Effects of milfoil weevils and weather on the control of Eurasian watermilfoil

We examined the response of Eurasian watermilfoil (Myriophyllum spicatum) coverage on Manistee Lake, Michigan (U.S.A.) in the presence of milfoil weevils (Euhrychiopsis lecontei). Among 150 sites, milfoil presence declined from 2008 levels of 34 (23%) sites to 2 (1%) sites by 2015 coincident with cumulative stocking of 259,500 weevils from 2007 to 2014. Severe winter temperatures also were associated with milfoil declines. Each 1°C decline in average low temperature during the preceding winter was associated with 3.4 (95% CI 0.8–6.1) fewer sites with milfoil. Impacts of weevil herbivory on watermilfoil may be accentuated by severe winter temperatures. Lake managers should, when possible, integrate weather conditions with weevil stocking regimes to control Eurasian watermilfoil.     

Oviposition specificity and behavior of the watermilfoil specialist Euhrychiopsis lecontei

Eurasian watermilfoil (Myriophyllum spicatum L.) is a nuisance aquatic weed, exotic to North America. The freshwater weevil Euhrychiopsis lecontei (Dietz) is a potential control agent of Eurasian watermilfoil and is a fully submersed aquatic specialist herbivore. Its presumed original host is the native northern watermilfoil (Myriophyllum sibiricum Komarov). We conducted a set of oviposition experiments to reveal first and second oviposition preference of Euhrychiopsis lecontei when presented with seven macrophytes. We tested differences between source (lake) populations of weevils, differences in behavior between weevils reared on the exotic Eurasian watermilfoil and the native northern watermilfoil and between weevils in the presence and absence of their preferred hostplant. Oviposition assays confirmed that E. lecontei is a watermilfoil specialist. Out of the 207 females that laid eggs, only three oviposited on a non-watermilfoil plant, Megalodonta beckii. The weevils' degree of specificity was influenced by the watermilfoil species on which they were reared. Weevils reared on Eurasian watermilfoil tended to oviposit on Eurasian watermilfoil, spent more time on Eurasian watermilfoil than on other plants, and spent more time off plants and took longer to oviposit when Eurasian watermilfoil was removed. Weevils reared on northern watermilfoil did not exhibit a preference for either watermilfoil species in oviposition or in time allocation, although they oviposited on and spent significantly more time on watermilfoils than on other species. Rearing of the two populations on their complementary watermilfoil hostplant resulted in responses typical of the rearing plant, not the original host. These results show that although both weevil populations are watermilfoil specialists, Eurasian-reared weevils prefer Eurasian watermilfoil in general host attraction and oviposition, whereas northern-reared weevils do not. The results support the contention that E. lecontei may be a good biocontrol agent for Eurasian watermilfoil because of its high specificity. The results also suggest that the current host range expansion of the weevil to Eurasian watermilfoil has the potential to become a host shift due to the increased specificity. Herbivory in freshwater systems is not well studied, and the E. lecontei-M. spicatum relationship is a rare example of submersed freshwater specialist herbivore-host-plant interactions.     

Gas chromatographic determination of flurprimidol in a submersed aquatic plant (Myriophyllum spicatum), soil,and water

Methods for the extraction and quantification of flurprimidol residues in Eurasian watermilfoil (Myriophyllum spicatum), soil, and water are described. The compound was detected and quantified by gas chromatography (GC) with a thermionic specific detector. Its identity was confirmed by gas chromatography-mass spectrometry (GCMS) with detection at m/e 40–320. Recoveries from samples spiked with flurprimidol at 10–10,000 ng ml^?1 or g^?1 averaged 86.8% for Eurasian watermilfoil shoots, 85.2% for roots, 79.3% for loam soil, and 93.3% for water. In a small-scale experiment under field conditions, approximately 88% of the applied flurprimidol dissipated in 4 weeks. The majority of recovered flurprimidol was found in the water and upper 5 cm soil layer. The half-life of the compound in water was 6.8–8 days during June/July 1989.     

