Monitoring the rust fungus, Puccinia jaceae var. solstitialis, for biological control of yellow starthistle (Centaurea solstitialis)

A.J. Fisher,1 D.M. Woods,2 L. Smith1 and W.L. Bruckart3

Summary Yellow starthistle, Centaurea solstitialis L., is a noxious weed that infests more than 7 million ha of rangeland in California. The rust fungus, Puccinia jaceae Otth var. solstitialis, was first released as a classical biological control for yellow starthistle in California in 2003. In 2005, a research program was initiated to monitor the life cycle and spread of P. jaceae solstitialis. The rust was released at two sites representing different climatic zones, the coastal hills and Central Valley, in January 2005 and 2006. Releases resulted in infected plants at both sites in both years. Natural urediniospore (infective spore) reinfection occurred throughout yellow starthistle’s growing season at the Central Valley site, but infection did not persist at the coastal hills site. P. jaceae solstitialis spread at least 100 m in 2005 at the Central Valley site but did not spread at the coastal hills site. The results of this study show that the spread of the rust is most concentrated in areas closest to release sites. Teliospores (dormant spores) were produced during plant senescence at both sites in 2005. Our results suggest that P. jaceae solstitialis is likely to establish and spread to new yellow starthistle populations in the Central Valley but not in the coastal hills, near Napa California.

Keywords: reinfection, establishment, spread.

Introduction ture. Scientists have introduced six insect biological control agents, all of which attack flower heads and Yellow starthistle (Centaurea solstitialis L., Astera- destroy developing seeds (Turner et al., 1995; Pitcairn ceae) is an invasive alien weed that infests more than et al., 2004). However, these have not reduced yellow 7 million ha of rangeland in California (Pitcairn et al., starthistle populations to acceptable levels (Balciunas 2006). Yellow starthistle displaces desirable plants in and Villegas, 1999; Woods et al., 2004). both natural and agricultural areas. Its spiny flowers In 2003, the rust fungus, Puccinia jaceae Otth var. deter feeding by grazing animals and lower the value solstitialis, was introduced to California for the biolog- of recreational lands (Sheley et al., 1999). Yellow ical control of yellow starthistle (Woods et al., 2003). starthistle is a winter annual adapted to the mild wet This is the first exotic plant pathogen to be approved for winters and dry summers of the Mediterranean (Mad- release for the classical biological control of a weed in dox, 1981). Seeds usually germinate soon after the be- the continental USA using the modern permitting proc- ginning of fall rains, rosettes develop slowly during the ess required by the Animal and Plant Health Inspection winter and plants bolt in late spring and flower con- Service (APHIS; Bruckart et al., 1999). tinuously until the plant senesces from lack of mois- P. jaceae var. solstitialis is macrocyclic (its life cy- cle includes five spore stages), and it conducts its entire life cycle on yellow starthistle (Savile, 1970). Plants infected during the growing season produce pustules 1 USDA, ARS, Exotic and Invasive Weeds Research Unit, Albany, CA 94710, USA. that release urediniospores. Urediniospores disperse 2 California Department of Food and Agriculture, Sacramento, CA aerially to infect other leaves and plants, resulting in 95832, USA. multiple generations within a growing season that may 3 USDA, ARS, Foreign Disease-Weed Science Research Unit, Fort Det- progressively increase the incidence, intensity and spa- rick, MD 21702, USA. Corresponding author: A.J. Fisher < [email protected] >. tial spread of infection. The latent period (time from © CAB International 2008 infection to spore formation) ranges from 10 to 15 days

