Hydrellia pakistanae and H. balciunasi, biological control agents of : boon or bust?

Michael J. Grodowitz,1 Michael Smart,2 Robert D. Doyle,3 Chetta S. Owens,2 Robin Bare,2 Christie Snell,2 Jan Freedman1 and Harvey Jones1

Summary

Of four insect species released in North America for the management of hydrilla ( Hydrilla verticillata, Hydrocharitaceae), only the two leaf-mining Hydrellia pakistanae and H. balciunasi have become established. While the flies have exhibited impressive range extensions since their first release in 1987, populations at most sites have remained below what was considered damaging. Recently, modest to large increases in populations followed by hydrilla declines have been observed at several sites including Lake Seminole, Florida, Coleto Creek Reservoir, Texas, and Sheldon Reservoir, Texas, United States of America (USA). Long-term, large tank experimentation has shown that even modest levels of fly damage can significantly reduce hydrilla biomass (50%) and tuber numbers (25%), appar- ently by reducing photosynthesis and thereby decreasing plant vigour and production. Field studies have also substantiated these findings where lower numbers of tubers (60%) were observed at sites on Lake Seminole impacted by fly feeding. While more detailed field evaluations are needed, it appears that these agents have the potential to suppress hydrilla populations over the long term. However, a complex of factors can influence their effectiveness, including temperature, plant nutrition, especially protein levels, crowding and the presence of a capable pupal parasite. Further research is needed, including overseas work to identify additional agents and the implementation of new release programs. Based on field surveys, fly releases may increase the likelihood of impact since US release sites now have as much as seven-fold higher fly numbers and associated damage than non-release sites.

Keywords: biological control, Hydrellia, Hydrilla verticillata.

Introduction subsequently in Louisiana, Alabama, Georgia, Texas and California, United States of America (USA). Its Beginning in 1987, two species of leaf-mining flies in distribution has expanded considerably, now extending the family were introduced to North throughout the Florida peninsula, upwards into the America for the management of hydrilla (Hydrilla vert- Florida panhandle and Georgia, mainly on Lake Semi- icillata, Hydrocharitaceae) (Center et al. 1997, nole, north and west into Alabama, and throughout Grodowitz, et al. 1997). The first species, Hydrellia many locations in eastern and south-eastern Texas. pakistanae Deonier, was introduced into Florida and Direct impact to hydrilla by H. pakistanae has been observed at several locations mainly in northern Alabama, Texas, and Florida (Grodowitz et al. 1995, 1 US Army Engineers Research and Development Center (ERDC), Grodowitz et al. 1999, 2000b), but long-term moni- CEERD-EE-A, 3909 Halls Ferry Road, Vicksburg, MS 39180 USA. toring for impact has been limited. In many areas, intro- 2 US Army Engineers Research and Development Center (ERDC), duced Hydrellia spp. population levels and associated CEERD-EE-A, Lewisville Aquatic Ecology Research Facility, Lewis- damage have been low (Grodowitz 1999, Wheeler & ville, TX 75056 USA. 3 Baylor University, Department of Biology, Waco, TX 76798 USA. Center 2001). Unfortunately, factors accounting for Corresponding author: Michael J. Grodowitz such low populations have not been quantified, but may . include high levels of parasitism, plant nutritional