Gas chromatographic determination of flurprimidol in a submersed aquatic plant (Myriophyllum spicatum), soil, and water

Methods for the extraction and quantification of flurprimidol residues in Eurasian watermilfoil (Myriophyllum spicatum), soil, and water are described. The compound was detected and quantified by gas chromatography (GC) with a thermionic specific detector. Its identity was confirmed by gas chromatography-mass spectrometry (GCMS) with detection at m/e 40–320. Recoveries from samples spiked with flurprimidol at 10–10,000 ng ml^–1 or g^–1 averaged 86.8% for Eurasian watermilfoil shoots, 85.2% for roots, 79.3% for loam soil, and 93.3% for water. In a small-scale experiment under field conditions, approximately 88% of the applied flurprimidol dissipated in 4 weeks. The majority of recovered flurprimidol was found in the water and upper 5 cm soil layer. The half-life of the compound in water was 6.8–8 days during June/July 1989.     

Vision is Important for Plant Location by the Phytophagous Aquatic Specialist Euhrychiopsis lecontei Dietz (Coleoptera: Curculionidae)

The aquatic milfoil weevil Euhrychiopsis lecontei Dietz (Coleoptera: Curculionidae) is a specialist on Myriophyllum spp. and is used as a biological control agent for Eurasian watermilfoil (Myriophyllum spicatum L.), an invasive aquatic macrophyte. We show evidence that visual cues are important for plant detection by these weevils. Weevils had difficulty locating plants in dark conditions and were highly attracted to plant stems in the light, even when the plant sample was sealed in a vial. However, weevils were equally attracted to both M. spicatum and another aquatic macrophyte, coontail (Ceratophyllum demersum L.) in vials. Turbidity (0–100 NTU) did not significantly influence visual plant detection by the weevils. This work fills a void in the literature regarding visual plant location by aquatic specialists and may help lead to a better understanding of when and where these weevils will find, accept, and damage their target host-plants.     

Species distribution models for invasive Eurasian watermilfoil highlight the importance of data quality and limitations of discrimination accuracy metrics

AimAvailability of uniformly collected presence, absence, and abundance data remains a key challenge in species distribution modeling (SDM). For invasive species, abundance and impacts are highly variable across landscapes, and quality occurrence and abundance data are critical for predicting locations at high risk for invasion and impacts, respectively. We leverage a large aquatic vegetation dataset comprising point‐level survey data that includes information on the invasive plant Myriophyllum spicatum (Eurasian watermilfoil) to: (a) develop SDMs to predict invasion and impact from environmental variables based on presence–absence, presence‐only, and abundance data, and (b) compare evaluation metrics based on functional and discrimination accuracy for presence–absence and presence‐only SDMs.LocationMinnesota, USA.MethodsEurasian watermilfoil presence–absence and abundance information were gathered from 468 surveyed lakes, and 801 unsurveyed lakes were leveraged as pseudoabsences for presence‐only models. A Random Forest algorithm was used to model the distribution and abundance of Eurasian watermilfoil as a function of lake‐specific predictors, both with and without a spatial autocovariate. Occurrence‐based SDMs were evaluated using conventional discrimination accuracy metrics and functional accuracy metrics assessing correlation between predicted suitability and observed abundance.ResultsWater temperature degree days and maximum lake depth were two leading predictors influencing both invasion risk and abundance, but they were relatively less important for predicting abundance than other water quality measures. Road density was a strong predictor of Eurasian watermilfoil invasion risk but not abundance. Model evaluations highlighted significant differences: Presence–absence models had high functional accuracy despite low discrimination accuracy, whereas presence‐only models showed the opposite pattern.Main conclusionComplementing presence–absence data with abundance information offers a richer understanding of invasive Eurasian watermilfoil''s ecological niche and enables evaluation of the model''s functional accuracy. Conventional discrimination accuracy measures were misleading when models were developed using pseudoabsences. We thus caution against the overuse of presence‐only models and suggest directing more effort toward systematic monitoring programs that yield high‐quality data.     

