Biostimulant Substances for Sustainable Agriculture: Origin, Operating Mechanisms and Effects on Cucurbits, Leafy Greens, and Nightshade Vegetables Species.

Climate change is a pressing matter of anthropogenic nature to which agriculture contributes by abusing production inputs such as inorganic fertilizers and fertigation water, thus degrading land and water sources. Moreover, as the increase in the demand of food in 2050 is estimated to be 25 to 70% more than what is currently produced today, a sustainable intensification of agriculture is needed. Biostimulant substances are products that the EU states work by promoting growth, resistance to plant abiotic stress, and increasing produce quality, and may be a valid strategy to enhance sustainable agricultural practice. Presented in this review is a comprehensive look at the scientific literature regarding the widely used and EU-sanctioned biostimulant substances categories of silicon, seaweed extracts, protein hydrolysates, and humic substances. Starting from their origin, the modulation of plants' hormonal networks, physiology, and stress defense systems, their in vivo effects are discussed on some of the most prominent vegetable species of the popular plant groupings of cucurbits, leafy greens, and nightshades. The review concludes by identifying several research areas relevant to biostimulant substances to exploit and enhance the biostimulant action of these substances and signaling molecules in horticulture.

Characterization of Starch in Cucurbita moschata Germplasms throughout Fruit Development.

Pumpkins (Cucurbita moschata; Cucurbitaceae) are the rich source of nutrients and valued for their biologically active substances to be used for the treatment of several diseases. The contents, composition, and conformation of starch are the significant quality traits of C. moschata. Two germplasms were targeted for analysis regarding the taste difference. Results indicated that the total starch contents and amylose/amylopectin ratio were high in CMO-X as compared to CMO-E during each fruit development stage. Scanning electron microscopy and transmission electron microscopy observations revealed that smooth surface starch granules fused together to enhance the starch accumulation. For a comparison of fruit development in CMO-E and CMO-X, the putative pathway for starch metabolism was developed and homologs were identified for each key gene involved in the pathway. GBSS and SBE were correlated with the difference in the amylose/amylopectin ratio of CMO-E and CMO-X. Conclusively, the developmental regulation of genes associated with starch accumulation can be considered as an important factor for the determination of fruit quality.

Hypoglycemic effect and bioactive compounds associated with the ripening stages of the Cucurbita ficifolia Bouché fruit.

BACKGROUND: The fruit of Cucurbita ficifolia Bouché is known in Mexico as 'chilacayote'. The scientific interest that C. ficifolia Bouché has acquired is due to its important hypoglycemic effect. The present research aimed (i) to discover whether this hypoglycemic property is present at different stages of development of this fruit, and (ii) to characterize some bioactive compounds with antioxidant or anti-inflammatory properties. Ethylene production, respiration rate, and maturity indices were determined during fruit development. The chemical characterization of the aqueous extracts of each stage of maturity studied was determined and their hypoglycemic effects were bioassayed using groups of normal mice with diabetes induced by streptozotocin at a dose of 500 mg-1 kg-1 body weight. RESULTS: Respiration rate and ethylene production showed a typical pattern for non-climacteric fruit and the quality parameters did not show significant changes. Phenolic compounds such as gallic acid and chlorogenic acid were found to have the highest concentration at 15 days of development. Extracts at 15 days showed a hypoglycemic effect that was 11% greater than that of glibenclamide in diabetized mice. CONCLUSION: All stages of development of C. ficifolia fruit had a hypoglycemic effect; however, the aqueous extract from the fruit at 15 days of development showed a better effect than glibenclamide. This finding highlights the potential of this maturity stage, and shows that it is appropriate for inclusion in treatments of type 2 diabetes mellitus. The results also indicate that phenolic compounds are mainly responsible for this effect and not d-chiro-inositol as previously thought. © 2020 Society of Chemical Industry.

Reynoutria sachalinensis extract elicits SA-dependent defense responses in courgette genotypes against powdery mildew caused by Podosphaera xanthii.