529 Proceedings of the XI International Symposium on Biological Control of Weeds quality and other forms of predation (Wheeler & Center than H. pakistanae and this may account for the differ- 1996, Dr Jim Cuda, pers. comm., unpublished data). ences in the release effort. The three other introduced insect agents have had limited, if any measurable success. These include the Expansion closely related leaf-mining fly H. balciunasi Bock (Grodowitz et al. 1997), the tuber-feeding weevil Another important measure of success for an agent Bagous affinis Hustache and the stem-feeding weevil B. is its ability to disperse extended distances after initial hydrillae O’Brien (Grodowitz et al. 1995). Of these releases have been discontinued. Hydrellia pakistanae three species, the only agent to become established has has exhibited impressive range expansion since 1987. been H. balciunasi, but expansion in distribution and Considering that this species was released at only about population size has been severely limited. Recent 30 locations in 5 states, it is impressive that it is found surveys have shown the presence of this species at sites in almost every location examined. During surveys in Texas that are substantially removed from the orig- conducted in 2000 (Grodowitz et al. 2000b), new popu- inal release sites in Texas. lations of H. pakistanae were located at sites on the Rio The purpose of this paper is to review the existing Grande near McAllen and Rio Grande City, Texas, that evidence and present new information on the impact of are well over 300 km and 400 km, respectively, from the introduced Hydrellia spp. in the US. Information on the nearest deliberately established populations. establishment success, expansion in distribution, popula- Surveys conducted in 2001 showed that it was present tion increase and ultimate impact will mainly be directed in 50% of non-release sites examined in Louisiana even toward H. pakistanae, since much of the current work though it was released in only one isolated system has focused on this species. The information has been (Lake Boeuf) south-west of New Orleans, Louisiana. obtained from a variety of published and unpublished Sites examined in Louisiana in 2001 ranged throughout research from both controlled experimentation and the state and encompassed almost every considerable actual field studies. Limited information on abiotic or type of hydrilla habitat. Wheeler & Center (2001) noted biotic factors that could possibly be influential, including the occurrence of H. pakistanae in almost every site plant nutrition and parasitism, is also included. examined in Florida. In contrast, Hydrellia balciunasi has exhibited only minimal range expansion. As indicated previously, H. Results and discussion balciunasi was established in only two Texas sites (i.e. Establishment Lake Raven, Huntsville State Park, and Sheldon Reser- voir, near Houston) located in the eastern portion of the Since the first release of H. pakistanae in North state. Surveys conducted in the early- to mid-1990s America in 1987, over 3 million individuals have been failed to reveal its presence in any other location, even released at close to 30 different sites from Florida to with extensive sampling. However, in 1997, H. balciu- California. Establishment success has been high, with nasi was discovered in locations north and north-east of at least 70% of the release attempts having H. pakis- the original two release locations, often in combination tanae present six months or longer after terminating the with H. pakistanae. These sites include ponds at the introductions (Center et al. 1997). Surveys conducted Lewisville Aquatic Ecosystem Research Facility during 2001 and 2002 at release sites in Louisiana and (LAERF) and Cypress Springs Lake near the town of Texas have shown that establishment success may be Mount Pleasant, Texas. Reasons for its recent expan- higher, since H. pakistanae has subsequently been sion are unknown, but offer encouragement for its found at sites where it was thought not to have initially continued expansion success. established. However, observing the agent after such an Determining mechanisms for such large expansions is extended period subsequent to termination of releases difficult, at best, for these species. First of all, they may be due to natural expansion from nearby popula- appear to be relatively weak fliers and are often seen tions. hopping from one resting place to another instead of Compared to H. pakistanae, H. balciunasi has flying. Human or transportation of hydrilla sprigs shown substantially lower establishment success. For containing immatures seems plausible, but established example, establishment success for H. balciunasi was sites and associated population size of the introduced only 18% in 1997, nearly four-fold lower than what was Hydrellia spp. was minimal during this time, hence the observed for H. pakistanae (Grodowitz et al. 1997). In odds of man or carrying Hydrellia spp. laden fact, only two release sites have had verified establish- sprigs seem unlikely. Additional research is warranted. ment of this species, both of which are in Texas. Reasons for such low establishment success for H. balciunasi are unknown. However, much less effort Population increase went into its release in comparison to H. pakistanae. Another important criterion of success is the ability For example, H. balciunasi was introduced in only 11 of the released agents to substantially increase in sites in two states with less than 300,000 individuals. population size. While large expansion in distribution Hydrellia balciunasi was always more difficult to rear is desirable, it is often more important to have corre-

