Potential Bioherbicide Research Articles

Interaction of the Bioherbicide Myrothecium verrucaria and Glyphosate for Kudzu Control

Kudzu is an exotic invasive weed in the southeastern U.S. that is difficult to control with current commercial herbicides. Some success for its control has been achieved using a bioherbicidal agent, Myrothecium verrucaria (MV). Spore and mycelial formulations of MV were tested alone and in combination with glyphosate for control of kudzu (Pueraria lobata) under greenhouse and field conditions in naturally-infested areas. In greenhouse and field experiments, kudzu control increased as the concentration of spores or mycelia increased. Glyphosate alone provided 10%, 35%, 50% and 60% control in field experiments at 0.25, 0.50, 0.75 and 1.0X rates, respectively and MV alone spores provided 15%, 50%, 65% and 85% control at 0.25, 0.50, 0.75 and 1.0X rates, respectively. However, when MV spores were combined with glyphosate, significantly higher control occurred than that caused by either component alone. Similar levels of control were observed for MV mycelial formulations applied alone or with glyphosate at equivalent concentrations of the fungus. The rate of disease progression was more rapid and severe at all fungal spore or mycelial formulations and herbicide rates when these propagules were applied in combination with glyphosate. In field tests, 24 h after application, only 20% of kudzu plants were severely damaged by MV alone (0.25X), whereas 80% were severely diseased when MV spores and glyphosate were mixed and applied at 0.25X rates each. A similar trend occurred with the MV mycelial formulation applied at these rates. Synergist interactions on kudzu control were observed, especially when lower levels of MV (spores or mycelia) and glyphosate were combined and applied to kudzu in the greenhouse or in the field. These results suggest that it may be possible to incorporate glyphosate to improve the bioherbicidal control potential and reduce herbicide and inoculum requirements of M. verrucaria spores or mycelium for controlling kudzu.

Screening of Photosynthetic Pigments for Herbicidal Activity with a New Computational Molecular Approach

There is an immense interest among the researchers to identify new herbicides which are effective against the herbs without affecting the environment. In this work, photosynthetic pigments are used as the ligands to predict their herbicidal activity. The enzyme 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase is a good target for the herbicides. Homology modeling of the target enzyme is done using Modeler 9.11 and the model is validated. Docking studies were performed with AutoDock Vina algorithm to predict the binding of the natural pigments such as β-carotene, chlorophyll a, chlorophyll b, phycoerythrin and phycocyanin to the target. β-carotene, phycoerythrin and phycocyanin have higher binding energies indicating the herbicidal activity of the pigments. This work reports a procedure to screen herbicides with computational molecular approach. These pigments will serve as potential bioherbicides in the future.

Leucaena leucocephala leachate compromised membrane integrity, respiration and antioxidative defence of water hyacinth leaf tissues.

BackgroundWater hyacinth is an invasive aquatic weed in many regions of the world. In this study, the bioherbicidal potential of allelopathic plant Leucaena leucocephala against water hyacinth was investigated using a leaf disc assay.ResultsL. leucocephala leachate enhanced electrolyte leakage from water hyacinth leaf discs in a concentration-dependent manner. Control experiments eliminated the possibilities that increased membrane permeability in the leachate-treated leaf discs was due to pH or osmotic effects of the leachate. Thus, the loss of membrane stability in the leachate-treated leaf discs was likely due to phytotoxins detected in the leachate, namely mimosine and phenolic constituents. Decline in tissue respiration was detected in leachate-treated water hyacinth leaf discs. This suggests that the L. leucocephala leachate may contain compounds which acted as respiratory inhibitors. Enhanced reactive oxygen species production coincided with inhibition of catalase and ascorbate peroxidase activities in the leachate-treated water hyacinth leaf tissues. The injurious effects of L. leucocephala leachate on water hyacinth leaf discs probably involved direct inhibition of antioxidant enzymes in addition to direct involvement of some allelochemicals in reactive oxygen species formation.ConclusionIn summary, the toxic effects of L. leucocephala leachate on water hyacinth leaf discs likely lay in its ability to effectively compromise the membrane integrity, tissue respiration and antioxidant defence of the latter.Electronic supplementary materialThe online version of this article (doi:10.1186/1999-3110-54-8) contains supplementary material, which is available to authorized users.