540 Monitoring the rust fungus, Puccinia jaceae var. solstitialis, for biological control of yellow starthistle at 25°C to 15°C, respectively (Bennet et al., 1991). rectangular wall of plastic sheeting was placed around Many rust fungi produce teliospores, dormant spores, each plot to prevent drift during application. Plots were during plant senescence. Teliospores germinate in the evaluated 30, 60, 90 and 120 days after inoculations presence of a host plant to produce basidiospores during and scored either as positive or negative for disease the wet season. To complete the life cycle, basid- symptoms. iospores are thought to infect yellow starthistle seed- In 2006, the experiment was repeated on a smaller lings to produce pycnia. Pycnia have been observed scale, and six additional 0.5 ´ 0.5 m plots were in- on yellow starthistle seedlings in California at a site stalled. Plots were inoculated on the same dates, using where P. jaceae solstitialis was released the previous the same methods, as in 2005. year (Fisher et al., 2006). Pycnia produce aecia, which produce urediniospores to continue the infection cycle Teliospore emergence (Savile, 1970). In 2005, a research program was initiated to monitor In addition to the 12 plots above, a separate 1-m2 plot the life cycle and spread of P. jaceae solstitialis in Cali- was inoculated at both sites, with 200 ml of deionized fornia. The goals of this study were to: (1) monitor ure- water containing 50-mg spores and 0.15% Tween 20, to diniospore natural reinfection over the yellow starthistle monitor teliospore emergence in 2005. Plots were rein- growing season, (2) monitor teliospore emergence and oculated once a month from February to June, if need- (3) monitor the spread of P. jaceae solstitialis at two ed, to maintain infection. Once a month, from March release sites in California. to September, five infected leaves were harvested from each site. In the lab, spores were scraped off leaves into 0.1% water agar, and the number of urediniospores and Materials and methods teliospores in a 200-spore sample from each leaf was counted using a compound microscope. The Proc GLM Fungal isolate and site characteristics procedure was used to evaluate the effect of location (Napa or Woodland) on teliospore production. Analy- P. jaceae var. solstitialis field isolate FDWSRU 84- ses were carried out using SAS Institute software ver- 71 was collected by S. S. Rosenthal in 1984 east of sion 9.1 (SAS Institute, 2000). Yarhisar and Hafik (near Šivas), Turkey. This isolate was used for host-specificity testing and was the isolate released from quarantine (Bruckart, 1989; Bruckart Monitoring P. jaceae solstitialis spread et al., 1999). Urediniospores for the present study were In a separate study to develop an optimal strategy propagated at the California Department of Food and to release P. jaceae solstitialis (Fisher et al., 2007), Agriculture, Sacramento, CA, using the methods de- 9 g of urediniospores were used to inoculate yellow scribed by Woods and Popescu (2004). starthistle at the two sites described above. Plots of Permanent experimental plots were established in yellow starthistle were inoculated up to five times January 2005 and 2006 at sites near Napa and Wood- from January to June. At the first inoculation, plants land, CA. The Napa site is an ungrazed rangeland in the were at the rosette stage, and during the last inocula- coastal hills near Napa, Napa County, 427-m elevation, tion, plants were beginning to flower. To document dominated by yellow starthistle and exotic European spread, permanent 0.5 ´ 2 m plots were installed 20, annual grasses, and surrounded by oak and manza- 40, 60, 80 and 100 m from inoculated plots at both nita forest. The Woodland site is an ungrazed range- sites in May 2005, positioned along four compass land, near Woodland, Yolo County, 5-m elevation. It directions. Because of yellow starthistle patchiness is dominated by exotic European annual grasses, sur- and site boundaries, there were a total of seven moni- rounded by actively grazed rangeland and agriculture. toring plots 20 m from release plots, five monitoring The Woodland site is in the Central Valley, which has plots 40 and 60 m from release plots, four monitoring a hot summer, and the Napa site is in the coastal hills, plots 80 m from release plots and six monitoring plots which is cooler. 100 m from release plots. In May 2005, we visually inspected plants within a metre on all four sides of Urediniospore natural reinfection over each of the six plots inoculated in January to observe natural urediniospore spread. In June 2005, we visu- a single growing season ally inspected all monitoring plots 20, 40, 60, 80 and In 2005, six permanent 1 ´ 0.5 m plots marked by 100 m from inoculated plots and recorded the number wooden stakes were installed at each site. In late Janu- of infected plants out of a maximum of 50 plants per ary 2005, each plot was uniformly sprayed with 200 ml plot. To determine if the rust fungus had spread be- of deionized water containing 50 mg urediniospores yond 100 m in July 2005, roadside yellow starthistle and 0.15% Tween 20 (=100 mg spores/m2) using plas- populations were inspected for P. jaceae solstitialis tic 250-ml finger pump spray bottles. This was suffi- infection at the Central Valley site up to a distance of cient to wet all plants in the plot to runoff. A portable approximately 5 km.