530 Insect biocontrol agents of hydrilla sponding increases in population levels to effect damage (Grodowitz et al. 1994). Collectively, these control. data demonstrate that substantial population increases In closed, controlled environment systems, H. paki- are possible for the introduced leaf-mining flies in stanae populations have been shown to increase signif- experimental or pond situations. icantly (Doyle et al. 2003). In this long-term study, Under field conditions, we see a few sites where examining the impact of herbivory and competition substantial population increases have taken place, but alone and in combination in a 14,000 L tank, large we also see very large variations observed from site to increases in population levels were observed over two site. For example, surveys conducted during 1999 at a growing seasons (Fig. 1). By the end of the 1999 variety of release and non-release sites indicated that growing season (i.e. October), mean immature numbers number of immatures and associated damage varied exceeded 3000 per kg fresh plant weight (Fig. 1a) and tremendously; e.g. 3000-fold for immatures and 40- close to 35% leaf damage (Fig. 1b). Similar results were fold for leaf damage (Fig. 2). Reasons for such large observed in nearby ponds at LAERF, where H. pakis- variations are unknown but may be related to various tanae in small pond systems increased substantially in abiotic and biotic factors as well as the numbers populations in only one growing season (2002) to released into an individual site. almost 5000 immatures per kg (Fig. 1c) with 40% leaf damage (Fig. 1d). Impact These results are similar to previously reported observations from ponds located at the Tennessee Larval feeding action directly impacts hydrilla Valley Authority Muscle Shoals, Alabama. There, internal leaf cellular material. Hence, it is not surprising population levels of H. pakistanae reached mean values that a decrease in light-saturated photosynthesis is posi- of almost 7000 immatures per kg with close to 70% leaf tively correlated with increasing leaf damage (Doyle et

7000 50 a b 6000 Mean Mean ± 1.96 × SE 40 5000 4000 30

3000 20

2000 % Leaf damage Immatures per kg 10 1000 0 0 8/98 2/99 4/99 6/99 8/99 8/98 2/99 4/99 6/99 8/99 10/98 12/98 10/99 10/98 12/98 10/99 Date Date 7000 50 c d 6000 40 5000 4000 30

3000 20

2000 % Leaf damage Immatures per kg 10 1000 0 0 Jan Jan Oct Oct Apr Apr Feb Feb Sep Sep Mar Mar July July Aug Aug May May June June

Figure 1. Population increases and associated changes in damage for Hydrellia pakistanae reared in closed systems (a and b) and small ponds (c and d). Note the large increases in immatures (a) and associated damage (b) for H. pakistanae reared in 14,000 L tanks for two growing seasons (1998 and 1999) as part of an experiment to assess herbivory/plant competition impacts. Only about 400 immatures were introduced into each tank to produce almost 3500 immatures per kg and 35% leaf damage (after Doyle et al. 2003). Similarly, substantial increases were observed in small pond experimentation again designed to assess herbivory/plant competition impacts during 2002. Note that in one growing season, immature levels (both H. pakistanae and H. balciunasi) reached 5000 per kg (c) with 40% leaf damage (d) (unpublished data).

531 Proceedings of the XI International Symposium on Biological Control of Weeds al. 2002). With 10% to 30% leaf damage, the maximum Decreases in biomass and tuber number have been rate of light saturated photosynthesis was reduced 30% observed in several closed-system experiments. Van et to 40% (Fig. 3). At these damage rates, total daily al. (1998) reported a 30% decrease in biomass with photosynthetic production was estimated to barely levels of herbivory that resulted in complete defolia- balance the daily respiratory needs of the stems. Based tion. Wheeler & Center (2001) achieved levels of about on this information, even relatively low leaf damage 4000 larvae per m2 within small enclosures that can be expected to significantly impact hydrilla growth. resulted in complete defoliation and significant hydrilla