Effects of eutrophication and snails on Eurasian watermilfoil (Myriophyllum spicatum) invasion

Exotic species can invade and establish new habitats both as a result of their own traits, and as a result of the characteristics of the environment they invade. Here, we show that the abundance of the invasive submerged aquatic plant, Myriophyllum spicatum (Eurasian watermilfoil) is highly dependent on the conditions of the environment in a mesocosm experiment. M. spicatum is allelopathic towards epiphytic algae, and in the absence of algivorous snails, we found that the abundance of both algae and M. spicatum significantly increased with experimentally increased nutrient loading, while the abundance of native submerged macrophytes declined. However, when snails were present, snail biomass increased with increasing nutrient loading, and M. spicatum biomass was consistently low while native submerged macrophyte biomass was consistently high. Our results stress the importance of the interaction between species traits and environmental conditions when considering the invasiveness of certain exotic species and the invasibility of certain environments.     

Geographic distribution and genotypic composition of invasive hybrid watermilfoil (<Emphasis Type="Italic">Myriophyllum spicatum</Emphasis> × <Emphasis Type="Italic">M. sibiricum</Emphasis>) populations in North America

The recent recognition of invasive hybrid watermilfoil (Myriophyllum spicatum × M. sibiricum) in North America has necessitated a more thorough evaluation of its overall distribution and occurrence in natural populations. A comprehensive survey of watermilfoil populations was conducted in five Minnesota lakes, three of which were suspected a priori to contain hybrid watermilfoil. DNA sequence data verified that hybrid plants between the nonindigenous M. spicatum L. and indigenous M. sibiricum Kom. occurred in three of the five lakes sampled. Myriophyllum spicatum was not detected in lakes where hybrids were prevalent. Further sampling of lakes in Idaho, Michigan, Minnesota, Wisconsin and Washington identified 30 additional hybrid watermilfoil populations. In only three of these populations the hybrid watermilfoil was found to co-occur with M. spicatum. To facilitate the field identification of the two parental species and their hybrid, morphological data from watermilfoil specimens collected across the United States were evaluated. We determined that leaf segment/leaf length measurements can effectively distinguish M. spicatum and M. sibiricum; however, hybrids are intermediate for these characters and such measurements frequently overlap with respect to their parental taxa. By incorporating a combined molecular and morphological approach to identifying watermilfoils, the hybrids can be identified readily and their distributions elucidated both within and between lakes. Because hybrids may respond differently to local ecological conditions than their parents, information on their presence and distribution should be of particular importance to management and conservation programs.     

Photosynthesis in Eurasian Watermilfoil (Myriophyllum spicatum L.)

Gas exchange of Eurasian watermilfoil (Myriophyllum spicatum L.) indicated a near-zero CO₂ compensation point and a high temperature optimum for photosynthesis. These properties are characteristic of plants fixing CO₂ by a β-carboxylation mechanism. Operation of the Calvin cycle was shown and no evidence for β-carboxylation was obtained. These results indicate that near-zero CO₂ compensation points are not dependent on a β-carboxylation mechanism.     

Hybrid watermilfoil (Myriophyllum spicatum × Myriophyllum sibiricum) exhibits traits associated with greater invasiveness than its introduced and native parental taxa

Hybridization has been associated with increased invasiveness in plants. In North America, the hybrid aquatic plant Myriophyllum spicatum?×?Myriophyllum sibiricum (hybrid watermilfoil, hereafter HWM) is a cross between non-native invasive Eurasian watermilfoil (M. spicatum, EWM) and native northern watermilfoil (M. sibiricum, NWM). Lab-based trials have demonstrated higher growth rates in HWM compared to EWM and NWM, but these patterns have not been systematically examined in the field. In this study, we compared the invasiveness of HWM to its parental taxa, EWM and NWM, by examining the amount and timing of: (1) flowering, (2) surface cover, and (3) biomass (using stem counts as a proxy). We conducted repeat surveys of Myriophyllum beds at eight lakes (2–3 lakes/taxon) in the Minneapolis–St. Paul Metropolitan area (Minnesota, USA) between June 2017 and November 2018. HWM produced more flower spikes earlier and overall, and maintained consistently more flower spikes throughout the growing season than EWM and NWM. In addition, surface cover reached greater annual peaks and was higher for longer throughout the growing season for HWM than for both parental taxa. We did not observe a significant difference in stem counts among the three taxa, but HWM did reach a higher maximum number of stems than either parental taxon. This study provides field-based evidence of increased invasiveness associated with hybridization between EWM and NWM; specifically, greater reproductive potential via flowering and greater surface cover may increase HWM spread, have greater impacts on native species, and pose more of a nuisance to lake users.