Bioprospecting of Freshwater Macrophytes for Bioactivity

The vast flora diversity of Malaysia is an important advantage where bioprospecting for bioactive natural products is concerned. Published work evaluating the bioactivities of local flora has often focused on popular medicinal or edible plants, which are largely terrestrial species. By contrast, macrophytes (aquatic plants growing near or in water), especially those not popularly used as folk remedies or food, are often underexplored. My research focuses on evaluating the therapeutic and bioherbicidal potential of freshwater macrophytes, which abound in the Kinta Valley region of the State of Perak, Malaysia. In this presentation, I will present the research findings of my group on two macrophytes: Typha domingensis and Eichhornia crassipes. Studies carried out in my laboratory revealed that these macrophytes are potential sources of water-soluble antioxidants, iron chelators, glucosidase inhibitors, as well as bioherbicidal agents. Such bioactivities can be correlated to the phytochemical contents, e.g. polyphenols, flavonoids, hydroxycinnamic acids, and proanthocyanidins, of the macrophytes. Notably, both of these macrophytes are invasive weeds of global importance. Discovery of bioactivity in these aquatic weeds may contribute towards promoting their use as bioresources for therapeutic and bioherbicidal agents, hence providing an alternative to the challenging task of eradicating these weeds.

Ecotoxicological characterisation of a mycoherbicide mixture isolated from the fungus Ascochyta caulina

Recent studies have highlighted the possibility of using a mixture of three fungal toxins produced by Ascochyta caulina as a bioherbicide to control noxious weeds. However, to assess the commercial potential of this new bioherbicide, it is necessary to increase knowledge regarding the environmental behaviour of these toxins, and particularly their effects on non-target organisms. A negative ecotoxicological profile of this potential bioherbicide could eventually hinder its registration. In this study, acute and chronic effects of the A. caulina toxins (ACTs) on non-target organisms were evaluated in the laboratory. The (96 h) calculated algal Er C50 and NOEC were 142.7 and <12.5 mg L(-1) respectively. For daphnids, the EC50 (48 h, acute test) was 20.1 mg L(-1) , whereas the NOEC (21 day chronic reproduction test) was 6.3 µg L(-1) . No cases of mortality, sublethal effects or abnormalities were observed in the acute and chronic tests for Brachydanio rerio at a concentration of 100 mg L(-1) , indicating a low toxicity of ACTs for this organism. The calculated LC50 (14 days, acute test) and NOEC (56 day chronic reproduction) for earthworms were >1000 and ≥250 mg kg(-1) soil respectively. The present study has provided baseline information needed to assess the potential hazard of the fungal toxins of A. caulina. In particular, the acute and chronic effects on aquatic (algae, Daphnia and fish) and terrestrial organisms (earthworms) have been highlighted. Finally, a comparison of the inherent toxicity of ACTs with other synthetic herbicides has shown comparative ecotoxicity of the tested mixture.

Allelopathic and bioherbicidal potential of Cladonia verticillaris on the germination and growth of Lactuca sativa

Responses to germination and initial growth of Lactuca sativa (lettuce) submitted to organic extracts and purified compounds of Cladonia verticillaris (“salambaia”) were analyzed in this work. The experiments were conducted in laboratory conditions using extracts and pure compounds at different concentrations. None of the assays showed any influence on the germination of L. sativa seeds using C. verticillaris extracts; however, modifications in leaf area and seedling hypocotyl and root development occurred. In the growth experiments, seedlings exposed to ether or acetone extract showed diminished hypocotyl growth in detriment to the root stimulus, compared to controls. Increases in extract concentrations led to the formation of abnormal seedlings. To determine the allelochemicals of C. verticillaris, its principal components, fumarprotocetraric and protocetraric acids, were isolated and then analyzed by high performance liquid chromatography (HPLC). When the seedlings were exposed to the two acids separately, presented increased leaf area at all concentrations. In contrast, hypocotyl and root stimulus was observed only in the presence of protocetraric acid at different concentrations. Fumarprotocetraric as well as protocetraric acids, isolated and purified from C. verticillaris and Parmotrema dilatatum respectively, influenced the development of L. sativa seedlings at high concentrations, indicating a possible bioherbicide potential of these acids.