7000 80 Mean ± Mean SE 70 6000 Mean ± 1.96 × SE

60 5000

50 4000 40

3000 %Leaf damage

Immatures per kg 30

2000 20

1000 10

0 0 PINES PINES CHOKE CHOKE CADDO CADDO TEXANA TEXANA MURVAL MURVAL CONROE CONROE CYPRESS CYPRESS RAYBURN RAYBURN PINKSTON PINKSTON SEMINOLE SEMINOLE FLAG LAKE FLAG LAKE LEWISVILLE LEWISVILLE LAKE RAVEN LAKE RAVEN SOMERVILLE SOMERVILLE RIO GRANDE RIO GRANDE PURTIS CREEK PURTIS CREEK TOLEDO BEND TOLEDO BEND GUNTERSVILLE GUNTERSVILLE COLETO CREEK COLETO CREEK LAKE FAIRVIEW LAKE FAIRVIEW NACOGDOCHES NACOGDOCHES Figure 2. Immatures per kg and percentage leaf damage for release and non-release sites surveyed during 1999 (unpublished surveys). Release sites include Lake Seminole, Florida; Coleto Creek Reservoir, Texas; Lewisville Aquatic Ecosystem Research facility (LAERF) ponds; and Guntersville Reservoir, Alabama. Note large variation from site to site.

Model: v1 = (0.647 + c * v2)^0.059 Pmax = ((0.647047) + (0.059096)*% Damage)^–0.40575) 1.4 r = 0.997 1.2

1.0

0.8 per gdw h) 2 0.6 (mg O

max 0.4 P

0.2

0 0 20 40 60 80 100 % Leaf damage

Figure 3. Light-saturated photosynthetic rate (Pmax) for various amounts of hydrilla leaf damage caused by the feeding action of immature Hydrellia pakistanae. Note that maximum rate of light saturated photosynthesis is reduced 30% to 40% for stems having 10% to 30% leaf damage (after Doyle et al. 2002).

532 Insect biocontrol agents of hydrilla impact; i.e. no formation of a surface canopy. However, et al. 2002). Similarly, in larger, long-term tank exper- such high levels of herbivory are not typical of what has iments, biomass and tuber number were decreased 45% been observed in the field. and 21%, respectively, at herbivory levels more typical Experiments designed to assess herbivory impact at of what is observed in the field (Fig. 5). much lower levels, more similar to what is observed at Impacts to field hydrilla infestations are, quite field locations, also show the same results. Mean understandably, harder to assess. However, evidence is hydrilla biomass and tuber number decreased in small, now emerging that indicates that long-term, sustained short-term tank experiments by 17% and 40%, respec- H. pakistanae herbivory can significantly impact tively with only 15% to 40% leaf damage (Fig. 4; Doyle hydrilla infestations in the field. Hydrilla at several

Biomass (g per tank) Tuber number per tank

30 a b

25 c

20 a 15 Number

10 ab b

5

0 0–5% 15–40% 50–75% % Leaf damage category Figure 4. Mean hydrilla biomass (g dry plant material) and tuber number per tank for small, short-term tank experiments measuring herbivory/plant compe- tition impacts to hydrilla. Columns for a specific parameter are not signifi- cantly different at p < 0.10 if followed by the same letter. Note that significant declines in biomass and tubers occurred with increasing leaf damage (after Doyle et al. 2002).

Biomass (g per tank) Tuber number per tank

1400 a 1200 a

1000 b

800 b

Number 600

400

200

0 NO YES Presence of Hydrellia pakistanae

Figure 5. Hydrilla biomass (g dry material per tank) and tuber number per tank over two growing seasons for large (14,000 L), long-term tank experiments designed to assess herbivory/plant competition. Means for a given parameter are signifi- cantly different at p = 0.11 if followed by different letter. Significant decreases in biomass and tuber number occurred; i.e. 3500 immatures per kg and 35% leaf damage (Figs 1a and 1b) (after Doyle et al. 2003).