Powdery mildew (PM) caused by Podosphaera xanthii is one of the most important courgette diseases with high yield losses and is currently controlled by fungicides and sulphur applications in conventional and organic production. Plant derived elicitors/inducers of resistance are natural compounds that induce resistance to pathogen attack and promote a faster and/or more robust activation of plant defense responses. Giant knotweed (Reynoutria sachalinensis, RS) extract is a known elicitor of plant defenses but its mode of action remains elusive. The aim of this study was to investigate the mechanisms of foliar RS applications and how these affect PM severity and crop performance when used alone or in combination with genetic resistance. RS foliar treatments significantly reduced conidial germination and PM severity on both an intermediate resistance (IR) and a susceptible (S) genotype. RS application triggered plant defense responses, which induced the formation of callose papillae, hydrogen peroxide accumulation and the Salicylic acid (SA) - dependent pathway. Increased SA production was detected along with increased p-coumaric and caffeic acid concentrations. These findings clearly indicate that RS elicits plant defenses notably as a consequence of SA pathway induction.

Impact of superparamagnetic iron oxide nanoparticles (SPIONs) and ionic iron on physiology of summer squash (Cucurbita pepo): A comparative study.

This study investigates the effect of SPIONs (superparamagnetic iron oxide nanoparticles, ∼12.5 nm in size) on summer squash plant (Cucurbita pepo) in the presence and absence of supplementary iron (Fe(II)-EDTA). The plants were grown in nutrient solution with different iron sources: (i) Fe(II)-EDTA, (ii) without Fe(II)-EDTA (iii) SPIONs only, and (iv) Fe(II)-EDTA with SPIONs. Plant growth and development were assessed after 20 days of soaking by measuring phenological parameters such as plant biomass, chlorophyll content, amount of carotenoids, and the catalase enzyme activity. Transmission electron microscopy, inductively coupled plasma atomic emission spectroscopy, X-ray diffraction, and vibrating sample magnetometer methods were used to detect uptake and translocation of SPIONs in plant tissues. Our results showed that SPIONs treatment (without Fe(II)-EDTA) caused growth retardation and decreased the plant biomass and chlorophyll content. Hence, they are not efficient sources to compensate for iron demand of squash plant. Electron microscopy observations, magnetization and elemental analyses revealed that SPIONs are taken-up by plant roots but not translocate to upper organs. In roots, SPIONs use a symplastic route for intercellular transfer. These findings suggest that as an iron source, SPIONs alone are not efficient for plant growth, but can contribute it together with Fe(II)-EDTA.

NMR-Based Metabolomics Approach To Study the Influence of Different Conditions of Water Irrigation and Greenhouse Ventilation on Zucchini Crops.

This study describes the approach of 1H NMR metabolomic profiling for the differentiation of zucchini produced under different conditions of water irrigation (desalinated seawater -0.397 dS/m, 0.52 €/m3 vs groundwater -2.36 dS/m, 0.29 €/m3) and ventilation (surface area of the vent openings/greenhouse area was 15.0% for one sector and 9.8% for the other). Overall, 72 extracts of zucchini ( Cucubirta pepo L. cv Victoria) under four different conditions were regularly analyzed during the spring-summer cycle from April to July 2017. We have found that zucchini plants irrigated with desalinated seawater increased the zucchini production yield, presented fruits with higher concentration of glucose, fructose, and vitamin B3, and displayed an increased antioxidant activity. On the contrary, plant groundwater irrigation produced the increment of sucrose level that could rise the sweetness perception of the fruits. Finally, the ventilation variable produced a higher concentration of trigonelline, histidine, and phenylalanine but only on those zucchinis irrigated with groundwater.

[The fatty acid composition of large pumpkin seed oil (Curucbitae maxima Dich) cultivated in Georgia].

The aim of the study was to identify qualitatively and quantitatively fatty acid composition of large pumpkin seed oil cultivated in Georgia (Cucurbitae maxima Duch) and evaluate its biological activities. Evaluation was conducted using high-performance liquid chromatography method. Fatty acids ranging from C12:0 to C22:0 were identified in the probe. The oil contained 0,2В±0,01mg% lauric, 0,3В±0,01 mg% miristic, 9,0В±0,7mg% palmitic, 5,5В±0,4 mg% stearic, 28,1В±1,0 mg% oleic, 40,2В±1,9 mg% linolic, 12,1В±1,0 mg% linolenic, 2,0В±0,2mg% arachinic and 1,2В±0,1 mg% begenic acids. The investigation showed that large pumpkin seed oil contains a range of biologically significant fatty acids, unique proportion of which attaches great value to the vegetative material.

Root foraging elicits niche complementarity-dependent yield advantage in the ancient 'three sisters' (maize/bean/squash) polyculture.