     

Experimental estimation of the possible use of submersed macrophytes for biotesting bottom sediments of the Yenisei River

The laboratory testing of bottom sediments (BSs) from the Yenisei River containing different concentrations of technogenic radionuclides, heavy metals, and biogenic elements (N and P) based on aquatic such plants as Elodea canadensis (Canadian waterweed) and Myriophyllum spicatum (Eurasian watermilfoil) has revealed a higher sensitivity of roots to the general quality of BSs than shoots: shoot length (9%) < root length (11%) < root number (15%) in M. spicatum; shoot length (22%) < root length (42%) < root number (44%) in E. canadensis. In contrast to M. spicatum, the growth parameters of roots and shoots in E. canadensis have differed in a significant statistical manner between most BS samples. A reverse correlation has been found between the increase in shoot length and the activity of technogenic radionuclides in BSs, which is mostly significant in E. canadensis (r ² = 0.90–0.95, p = 0.05). Since the growth of shoots and roots in E. canadensis has turned out to be more sensitive to changes in the quality of BSs than that in M. spicatum, E. canadensis can be considered more prospective for biotesting BSs.     

Glycolate Metabolism in Eurasian Watermilfoil (Myriophyllum spicatum)

The metabolism of glycolate by Eurasian watermilfoil (Myriophyllum spicatum L.), a submersed angiosperm, was studied by feeding radioactive glycolate and glyoxylate and by analysis of glycolate and glycolic acid oxidase. Evidence for operation of the glycolate pathway is given. Glycolic acid oxidase occurs at levels comparable to amounts in species showing photorespiration. This species has a high affinity for CO₂ and a possible mechanism for it is described.     

Visual Active Space of the Milfoil Weevil, <Emphasis Type="Italic">Euhrychiopsis lecontei</Emphasis> Dietz (Coleoptera: Curculionidae)

Euhrychiopsis lecontei Dietz (Coleoptera: Curculionidae), a native weevil, is used as a biological control agent for the invasive aquatic macrophyte, Eurasian watermilfoil (Myriophyllum spicatum L.). Because E. lecontei overwinters on land in the adult stage and must find plants in lakes each spring, plant finding behaviors are essential to eventually understanding and predicting long term biological control. Our research showed that E. lecontei is visually attracted to M. spicatum at up to 17.5 cm, and is more attracted to plants than other visual stimuli within 15 cm. We also showed that turbidity may affect visual plant finding at 15 cm. Using available data from this and other previous studies involving chemical cues and other life history traits, we propose a testable conceptual model for how E. lecontei finds plants each year, especially while underwater. This model may also be used to explain plant finding by aquatic phytophagous insects in general.     

Experimental Tests of Priority Effects and Light Availability on Relative Performance of Myriophyllum spicatum and Elodea nuttallii Propagules in Artificial Stream Channels