533 Proceedings of the XI International Symposium on Biological Control of Weeds locations on Lake Seminole, Florida, apparently suffered substantial declines six to seven years after H. pakistanae releases were terminated (Grodowitz et al. 2003). In the summer of 1999, Lake Seminole managers reported significant declines that were possibly related to H. pakistanae feeding action. Surveys conducted in September 1999 showed rela- tively high field populations of about 2500 immatures per kg with leaf damage approaching 16%; higher than what was reported previously. Quantitative surveys in November 1999 showed significant reductions in tubers (three-fold) compared to sites with lower insect impact (Grodowitz et al. 2003). Also, a strong relationship between tuber numbers and leaf damage with decreased number of tubers associated with higher leaf damage were noted (Fig. 6). The ThreeRiv and Wingates sites served as controls. ThreeRiv was far removed from the original releases and typically showed lower insect numbers and the hydrilla at Wingates was recently recovering from a herbicide treatment and thus exhibited low insect numbers. The declines were quite evident. Hydrilla present on Lake Seminole prior to insect impact appeared healthy with little evidence of insect feeding (Fig. 7a). Hydrilla grew consistently as a monoculture and appeared lush Figure 7. Hydrilla on Lake Seminole during 1994 (a) and fully canopied during the peak of the growing and 1999 (b). In 1994, before insect impact, season. However, during 1999, hydrilla was often hydrilla grew in monoculture, but by 1999 it found growing in association with other native plants was part of a mixed aquatic plant bed and and often made up only a small proportion of the total onbviously stressed, coinciding with increased plant community (Fig. 7b). It was noticeably stressed insect populations. with fewer and smaller leaves.

Model: Exponential growth (y = c + exp(b0 + b1*x1 + b2*x2…)) y = 33.1888 + exp(9.25419 + (–1.6497)*x) 160 120 Mean Variance explained = 91.3% 140 Mean ± SE Mean ± 1.96 × SE 100 Wingates 120 Three Riv 2 2 110 80

100

60 80 Tuber number per m Tuber number per m a 60 Camp 40 FoxIsl ab Flats 40 bc SprCrk c c bc 20 20 Three Riv Flats FoxIsl 2 4 6 8 10 12 14 16 18 Wingates SprCrk Camp % Leaf damage Site

Figure 6. Tuber numbers per m2 for sites sampled on Lake Seminole during November 1999. Both ThreeRiv and Wingates sites served as controls, since insect numbers and impact were reduced in comparison to the other sites. Means followed by the same letter are not significantly different at p = 0.05. Also shown is the exponential growth relation- ship between tuber numbers and percentage leaf damage. Lower tuber numbers were associated with sites with higher mean leaf damages (after Grodowitz et al. 2003).

534 Insect biocontrol agents of hydrilla

The following year (2000), more extensive sampling The presence of higher numbers of broken stem pieces was conducted to assess H. pakistanae impact. Insect within the canopy is a common occurrence during times of numbers remained suppressed throughout the year, high H. pakistanae populations. Apparently, the stem which is not surprising after the high numbers and becomes more brittle at the point where several leaves damage exhibited in 1999. Still, decreases in species- within a single whorl are damaged (unpublished data). richness were found related to insect numbers and With increased stem breakage the canopy is opened and damage. Using a three-dimensional linear graphing may lead to the recolonization of native species due to analysis to observe data trends, increases in species- increased light penetration below the hydrilla. On Lake richness (i.e. number of plant species as a measure of Seminole, during September 2000, significant correlations diversity) occurred. Species-richness was highest in (p < 0.05) were observed between species-richness and those samples with high insect numbers and damage; i.e. percentage damaged stems where higher numbers of almost four species for high insect numbers and damage native plant species occurred in those samples (i.e. and only one species, hydrilla, in those samples species-richness) with highest number of stems containing containing minimal insect numbers and damage (Fig. 8). H. pakistanae feeding damage (Fig. 9).