BACKGROUND AND AIMS: Since ancient times in the Americas, maize, bean and squash have been grown together in a polyculture known as the 'three sisters'. This polyculture and its maize/bean variant have greater yield than component monocultures on a land-equivalent basis. This study shows that below-ground niche complementarity may contribute to this yield advantage. METHODS: Monocultures and polycultures of maize, bean and squash were grown in two seasons in field plots differing in nitrogen (N) and phosphorus (P) availability. Root growth patterns of individual crops and entire polycultures were determined using a modified DNA-based technique to discriminate roots of different species. KEY RESULTS: The maize/bean/squash and maize/bean polycultures had greater yield and biomass production on a land-equivalent basis than the monocultures. Increased biomass production was largely caused by a complementarity effect rather than a selection effect. The differences in root crown architecture and vertical root distribution among the components of the 'three sisters' suggest that these species have different, possibly complementary, nutrient foraging strategies. Maize foraged relatively shallower, common bean explored the vertical soil profile more equally, while the root placement of squash depended on P availability. The density of lateral root branching was significantly greater for all species in the polycultures than in the monocultures. CONCLUSIONS: It is concluded that species differences in root foraging strategies increase total soil exploration, with consequent positive effects on the growth and yield of these ancient polycultures.

Cloning and characterisation of a putative pollen-specific polygalacturonase gene (CpPG1) differentially regulated during pollen development in zucchini (Cucurbita pepo L.).

Studies in zucchini (Cucurbita pepo L. spp. pepo) pollen have been limited to the viability and morphology of the mature pollen grain. The enzyme polygalacturonase (PG) is involved in pollen development and pollination in many species. In this work, we study anther and pollen development of C. pepo and present the cloning and characterisation of a putative PG CpPG1 (Accession no. HQ232488) from pollen cDNA in C. pepo. The predicted protein for CpPG1 has 416 amino acids, with a high homology to other pollen PGs, such as P22 from Oenothera organensis (76%) and PGA3 from Arabidopsis thaliana (73%). CpPG1 belongs to clade C, which comprises PGs expressed in pollen, and presents a 34 amino acid signal peptide for secretion towards the cell wall. DNA-blot analysis revealed that there are at least another two genes that code for PGs in C. pepo. The spatial and temporal accumulation of CpPG1 was studied by semi-quantitative- and qRT-PCR. In addition, mRNA was detected only in anthers, pollen and the rudimentary anthers of bisexual flowers (only present in some zucchini cultivars under certain environmental conditions that trigger anther development in the third whorl of female flowers). However, no expression was detected in cotyledons, stem or fruit. Furthermore, CpPG1 mRNA was accumulated throughout anther development, with the highest expression found in mature pollen. Similarly, exo-PG activity increased from immature anther stages to mature anthers and mature pollen. Overall, these data support the pollen specificity of this gene and suggest an involvement of CpPG1 in pollen development in C. pepo.

Nutritional evaluation and health promoting activities of nuts and seeds cultivated in Greece.

Available data suggest that genetic as well as environmental factors may influence nuts and seeds nutrients content. In this context nuts and seeds cultivated in Greece were studied. Macronutrients content was in agreement with that from other areas. Total phenolics content was in the range of 43.0 ± 2.1-1512.7 ± 60.7 mg GAE/100 g for chestnut and walnut, respectively. Thirteen to 22 individual phenolics were identified in the studied species. Oleanolic acid was in the range of 0.10-9.03 mg/100 g. Pumpkin seeds contained the higher squalene content (71.6 mg/100 g). ß-Sitosterol predominated in all samples except pumpkin seeds. Tocopherols ranged from 8.9 mg/100 g (chestnut) to 29.3 mg/100 g (almond). Nuts and seeds hydrophilic extracts at quantities corresponding to the estimated daily consumption by the Greeks succeeded in inhibiting LDL oxidation in vitro by increasing lag time 1.1-14.1 times. One serving of nuts or seeds may cover a significant fraction of health promoting microconstituents daily intake.

Bioactive compounds in lipid fractions of pumpkin (Cucurbita sp) seeds for use in food.