Submersed macrophytes have important ecological functions in many streams, but fostering growth of beneficial native species while suppressing weedy invasives may be challenging. Two approaches commonly used in management of terrestrial plant communities may be useful in this context: (1) altering resource availability and (2) establishing desirable species before weeds can invade (priority effects). However, these approaches are rarely used in aquatic systems, despite widespread need for sustainable solutions to aquatic weed problems. In artificial stream channels in California, USA, I conducted experiments with asexual propagules of non-native invasive Myriophyllum spicatum (Eurasian watermilfoil) and native Elodea nuttallii (western waterweed) to address the questions: (1) How does light availability affect relative performance of the two species?; (2) Does planting the native earlier than the invasive decrease survival or growth rate of the invasive?; and (3) Do light level and priority effects interact? The relative performance between E. nuttallii and M. spicatum had an interesting and unexpected pattern: M. spicatum had higher growth rates than E. nuttallii in the zero and medium shade levels, but had similar performance in the low and high shade levels. This pattern is most likely the result of E. nutallii’s sensitivity to both very low and very high light, and M. spicatum’s sensitivity to very low light only. Native priority did not significantly affect growth rate or survival of M. spicatum, possibly because of unexpectedly poor growth of the E. nuttallii planted early. This study suggests that altering light levels could be effective in reducing growth of an invasive macrophyte, and for changing the competitive balance between a native and a non-native species in the establishment phase. Further investigations into the use of priority effects and resource alteration for submersed macrophyte management are warranted, given their mixed results in other (limited) studies.     

Variation in hostplant preference and performance by the milfoil weevil, Euhrychiopsis lecontei Dietz, exposed to native and exotic watermilfoils

Exotic plants often displace native plants and thus alter the availability of native hostplants for specialist herbivorous insects. The submersed aquatic weevil Euhrychiopsis lecontei Dietz is endemic to North America, but there are now source populations on the exotic Eurasian watermilfoil (Myriophyllum spicatum L.) as well as on the weevil's ancestral host, northern watermilfoil (Myriophyllum sibiricum Komarov). This provides an opportunity to examine a host range expansion in progress. To further define the host range of the weevil and to determine how population source and rearing plant influence host plant preference and performance, we conducted a series of preference and rearing experiments with weevils from two source populations reared on northern milfoil, on Eurasian milfoil, switched late in larval development from northern to Eurasian milfoil, and vice versa. We also included two rearing treatments with milfoils on which the weevil has not been documented: the native M. verticillatum L. and the exotic M. aquaticum Verd. Preference by weevils in the switched rearing treatments was similar to preferences exhibited by weevils reared solely on the second (later) milfoil species and an increase in preference for Eurasian milfoil was induced by adult exposure to Eurasian milfoil for 2 weeks. In contrast, sizes and development times of weevils in the switched rearing treatments were similar to sizes and development times exhibited by weevils reared solely on the first (early) milfoil species. These results indicate that preference by the milfoil weevil is determined late in larval development or later and Hopkins' host selection principle is not supported. However, size and development time were most affected by hostplant quality during larval development when larvae must acquire the resources needed for pupation. Oviposition preference in the milfoil weevil was a population attribute, not a fixed individual attribute and there was no significant variation in preference among individuals reared on northern milfoil, but significant variation in preference was detected among weevils reared on Eurasian milfoil. Weevils oviposited on all four milfoil species and completed development on three of them, but did not develop beyond the larval stage on M. aquaticum. Weevils reared on Eurasian milfoil developed faster and reached larger adult sizes than weevils in any other rearing treatment. The smallest sizes and longest development times were for weevils reared on the natives, northern milfoil and M. verticillatum. The milfoil weevil oviposits on an array of milfoil species and is unable to distinguish an unsuitable host (M. aquaticum) within this genus. The influence of rearing plant and adult exposure to Eurasian milfoil on hostplant preference suggests that host range expansion to novel congeners may occur more rapidly than predicted by models which assume that genetic variation is required. Significant variation among individuals in hostplant preference suggests the potential for a host shift to a plant for which E. lecontei appears pre-adapted.     

Interactions between invasive Eurasian watermilfoil and native water stargrass in Cayuga Lake,NY, USA