Hemp sesbania (Sesbania exaltata) control in rice (Oryza sativa) with the bioherbicidal fungus Colletotrichum gloeosporioides f. sp. aeschynomene formulated in an invert emulsion

In greenhouse and field experiments, an invert emulsion (MSG 8.25) was tested with dried, formulated spores of the bioherbicidal fungus Colletotrichum gloeosporioides f. sp. aeschynomene, a highly virulent pathogen of the leguminous weed Aeschynomene virginica (northern jointvetch), but considered ‘immune’ against another more serious leguminous weed, Sesbania exaltata (hemp sesbania). A 1:1 (v/v) fungus/invert emulsion mixture resulted in 100% infection and mortality of inoculated hemp sesbania seedlings over a 21-day period under greenhouse conditions. In replicated field tests of the fungus/invert formulation conducted in Stuttgart, AR, and Stoneville, MS, hemp sesbania was controlled 85 and 90%, respectively. These results suggest that this invert emulsion expands the host range of C. gloeosporioides f. sp. aeschynomene, with a concomitant improvement of the bioherbicidal potential of this pathogen.

Molecular and morphological differentiation of Colletotrichum truncatum from scentless chamomile and selected crop species

The fungal pathogen Colletotrichum truncatum is a potential bioherbicide agent for management of the noxious weed scentless chamomile (Metricaria perforate) in western Canada. Isolates of the potential biocontrol agent from scentless chamomile cannot be separated morphologically from the pathogen that causes anthracnose on lentil and field pea, raising questions about crop safety for its use as a bioherbicide agent in the region. Internal transcribed spacer (ITS) regions of ribosomal DNA (rDNA) of C. truncatum from scentless chamomile and several crop species were examined for genetic differences. ITS-rDNA fragments of 63 isolates were amplified using the universal primers Its4 and Its5 and sequenced. The rDNA fragment was 571 bp in size for isolates from scentless chamomile, 566 bp from lentil, 558 or 559 bp from field pea isolates and 555 bp from soybean. Phylogenetic analysis using parsimony and neighbour-joining methods separated these putative C. truncatum isolates into four groups that coincided with host origin. All of the isolates of scentless chamomile fell within a single cluster regardless of their geographic origins, and were most closely related to C. truncatum isolates from lentil. Morphologically, the isolates from scentless chamomile, lentil and field pea were similar in conidial shape and dimension, whereas soybean isolates had substantially longer and more falcate conidia and slightly bigger appressoria. Based on the sequence information, strain-specific PCR primers were designed for isolates of C. truncatum from each host and used to amplify specific rDNA fragments (markers). This molecular technique was sensitive and strain-specific for detection and differentiation of Colletotrichum isolates from scentless chamomile, lentil, field pea and soybean, and may be used as a tracking and monitoring tool in environmental risk assessment for the bioherbicide agent.

Effect of environmental factors and precursors on oxalic acid production, mycelial biomass and virulence of a potential bioherbicide isolate of Sclerotium rolfsii SC64 produced in liquid culture

The fungus Sclerotium rolfsii is presently under development as a bioherbicide for broadleaf weed species using fungus-infested substrates as application material in this laboratory. The effect of environmental factors and three precursors (citric acid, ascorbic acid, and sodium succinate) on mycelial growth, oxalic acid production, and virulence by SC64 in liquid culture were investigated. The results showed that for mycelia growth the optimum liquid medium was Modified Richard's solution (MRS) among the five tested media, but potato dextrose broth (PDB) produced the maximum oxalic acid production and virulence on detached Solidago canadensis leaves. When PDB was used as the basic medium, the oxalic acid/mycelial dry weight (mg g–1) ratio reached the peak 4 days after inoculation. The optimum temperature for oxalic acid production was at 27°C, but increased mycelial dry weight and virulence were observed at 30°C. The optimum range of initial pH value for oxalic acid accumulation was 4.0–6.0, with the optimal pH 5.0; highest mycelial growth was with an initial pH 3.5–6.0 (optimum pH 5.0) and subsequently pH 3.5–5.5 (maximum at pH 3.5). Both mycelial dry weight and oxalic acid production showed a decreasing trend as a result of the precursor of oxalic acid being added to PDB. Among the three precursors, the greatest decrease in mycelial dry weight, and oxalic acid production was caused by sodium succinate. This clarification of optimal conditions for production of mycelial biomass while insuring high concentrations of oxalic acid and high virulence should be useful for further development of this fungus as biocontrol agent.