UNLABELLED: Seeds are considered to be agro-industrial residues, which can be used as source of macronutrients and/or raw material for extraction of vegetable oils, since they present great quantities of bioactive compounds. This study aimed to characterize the lipid fractions and the seeds of pumpkin (Cucurbita sp) varieties Nova Caravela, Mini Paulista, Menina Brasileira, and Moranga de Mesa aiming at using them in food. The chemical composition of the seeds was performed according to the official methods of American Oil Chemists' Society and Association of Official Analytical Chemists. Total carotenoids and phenolic compounds were determined by spectrophotometry, while the levels of tocopherols were analyzed by high efficiency liquid chromatography. It was noted that the seeds contain high amounts of macronutrients that are essential for the functioning of the human organism. As to total carotenoids, Mini Paulista and Menina Brasileira pumpkin varieties presented significant amounts, 26.80 and 26.03 µg/g, respectively. Mini Paulista and Nova Caravela pumpkin varieties showed high amounts of total phenolic compounds in the lipid fractions and in the seeds. It was also found that γ-tocopherol is the isomer that stood out in the lipid fractions and in the seeds, mainly in Menina Brasileira. Finally, the consumption of these seeds and use of lipid fractions provide the supply of large quantities of compounds that are beneficial for health and that may be potentially used in food, besides representing an alternative to better use of agro-industrial residues. PRACTICAL APPLICATION: Bioactive compounds, besides presenting basic nutritional functions, provide metabolic and physiological health benefits when consumed as part of the usual diet. Therefore, there is a growing interest in vegetable oils of special composition, such as the ones extracted from fruit seeds. The seeds of Cucurbita sp are shown to be promising sources of oils, and especially the Cucurbita moschata and maxima species have not yet been fully elucidated. For this reason, it becomes important to investigate the chemical composition and lipid fractions of these seeds, aiming to use them in food.

Complementarity in root architecture for nutrient uptake in ancient maize/bean and maize/bean/squash polycultures.

BACKGROUND AND AIMS: During their domestication, maize, bean and squash evolved in polycultures grown by small-scale farmers in the Americas. Polycultures often overyield on low-fertility soils, which are a primary production constraint in low-input agriculture. We hypothesized that root architectural differences among these crops causes niche complementarity and thereby greater nutrient acquisition than corresponding monocultures. METHODS: A functional-structural plant model, SimRoot, was used to simulate the first 40 d of growth of these crops in monoculture and polyculture and to determine the effects of root competition on nutrient uptake and biomass production of each plant on low-nitrogen, -phosphorus and -potassium soils. KEY RESULTS: Squash, the earliest domesticated crop, was most sensitive to low soil fertility, while bean, the most recently domesticated crop, was least sensitive to low soil fertility. Nitrate uptake and biomass production were up to 7 % greater in the polycultures than in the monocultures, but only when root architecture was taken into account. Enhanced nitrogen capture in polycultures was independent of nitrogen fixation by bean. Root competition had negligible effects on phosphorus or potassium uptake or biomass production. CONCLUSIONS: We conclude that spatial niche differentiation caused by differences in root architecture allows polycultures to overyield when plants are competing for mobile soil resources. However, direct competition for immobile resources might be negligible in agricultural systems. Interspecies root spacing may also be too large to allow maize to benefit from root exudates of bean or squash. Above-ground competition for light, however, may have strong feedbacks on root foraging for immobile nutrients, which may increase cereal growth more than it will decrease the growth of the other crops. We note that the order of domestication of crops correlates with increasing nutrient efficiency, rather than production potential.

Differential responses in yield of pumpkin (Cucurbita maxima L.) and nightshade (Solanum retroflexum Dun.) to the application of three animal manures.

Crop responses to different manures differs considerably, however, the factors responsible for it have not been conclusively elucidated. Consequently, this study examined the biomass response of Cucurbita maxima and Solanum retroflexum to application rates of chicken and kraal manures of cattle and goat, and soil factors related to salinity. The crops' biomass yield increased linearly with increase in application rates of kraal and chicken manures, but steeper in the latter. Results showed that significant decline in biomass yield in chicken manure at rates above 8.5 tons ha(-1) were not due to salinity. The crops' response to cattle and goat kraal manures was linear but polynomial (cubic) in layer chicken manure. It was concluded that the yield decline in chicken manure was due to other manure factors except salinity, probably toxicity effect of the manure fatty acids. Further research was however, recommended to elucidate this claim.

Effect of ripe fruit pulp extract of Cucurbita pepo Linn. in aspirin induced gastric and duodenal ulcer in rats.

A significant decrease in alkaline phosphatase (AP) activity and mucosal thickness and increase in ulcer index (UI) was observed in aspirin treated stomach and duodenum of albino rats. However, pretreatment with C. pepo fruit pulp extract for 14 consecutive days showed increase in AP activity and mucosal thickness along with decrease in UI, suggesting gastroduodenal protective and anti-ulcerogenic properties of C. pepo.