Richness = 1.6578 + 0.0008x + 0.087y

Species richness

% Leaf damage Immatures per kg

Figure 8. Linear relationship between species-richness and Hydrellia pakistanae numbers and leaf damage using a statistical graphing technique to illustrate overall trends.

Richness = –1.9616 + 0.1107(% Damaged stems) 6 r = 0.9393

5

4

3

Species richness 2

1

0 25 30 35 40 45 50 55 60 65 % Damaged stems Figure 9. Species-richness and percentage damaged stems for samples collected from Lake Seminole during September 2000. Means were calculated based on each species-richness category. Corre- lation is significant at p < 0.05.

535 Proceedings of the XI International Symposium on Biological Control of Weeds

Possible regulators by simply moving from the top portion of the canopy to lower levels during the hotter parts of the day where Based on the evidence presented so far, establish- temperature decreases relatively quickly. Hence, ment has been highly successful for H. pakistanae and temperature may not be a very important determinant of minimally successful for H. balciunasi. Range expan- survival and more research is warranted. sion for H. pakistanae has been impressive, with popu- lations located upwards of 500 km away from the Another possibility is the presence of a pupal para- nearest release sites. Hydrellia balciunasi, however, site, Trichopria columbiana Ashmead, commonly has had only limited range expansion. Population found in association with native Hydrellia spp., that is increase for both species is highly variable from field now parasitizing both species of introduced Hydrellia. site to field site with higher numbers more typically While more research is needed, parasitism rates appear observed for H. pakistanae. Increases have been slow to be relatively low for H. pakistanae under field condi- to occur, with some release sites taking from six to eight tions, especially in the early part of the growing season years for H. pakistanae to develop significant popula- and tend to increase roughly proportionally to H. paki- tions. Impact, while impressive under controlled condi- stanae population increases later in the growing season tions (even at relatively low population sizes), is much (Fig. 10). Note that even with 35% parasitism occurring more variable under field conditions. Nonetheless, veri- during October 2001, immature numbers were still fied impact has been observed in at least three sites, high; 6000 immatures per kg. including Sheldon Reservoir and Coleto Creek Reser- Another important determinant for success under voir, Texas, and Lake Seminole, Florida. field conditions is plant nutritional composition, espe- Reasons for limited population increase and impact cially protein content. Wheeler & Center (1996) at field locations, especially in comparison to substan- showed that H. pakistanae larval development was tial increases observed for controlled experimentation, significantly reduced when reared on hydrilla with are unknown but several reasons appear plausible. harder leaf cuticles containing lower nitrogen levels. Temperature is often cited as a possible limiting factor Recent experiments have shown that hydrilla with low since the top portion of the hydrilla canopy during the nitrogen content appears to impact not only larval summer months in the southern US can reach 35°C to development time but the number of eggs oviposited 40°C relatively rapidly. Unpublished information per female (Fig. 11). Eggs per females were over two- suggests high larval mortality occurs after temperatures fold higher for larvae reared on hydrilla containing 2.4- reach 35°C. While this may be an important regulator, fold more protein (as estimated from nitrogen content). it would not be unusual for the larvae to thermoregulate However, nitrogen content is only part of the story,

7000 40

% Parasite emergence 35 6000 Immatures per kg

30 5000 % Parasite emergence 25 4000 20 3000

Immatures per kg 15

2000 10

1000 5

0 0 June July Aug Sep Oct Month

Figure 10. Percentage of parasite emergence from isolated Hydrellia pakistanae pupae collected from small ponds in Lewisville, Texas. Also shown is the corresponding number of immatures per kg. Note that parasitism roughly follows population increases with highest immature number and highest parasitism occurring during the latter part of the growing season (unpub- lished data, C. Snell and M. Grodowitz).