Aims Eurasian watermilfoil (Myriophyllum spicatum L.) is a common invasive plant in American lakes and has many negative impacts on invaded ecosystems. Drastic decline of this plant at the northern end of Cayuga Lake in the New York State has occurred since the 1980s, with a much smaller magnitude of decline of Eurasian watermilfoil at the southern end (Johnson et al. 2000) During the same period, increases in the abundance of native plants, particularly water stargrass (Heteranthera dubia (Jacq.) MacMill.), have been observed (Johnson et al. (1998)) We aimed to investigate the mechanisms responsible for the decline of Eurasian watermilfoil and evaluate the responses of co-occurring plants at the two ends of Cayuga Lake over time. We hypothesized that plant interactions might have contributed to the drastic decline of Eurasian watermilfoil, particularly allelopathy by native water stargrass.Methods A lake survey was conducted to assess distribution and abundance of plant communities at the northern end and the southern end of Cayuga Lake. Additionally, two sets of greenhouse experiments were conducted to investigate the interactions between invasive Eurasian watermilfoil and native water stargrass. A competition experiment evaluated intra- versus inter-specific competition among plants grown together; an allelopathy experiment examined the responses of plants to each other's extracts.Important findings The lake survey showed that water stargrass was extremely abundant at the northern end, whereas Eurasian watermilfoil was sparse at the northern end but one of the most common species at the southern end. The survey also revealed that water stargrass was more abundant than Eurasian watermilfoil at sites where the two species coexisted in the lake. Results from greenhouse experiments revealed no effects of Eurasian watermilfoil on water stargrass growth. However, Eurasian watermilfoil biomass was reduced by 46% when treated with high concentration of water stargrass extracts. This is likely due to osmotic effects rather than allelopathic effects of water stargrass. We proposed several possible reasons for the drastic decline of Eurasian watermilfoil and the increase in water stargrass abundance at the northern end of Cayuga Lake, including space competition, nutrients, substrates, wind exposure and water clarity in addition to insect herbivory and mechanic harvesting.     

Nectar and pollen sources for honeybees in Kafrelsheikh province of northern Egypt

This study was conducted at the apiary of the Beekeeping Research Section at the Sakha Agricultural Research Station, ARC, Kafrelsheikh, and other apiaries in Kafrelsheikh province, during two successive years 2015 and 2016. The study aimed to survey nectar and pollen floral resources in Kafrelsheikh province. Ninty seven plant species belonging to 33 families were recorded as nectar sources, and 82 plant species belonging to 36 families were recorded as pollen sources during the whole year. The largest amount of monthly trapped pollen was obtained during May followed by August. It can be concluded that, beekeepers in Kafrelsheikh province can harvest good honey yield at the end of blooming seasons of citrus (Citrus spp.) during March and April, Egyptian clover (Trifolium alexandrinum L.) during May and June, loofah (Luffa aegyptiaca Mill.) during June to October, cotton (Gossypium spp.) during July and August, and banana (Musa spp.) during August and September. They also, could be trapping pollen loads collected from faba bean (Vicia faba L.) and flax (Linum usitatissimum L.) during January to March, date palm (Phoenix dactylifera L.) during March and April, Egyptian clover during May and June, summer seed watermelon (Citrullus lanatus var. colothynthoides L.) during June and July, loofah and maize (Zea mays L.) during June to November.     

Host Range of Mycoleptodiscus terrestris, a Microbial Herbicide Candidate for Eurasian Watermilfoil, Myriophyllum spicatum

A native strain of the fungal plant pathogen Mycoleptodiscus terrestris is capable of causing under experimental conditions necrotic shoot lesions and a generalized decline and disintegration of Eurasian watermilfoil (Myriophyllum spicatum), an exotic submerged aquatic weed in North America. The potential of this fungus to cause disease on nontarget plants was evaluated on 33 species and cultivars in 11 families. The nontarget selection, intended as a Tier I group, was biased toward probable suscepts consisting of submerged, floating, and emergent aquatic species and terrestrial crop plants. The plants were exposed to the fungal mycelium formulated in alginate beads. Pathogenicity rather than phytotoxicity was the primary mode of attack by the fungus. Of the 16 nontarget aquatic species tested, the fungus was pathogenic to Hydrilla verticillata (hydrilla), Myriophyllum aquaticum (parrotfeather), and Ceratophyllum demersum (coontail), but only in hydrilla did it cause plant mortality comparable to that in Eurasian watermilfoil. The remaining 13 submerged, floating, and emergent aquatic species were unaffected. Of the 17 terrestrial species screened, none was significantly affected by the fungus in germination studies, but postemergent disease symptoms developed on seedlings of 10 species. Four of the 10, Medicago sativa (alfalfa), Lotus corniculatus (birdsfoot trefoil), Trifolium hybridum (alyce clover), and T. repens (white clover), developed disease affecting 26-50% of their tissues. Thus, this host range testing scheme, based on the concept of screening plants under the maximum hazard potential, helped to identify susceptible species. Nonetheless, it exaggerated the nontarget risk due to the small number of host species tested. Screening a taxonomically diverse and larger selection of plants as well as testing under conditions of less severe exposure to the fungal inoculum are necessary to obtain a more realistic view of the host range than presently indicated.     