Efficacy of Exserohilum longirostratum on barnyard grass (Echinochloa crus-galli spp. crusgalli) under field conditions

An isolate of Exserohilum longirostratum obtained from Rottboelia cochinchinensis in Malaysia was highly pathogenic to barnyardgrass (Echinocloa crus-galli,) a serious weed in rice fields in Malaysia. In glasshouse trials, high levels of barnyardgrass control were achieved when E. longirostratum was applied as a conidial concentration at 105 conidia/mL. This conidial concentration may not be sufficient for field use; therefore, a mini plot trial was carried out to assess the bioherbicidal potential of the fungus and the efficacy of different inoculum types under field conditions. Excellent barnyard grass disease as indicated by Area Under Disease Progress Curve (AUDPC) was achieved with mycelium (AUDPC = 583.8 unit2); mycelium + pretichlaclor (AUDPC = 610.4 unit2) and conidia alone (AUDPC = 468.3 unit2) compared to conidia + pretichlaclor (AUDPC = 395.8 unit2). Although conidia caused lower disease severity on barnyard grass compared to mycelium, this inoculum equally reduced the competition from barnyard grass as indicated by lower mean dry weight and fewer barnyard grass tiller numbers. Positive correlations between AUDPC of rice and its tiller number and AUDPC of rice and its dry weight were recorded while negative correlations were observed for the AUDPC of barnyard grass and its tiller number and AUDPC of barnyard grass and its dry weight. These findings confirmed that E. longirostratum applied either in the form of conidia or mycelium was effective against barnyard grass under field conditions.

Use of Selenium-enriched Mustard and Canola Seed Meals as Potential Bioherbicides and Green Fertilizer in Strawberry Production

New plant-based products can be produced from seed harvested from Brassica species used for phytomanaging selenium (Se) in the west side of central California. Se-enriched seed meals produced from canola (Brassica napus) and mustard (Sinapis alba) plants were tested as potential bioherbicides and green fertilizers in strawberry production under controlled and field conditions for two seasons. Treatments consisted of adding each meal (containing 2.2 mg Se/kg dry mass) to the soil at rates equivalent to 0, 2, and 6 t/acre, respectively, 7 days before planting. In growth chamber studies, the highest rates of either meal lowered berry yields by a high of 30% compared with no application (control). Among the nutrient accumulation, berry Se, calcium (Ca), manganese (Mn), and zinc consistently increased with most Brassica meal treatments and most significantly with the mustard meal. In the field studies, mustard treatments lowered the emergence of summer-germinating and resident winter annual weeds more than canola and control treatments. Strawberry fruit yields increased with all Brassica treatments, except a 42% fruit yield reduction was observed at a 6 t/acre rate of mustard meal. Increases in fruit Se concentrations and increases in Ca, phosphorus, and Mn were often observed for all Brassica treatments. Amending soils with Brassica seed meals may have more practical viability in organic agriculture as a potential bioherbicide and green fertilizer.

유채(Brassica napus L.) 종자의 발아와 초기생장을 억제하는 식물정유의 선발

This study was conducted to investigate the phytotoxic effects of 19 essential oils on seed germination and initial growth of rapeseed (Brassica napus L.). We found that anise, cinnamon, citronella, clove, geranium, lemongrass, mustard and pine oils completely inhibited germination of rapeseed at 100× dilute solution. Based on the inhibition rates of rapeseed emergence and initial growth, three essential oils (cinnamon, clove, and geranium) were selected as potential bio-herbicides. Under pre-emergence applications of cinnamon, clove, and geranium oils at 90 kg ai ha -1 , rates of rapeseed emergence were 7.1, 25.0, and 3.6% and its initial growth were 22.0, 9.9 and 11.0%, respectively.

Use of Local Plant Aqueous Extracts as Potential Bio-herbicides against Striga hermonthica (Del.) Benth. in Burkina Faso

This study was conducted to evaluate the allelopathic properties of endogenous plant species against Striga hermonthica (Del.) Benth., a root parasitic weed. In this respect, twenty five water extracts and eight freeze dried water extracts from sixteen plant species were screened in bio-assays to test their ability to induce or inhibit the germination of S. hermonthica seeds. Striga seeds were conditioned either in 10% water extracts or in 10% diluted lyophilisats to check their inhibition effect on Striga seed germination. Three doses, 1, 5 and 10% of water extracts were applied on conditioned Striga seeds to test their ability to induce Striga germination. Aqueous extracts from four plant species reduced Striga seed germination by 95.8 to 99.8% compared to the untreated control. Aqueous extracts from two others also significantly reduced Striga germination by 93.1 and 86.3%, respectively. Lyophilisats from four species inhibited Striga seeds germination, whereas that of two others reduced Striga seeds germination by 93.5 and 99.6%, respectively. Only 1% aqueous extracts of Ceiba pentandra and Eucalyptus camaldulensis significantly stimulated Striga seed germination by 39.2 and 38.9%. These results pointed out that the metabolites produced by some of the local plant species may have the potential to be used as bio-herbicides to control S. hermonthica and enhance cereals yield.