541 XII International Symposium on Biological Control of Weeds

Results and discussion would allow us to determine if and when these criteria are met in the field. P. jaceae var. solstitialis pustules emerged on yellow As the season progressed, temperatures rose and yel- starthistle at both sites in 2005 and 2006 within 30 days low starthistle senesced; P. jaceae solstitialis produced after inoculations (Fig. 1). In the coastal hills (Napa more teliospores (dormant spores that germinate the site) in 2005, rust pustules were present in all plots next winter) as a proportion of all spores. Production of after 60 days but declined to three of six plots after 90 teliospores gradually increased at both sites in August days and then to two of six plots after 120 days. In 2006 and September 2005 (Fig. 2). A higher percentage of at the Napa site, there was no natural reinfection at spores sampled in the Central Valley were teliospores plots inoculated in 2005, and plots inoculated in 2006 (P < 0.001) compared to the coastal hills. had pustules only at 30 days after inoculation. All six The rust spread up to 100 m at the Woodland site in plots in the Central Valley (Woodland site) contained 2005 (Fig. 3). In May of 2005, all of the plots inocu- infected plants at each sampling date in both years. It is lated in January had pustules at least 1 m outside of not clear why P. jaceae solstitialis did not consistently plots at the Woodland site. By July, the rust had spread persist in release plots at the Napa site. To identify fac- to one of the five monitoring plots 100 m from the near- tors that limit the success of the pathogen, it will be est release plot. Of the monitoring plots, the highest necessary to monitor local climate conditions. Bennett percentage with rust, aside from the metre surrounding et al. (1991) estimated that optimal P. jaceae solstitialis plots, occurred at 60 m (Fig. 3). It is not clear why a infection occurs at 15–20°C with dew periods between greater percentage of plots had rust at 60 m compared 8 and 16 h. Data on dew periods at these locations to 20 or 40 m. When we compare the percentage of

Figure 1. Occurance of Puccinia jaceae var. solstitialis pustules indicating recent infection of yellow starthistle plants in permanent plots that were inocu- lated in late January in 2005 and 2006.

542 Monitoring the rust fungus, Puccinia jaceae var. solstitialis, for biological control of yellow starthistle

Figure 2. Natural production of Puccinia jaceae var. solstitialis teliospores (dormant spores), as a proportion of all spores (means ± SE), at the Napa and Woodland, CA, sites in 2005 after inoculations in release plots start- ing in January 2005.

plants in each plot with at least one pustule, 26% of the Evans, released for mist flower,Ageratina riparia (Re- plants in plots 20 m from release plots were infected, gel) King and Robinson, spread 80 km over water in 2 compared with 2% in 40-m plots, 6% in 60-m plots, 2% years (Barton et al., 2007). in 80-m plots and 2% in 100-m plots. These data show The year after plots were inoculated, P. jaceae sol- that the spread of the rust is most concentrated in areas stitialis naturally reinfected yellow starthistle at the site closest to the release plots. No rust fungus was found in the Central Valley but not in the coastal hills (Fisher during visual inspection of roadside yellow starthistle et al., 2007). For P. jaceae solstitialis to reinfect yel- populations surrounding the release site (up to 5 km low starthistle after a dormant season, it must produce away). Spread was modest compared to other success- teliospores that persist and germinate in the winter to ful biological control pathosystems, for example the complete the life cycle. Teliospores were observed in white smut fungus, Entyloma ageratinae Barreto and the summer, and pycnia were observed in release plots

Figure 3. Spatial spread of Puccinia jaceae var. solstitialis, as represented by pustules on yellow starthistle plants during one growing season in Woodland, CA, in 2005.

543 XII International Symposium on Biological Control of Weeds the following February (Fisher et al., 2006). Soon after Fisher, A.J., Woods, D.M., Smith, L. and Bruckart, W.L. pycnia were observed, in March, urediniospores were (2007) Developing an optimal release strategy for the rust identified in the 2005 release plots. fungus Puccinia jaceae var. solstitialis for biological con- In general, the rust fungus was more productive in trol of Centaurea solstitialis (yellow starthistle). Biologi- the Central Valley (Woodland site) than the coastal hills cal Control 42, 161–171. Maddox, D. M. (1981) Introduction, phenology, and density (Napa site), with higher rates of reinfection, both within of yellow starthistle in coastal, intercoastal, and central a single growing season and after a dormant season. valley situations in California. US Department of Agri- In addition, P. jaceae solstitialis spread at least 100 m culture, Agricultural Research Results ARR-W-20. Agri- during the first year at the Central Valley site. In the cultural Research Service, Oakland, CA, USA, 33 pp. coastal hills, yellow starthistle exhibited disease symp- Pitcairn, M.J., Piper, G.L. and Coombs, E.M. (2004) Yellow toms soon after inoculation, but infection decreased starthistle. In: Coombs, E.M., Clark, J.K., Piper, G.L. and over time and did not return the following year. These Cofrancesco, A.F., Jr. (eds) Biological Control of Inva- results suggest that this isolate of P. jaceae solstitia- sive Plants in the United States. Oregon State University lis should establish well in California’s Central Valley Press, Corvallis, USA, pp. 421–435. but not at sites with climate conditions similar to the Pitcairn, M.J., Schoenig, S., Yacoub, R. and Gendron, D. coastal hills. (2006) Yellow starthistle continues to spread in Califor- nia. California Agriculture 60, 83–90. SAS Institute (2000) SAS Statistics User’s Guide. SAS Insti- Acknowledgements tute, Cary, NC, USA. 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