536 Insect biocontrol agents of hydrilla since higher-weight females (typically indicative of tion on Hydrellia population increases and associated having higher egg production) occurred in samples impact on hydrilla. containing higher protein and less crowding as indi- One of the more important determinants of popula- cated by lower fly emergence. More research is needed tion size may be whether or not were released to understand the complexities of nutritional composi- into a specific area. Significantly (p < 0.05) higher

Current effect: F(1, 32)=20.199, p=.00009 Current effect: F(1, 32)=1143.6, p=0.0000 Vertical bars denote 0.95 confidence intervals Vertical bars denote 0.95 confidence intervals 22 22

20 a 20 b

18 18 16 16 14 14 12 12 10 10 8

Eggs per female 8 6 6

4 % PR OTE(dry IN ma basi tter s)

2 4 Used Fertilised Used Fertilised Fertilisation level Fertilisation level

Model: Exponential growth (y = c + exp(b0 + b1x1 + b2x2…)) z = 77.398 + exp(4.5408 + 0.075909x + (–0.11879y))

Variance explained = 48.2%

Female weight µg

% Protein (dry matter basis) Total number emerged

Figure 11. Eggs per female (a) and percentage protein (b) for hydrilla grown in sediment where plants were repeatedly grown in an effort to remove excess nitrogen (Used) and sediment amended with nitrogen (Fertilized; Grodowitz & McFarland 2002). Also, three-dimen- sional surface plot showing relationship between percentage protein and total number of adults emerged versus female Hydrellia pakistanae weight in µg (c).

1400 18 Relase Non release 16 1200

14 1000 12

800 10

600 8

6 400 % Leaf damage Immatures per kg

4

200 2 * * * * 0 0

1998 1999 2000 1998 1999 2000 Year Year Figure 12. Number of immatures of Hydrellia pakistanae per kg and percentage leaf damage for release and non-release sites sampled in 1998, 1999, and 2000. Columns separated by an asterisk are signif- icantly different at p < 0.05 level. Note that higher number of immatures and percentage leaf damage occurred for release sites.

