying the initial culture of 0. mrrsprutfi.
REFERENCES MUSPRA~T, J. 1945. Observations on the larvae of tree-hole breeding culicini (Diptera:Culicidae) and two of their parasites. J. Entomol. Sot. Afv.. 8, 1320. MUSPRATT, J. 1947. The laboratory culture of a nematode parasite of mosquito larvae. J. ~~ro~o~. Sot. Afr.. 10, 131-132. MUSPRATT, J. 1965. Techniques for infecting larvae of the C&x pipiens complex with a mermithid nematode and for culturing the latter in the taboratory. Bull. WHO, 33, 140- 144. OBIAMIWE, B. A., AND MACDONALD, W. W. 1973. A new parasite of mosquitoes, Reesimermis musprat?i sp. nov. (Nematoda:Mermithidae), with notes on its life-cycle. Ann. Trap. Med. Parasitol.. 67, 439-444. PETERSEN, J. J. 1972. Factors affecting sex ratios of a mermithid parasite of mosquitoes. J. Nematol., 4, 83-87. PETERSEN. J. J. 197.5.Status of nematodes as mosquito control agents in North America. In Proc. Mosq. Abat. Sym., Feb. 25-27, pp. 181-190. Edmonton, Alberta, Canada. PETERSEN, J. J., AND WILLIS, 0. R. 1970. Some factors affecting parasitism by mermithid nematodes in southern house mosquito iarvae. J. Econ. Entomol., 63, 175-178. PETERSEN. J. J.. AND WILLIS, 0. R. 1972. Procedures for the mass rearing of a mermithid parasite of mosquitoes. ~5squ~~oNews, 32, 226-230.