Colonization of a Submersed Aquatic Plant, Eurasian Water Milfoil (Myriophyllum spicatum), by Fungi under Controlled Conditions

A laboratory assay to assess colonization of a submersed aquatic plant, Eurasian water milfoil (Myriophyllum spicatum), by fungi was developed and used to evaluate the colonization potential of Colletotrichum gloeosporioides, Acremonium curvulum, Cladosporium herbarum, Aureobasidium pullulans, a Paecilomyces sp., and an unidentified sterile, septate fungus. Stem segments of plants were first immersed in suspensions of fungal propagules for 24 h and then washed to remove all but the tightly attached component of the population. Inoculation was followed by two growth cycles of 3 days each. At the start of each cycle, washed plants were transferred to a mineral salts medium to provide an opportunity for the attached fungal populations to grow. After each growth period, plants were again washed, and fungal populations in the medium (nonattached), loosely attached and tightly attached to the plant, and within the plant (endophytic) were assayed by dilution plating. The fungi differed in the extent to which they attached to water milfoil and in their ability to grow in association with it. There were relatively few significant differences among the tightly attached fungal populations after 24 h, but growth of the better colonizers led to a greater number of significant differences after 4 and 7 days. In addition, the better colonizers showed sustained regrowth of loosely and nonattached fungal propagules in the face of intermittent removal by washing. A milfoil pathogen, C. gloeosporioides, was the only endophytic colonizer; it was also among the best epiphytic colonizers but was not demonstrably better than A. curvulum, a fungus commonly found as an epiphyte on watermilfoil. The yeastlike hyphomycete Aureobasidium pullulans was the only fungus that consistently failed to establish an increasing population on the plant.     

Isolation and Identification of a Paenibacillus polymyxa Strain That Coproduces a Novel Lantibiotic and Polymyxin

A new bacterial strain, displaying potent antimicrobial properties against gram-negative and gram-positive pathogenic bacteria, was isolated from food. Based on its phenotypical and biochemical properties as well as its 16S rRNA gene sequence, the bacterium was identified as Paenibacillus polymyxa and it was designated as strain OSY-DF. The antimicrobials produced by this strain were isolated from the fermentation broth and subsequently analyzed by liquid chromatography-mass spectrometry. Two antimicrobials were found: a known antibiotic, polymyxin E1, which is active against gram-negative bacteria, and an unknown 2,983-Da compound showing activity against gram-positive bacteria. The latter was purified to homogeneity, and its antimicrobial potency and proteinaceous nature were confirmed. The antimicrobial peptide, designated paenibacillin, is active against a broad range of food-borne pathogenic and spoilage bacteria, including Bacillus spp., Clostridium sporogenes, Lactobacillus spp., Lactococcus lactis, Leuconostoc mesenteroides, Listeria spp., Pediococcus cerevisiae, Staphylococcus aureus, and Streptococcus agalactiae. Furthermore, it possesses the physico-chemical properties of an ideal antimicrobial agent in terms of water solubility, thermal resistance, and stability against acid/alkali (pH 2.0 to 9.0) treatment. Edman degradation, mass spectroscopy, and nuclear magnetic resonance were used to sequence native and chemically modified paenibacillin. While details of the tentative sequence need to be elucidated in future work, the peptide was unequivocally characterized as a novel lantibiotic, with a high degree of posttranslational modifications. The coproduction of polymyxin E1 and a lantibiotic is a finding that has not been reported earlier. The new strain and associated peptide are potentially useful in food and medical applications.