537 Proceedings of the XI International Symposium on Biological Control of Weeds numbers of immatures and associated leaf damage were Doyle, R.D., Grodowitz, M.J., Smart, R.M. & Owens, C. (2002) found at sites where immatures were released previ- Impact of herbivory by Hydrellia pakistanae (Diptera: ously (Fig. 12). This occurred for sites sampled in 1999 Ephydridae) on growth and photosynthetic potential of and 2000. The difference may be quite dramatic, as in Hydrilla verticillata. Biological Control 24, 221–229. 1999 when immatures were 9-fold greater and leaf Doyle, R.D., Grodowitz, M.J., Smart, R.M. & Owens, C. (2003) Separate and interactive effects of competition and damage was 11-fold higher in release versus non- herbivory on the growth, expansion, and tuber formation of release sites. Hydrilla verticillata. Journal of Ecology (in press). It is apparent that additional releases are needed to Freedman, J.E., Grodowitz, M.J., Cofrancesco, A.F. & Bare, R. bolster Hydrellia spp. populations. However, since (2001) Mass-rearing Hydrellia pakistanae Deonier, a populations have remained low in the field, it has been biological control agent of Hydrilla verticillata (L.f.) Royle, next to impossible to collect high enough numbers at for release and establishment, ERDC/EL TR-01-24, US field sites for adequate additional releases. Also, mass- Army Engineer Research and Development Centre, Vicks- rearing under greenhouse conditions, while adequate, is burg. prohibitively expensive with each individual costing Grodowitz, M.J., Center, T.D., Cofrancesco, A.F. & Freedman, upwards of US$0.50 (Freedman et al. 2001). Recently, J.E. (1997) Release and establishment of Hydrellia balciu- a mass-rearing facility developed using small ponds at nasi (Diptera: Ephydridae) for the biological control of the submersed aquatic plant Hydrilla verticillata (Hydrocharita- LAERF has allowed the production of large numbers of ceae) in the United States. Biological Control 9, 15–23. flies at low cost. Over the last two growing seasons, Grodowitz, M.J., Center, T.D. & Snoddy, E. (1995) Current over one million flies were produced and released at status on the use of insect biological control agents for the sites in Texas and Florida at a cost of less than US$0.02 management of hydrilla. In Proceedings, 29th Annual per individual. Such capabilities should allow the Meeting, Aquatic Plant Control Research Program. Miscel- release and subsequent increase in field populations at laneous Paper A-95-3, 134–141, Vicksburg, MS: US Army many hydrilla sites across the US. Engineer Waterways Experiment Station. In summary, it appears that the introduced Hydrellia Grodowitz, M.J., Center, T.D., Snoddy, E. & Dray, F.A. (1994) spp. are capable of severely impacting hydrilla. While Release and establishment of insect biocontrol agents for the th more research is needed to understand possible popula- management of hydrilla. Proceedings, 28 Annual Meeting, tion regulators, the use of these agents should be Aquatic Plant Control Research Program, Miscellaneous considered operational and used as part of any hydrilla Paper A-94-2, 181–201, Vicksburg, US Army Engineer Waterways Experiment Station. management program. Grodowitz, M.J., Cofrancesco, A.F., Stewart, R.M. & Madsen, J. (2003) Possible Impact of Lake Seminole Hydrilla by the Acknowledgements Introduced Leaf-Mining Fly Hydrellia pakistanae. Technical report, US Army Engineers Research and Development The data presented herein, unless otherwise noted, were Center, Vicksburg, MS. obtained from research funded by the US Army Engi- Grodowitz, M.J., Freedman, J.E., Cofrancesco, A.F. & Center, neer Research and Development Centre, Aquatic Plant T.D. (1999) Status of Hydrellia spp. (Diptera: Ephydridae) Control Research Program. Permission was granted by release sites in Texas as of December 1998. Miscellaneous the Chief of Engineers to publish this information. Paper A-99-1, US Army Engineer Research and Develop- Many individuals contributed to the ideas and informa- ment Center, Vicksburg. tion summarized in this paper. These include Dr M. Grodowitz, M.J., Freedman, J.E., Jones, H., Jeffers, L., Lopez, Smart, Ms C. Owens, Ms R. Bare, and C. Snell of the C. & Nibling, F. (2000) Status of waterhyacinth/hydrilla infestations and associated biological control agents in LAERF; Dr R. Doyle (Baylor University); and Dr A. lower Rio Grande valley cooperating irrigation Cofrancesco, Ms J. Freedman and Mr H. Jones of the districts.ERDC/EL SR-00-11, US Army Engineer Research ERDC. I would also like to thank Dr J. Shearer and Ms and Development Center, Vicksburg. Sherry Whitaker for their critical review of the manu- Van, T.K., Wheeler, G.S. & Center, T.D. (1999) Competition script (ERDC). between Hydrilla verticillata and Vallisneria americana as influenced by soil and fertility. Aquatic Botany 62, 225–233. References Wheeler, G.S. & Center, T.D. (1996) The influence of hydrilla leaf quality on larval growth and development for the Center, T.D., Grodowitz, M.J., Jubinsky, A.F., Snoddy, E. & biological control agent Hydrellia pakistanae (Diptera: Freedman, J.E. (1997) Establishment of Hydrellia pakis- Ephydridae). Biological Control 7, 1–9. tanae (Diptera: Ephydridae) for the biological control of the Wheeler G.S. & Center T.D. (2001) Impact of the biological submersed aquatic plant Hydrilla verticillata (Hydrocharita- control agent Hydrellia pakistanae (Diptera: Ephydridae) on ceae) in the southeastern United States. Biological Control the submersed aquatic weed Hydrilla verticillata (Hydro- 8, 65–73. charitaceae). Biological Control 21, 168–181.

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