Induction of infection in Sesbania exaltata by Colletotrichum gloeosporioides f. sp. aeschynomene formulated in an invert emulsion

In greenhouse experiments, an invert emulsion (MSG 8.25) was tested with spores of the mycoherbicidal fungus Colletotrichum gloeosporioides f. sp. aeschynomene, a highly virulent pathogen of the leguminous weed Aeschynomene virginica (northern jointvetch), but considered “immune” against another leguminous weed, Sesbaniaexaltata (hemp sesbania). A 1:1 (v/v) fungus/invert emulsion mixture resulted in 100% infection and mortality of inoculated hemp sesbania seedlings over a 21 day period. Microscopic examinations revealed that the fungus proliferated within the cells of hemp sesbania and produced anthracnose lesions containing acervuli on infected stems. The fungus was reisolated and found to infect and kill northern jointvetch seedlings, thus fulfilling Koch’s postulates for disease identification. These results suggest that this invert emulsion expands the host range of C. gloeosporioides f. sp. aeschynomene, and possibly improves the bioherbicidal potential of this pathogen.

Fungal transformation of Colletotrichum coccodes with bacterial oahA to suppress defenses of Abutilon theophrasti

Abutilon theophrasti (Malvaceae) is a widely distributed weed that infests a variety of agricultural crops. Cotton is in the same botanical family, precluding the use of selective chemical herbicides. An Abutilon-specific pathovar of Colletotrichum coccodes has been considered as a potential bioherbicide for the selective control of A. theophrasti. An evolutionary balance between microorganism and weed maintains a relatively low virulence of the microorganism, such that it does not destroy its food supply by rapidly killing its host. We overexpressed the oahA (oxaloacetate acetylhydrolase) gene in C. coccodes to increase oxalate production, as oxalate inhibits plant-defensive callose synthase by chelating calcium, an obligate co-factor of this enzyme. C. coccodes oahA transformants acidified the surrounding medium, an indication of acid production by hydrolysis of oxaloacetate. Less callose was seen visually around lesions caused by C. coccodes oahA transformants than the wild type in preliminary experiments, but this was not biochemically quantifiable. Further optimization might be obtained through gene stacking, or the addition of multiple virulence genes to this potential biocontrol agent.

Susceptibility of silver birch pruning wounds to infection by white-rot fungus (Chondrostereum purpureum), a potential bioherbicide

We artificially inoculated pruning wounds of silver birch (Betula pendula Roth) to study seasonal variation in their vulnerability to infection by the fungal decomposer Chondrostereum purpureum (Pers. ex Fr.) Pouzar. This information is critical to the assessment of incidental infection risks in areas where C. purpureum may be used as a bioherbicide. On seven monthly occasions between April and October 2005, 30 birch trees were pruned to yield a total of 210 experimental trees. On each occasion, 10 trees were inoculated immediately with C. purpureum mycelium, 10 were inoculated with blank inoculum and 10 were only pruned. In the summer of 2007, a survey of 129 experimental trees showed that pruning wounds were most susceptible to infection during May. Treatment with C. purpureum at other times during the growing season also increased the extent of discoloration or decay but the effect was considerably less.

The essential oils in agriculture as an alternative strategy to herbicides: a case study

In this study we investigated the allelopathic effect of some essential oils extracted from aromatic plants to inhibit weed seed germination. The study was performed to evaluate the potential genotoxic effect of lavender on Vicia faba root meristems by genotoxicity tests (comet assay and micronuclei test) and to evaluate microbial diversity fluctuation by Restriction Fragment Length Polymorphism (RFLP) molecular analysis on ecological, biological and conventional soil. The results showed a phytotoxicity effect of the essential oil for all the concentration tested, while there is not a dose-dependent relation between oil concentrations and the genotoxic effect. A fluctuation of microbial communities in relation to the release of essential oils by Lavandula officinalis was also observed. Then we suggest that the essential oils could be useful as potential bioherbicides as an alternative strategy to the chemical remedy, but further studies are necessary to evaluate their appropriate use in the field.

Enzyme‐Linked Immunosorbent Assay Detection of Trichothecenes Produced by the Bioherbicide Myrothecium verrucaria in Cell Cultures, Extracts, and Plant Tissues

A rapid technique for trichothecene detection was needed in screening tests of the potential bioherbicide Myrothecium verrucaria (MV), in order to select strains, mutants, or formulations that were void of or that possessed low amounts of these undesirable mycotoxin compounds. Commercially available enzyme‐linked immunosorbent assay (ELISA) plates for trichothecene detection, possessing cross‐reactivity with several trichothecene mycotoxins (e.g., verrucarin A, and J, roridin A, L‐2, E, and H), were tested for their ability to detect trichothecenes produced by a strain of Myrothecium verrucaria (MV) in cell cultures, in plant tissues (hemp sesbania and kudzu) treated with purified roridin A, or ethyl acetate fractions of MV cultures. Evaluations of ELISA assays showed linear responses for standards of verrucarin A and roridin A over a concentration range of 0.2 to 20 ppb. Ethyl acetate or aqueous extractions were used to obtain samples from MV cultures and plant tissues for testing. Trichothecenes were detected in conidia and mycelia of MV, and in agar upon which wild‐type MV was grown, indicating secretion into the growth media. Two MV sectors (morphological variants of wild type) also tested positive for trichothecenes. Purified roridin A and concentrated extracts containing trichothecenes from MV spore cultures exhibited phytotoxicity (growth inhibition or necrosis) when applied to excised shoots of hemp sesbania seedlings and intact kudzu leaf tissues. Evidence of some translocation of trichothecenes from the application point in kudzu was found, but translocation to the upper shoot portion of hemp sesbania was not detected at the lowest limit of detection in this assay (0.14 ppb). This assay is also being employed to identify induced mutants and/or other naturally occurring sectors deficient in trichothecene mycotoxin production. Results indicated that ELISA is a sensitive and rapid assay method to quantify trichothecenes produced by this bioherbicidal fungus and in certain plant tissues treated with trichothecenes.

Role of native soil biology in Brassicaceous seed meal-induced weed suppression

Biologically based weed control strategies are needed in organic and low-input systems. One promising practice is the application of Brassicaceous seed meal (BSM) residue, a byproduct of biodiesel production. When applied as a soil amendment, BSM residue has exhibited potential bioherbicide activity. In this study, tree fruit orchard soils were treated with various BSMs and the impact of Pythium on weed suppression was examined in field and greenhouse studies. Although weed control obtained in response to Brassicaceous residue amendments has been repeatedly attributed solely to release of allelopathic phytochemicals, multiple lines of evidence acquired in these studies indicate the involvement of a microbiological component. Reduced weed emergence and increased weed seedling mortality were not related to BSM glucosinolate content but were correlated with significant increases in resident populations of Pythium spp. in three different orchard soils. Seed meal of Brassica juncea did not amplify resident Pythium populations and did not suppress weed emergence. Application of Glycine max SM did stimulate Pythium spp. populations and likewise suppressed weed emergence. Application of a mefenoxam drench to Pythium-enriched soil significantly reduced weed suppression. These studies indicate that a microbial mechanism is involved in SM-induced weed suppression and that selective enhancement of resident pathogenic Pythium spp. can be utilized for the purpose of weed control.

The interaction of two potential fungal bioherbicides and a sub-lethal rate of glyphosate for the control of shattercane

Greenhouse and laboratory experiments were conducted with the potential bioherbicides Colletotrichum graminicola (Cg) and Gloeocercospora sorghi (Gs) for control of shattercane weed. Single-spray tank mixture applications containing different ratios of the two fungi resulted in additive percent weed biomass losses. Intraspecific (Cg + Cg or Gs + Gs) and interspecific (Cg + Gs or Gs + Cg) sequential applications 1- or 7-days apart indicated antagonistic interactions in percent biomass loss. Application of either fungus with, or 1–3 days prior to, a sub-lethal concentration of glyphosate resulted in an antagonistic percent biomass loss; while application of glyphosate prior to either potential bioherbicide resulted in a synergistic weed disease response. Conidia germination studies conducted both in vitro on agar plates and with leaf impression peels suggest that antagonistic interactions observed in weed disease severity are probably due to the host–pathogen response following infection.