Tritrophic effects on survival and development in a Coccidophagous coccinellid.

Chilocorus nigritus is currently considered one of the most successful biological control agents of armoured scale insects. However, establishment of this beetle in crop pest situations has not always been successful and there are still gaps in our knowledge of its ecology and behaviour. The research involved an examination of tritrophic effects on the survival and development of this common diaspid predator. The effect of a forced change in host plant on the developmental time of the juvenile stages was also examined. The prey and host plants used were the armoured scales Aspidiotus nerii Bouché Homoptera: Diaspididae and Abgrallaspis cyanophylli (Signoret) Homoptera: Diaspididae, on potatoes (Solanum tuberosum L.) and Butternut squashes (Cucurbita moschata Duchesne ex Lamarck). C. nigritus eggs were incubated on four treatments of scales on potatoes or squashes for ten days, half the second instar larvae were then switched to the same scales on the other host plant. Daily observations were made during development to adult emergence. C. nigritus larvae survived and completed development on two species of diaspid scales and the two host plants examined with varying levels of success. Larvae were able to switch from feeding on A. nerii on potatoes to A. nerii on squashes or A. cyanophylli on potatoes to A. cyanophylli on squashes and vice versa with little or no deleterious effects when compared to those beetles reared on one prey and host plant throughout. There were significant differences in survival of larvae reared to the adult stage on both A. nerii and A. cyanophylli on potatoes when compared to larvae reared on these scales on squashes. Squash appears to be a less desirable and potato a more favourable host plant for survival and development. The results have implications for rearing programmes, and the release and establishment of C. nigritus, in fields and glasshouses where scale pests may be present on a variety of host plants, or may be on host plants different to those used in the insectary.

Soil treatment with hot air (Cultivit) as alternative to methyl bromide.

A new development in physical soil treatment is the application of hot air. Hot air treatment is based on blowing extremely hot air into rotavating humid soil. The method has been developed and applied commercially in Israel for the last few years. An increased growth response (IGR) was observed in several crops like potato, cauliflower, kohlrabi and the flower Esclepia, when the soil was treated with hot air prior to planting. Scientific trials were performed in Israel and Cyprus to quantify IGR and to evaluate the efficacy against plant-parasitic nematodes. Squash was grown in tunnels on root-knot nematodes (Meloidogyne javanica and M. incognita) infested fields in sandy (Israel) and clay loam (Cyprus) soils. In Israel hot air treatment was compared with metam sodium and methyl bromide and a cold air treated control. In Cyprus hot air treatment was compared with untreated control. Hot air treatment increased squash yield in Israel with 90 % and in Cyprus with 150%. Root assessments showed that after hot air treatment the root-knot nematodes were still able to infest plants and cause galling damage. Nematode counts were not reduced by hot air treatment. It may be concluded that the general concept of soil disinfestation is not applicable to hot air treatment. Any positive effect in yield could not be explained by reduction in nematode populations in soil. Possible chemical and biological changes in the hot air treated soils need to be identified. Further research will determine the possibilities and limitations of this method in other crops and under various climatic conditions.

[Allelopathic effects of cultured Cucurbita moschata root exudates].

By using the techniques of tissue culture, bio-assay and laboratory analysis, this paper studied the effects of the allelopathic chemicals from pumpkin (Cucurbita moschata) roots on the seed germination and seedling growth of pumpkin, wheat (Triticum aestivum), and radish (Raphanus sativus). The pumpkin root was cultured on a sterile B5 media, and the concentrations of macro- and microelements, organic supplements and hormones in the media were adjusted by using an orthogonal design. After culturing, the culture media was filtered and used in a bioassay to test the autotoxicity and allelopathic effects. The results showed that the pumpkin had both autotoxic and allelopathic effects, and the media having been used to culture the pumpkin roots contained the chemicals that significantly inhibited the seedling growth of wheat and radish. The allelopathic effect decreased when the culture media was diluted. The production of allelochemicals seemed to be related to the growth rate of the pumpkin roots. When the root growth was rapid, the concentration of allelochemicals was high. The allelopathic effect was stronger on radish than on wheat. The optimum concentrations of macro- and microelements, vitamins and hormones for culturing pumpkin root were determined, and the effect of pumpkin root nutrition on the production of allelochemicals was tested. The results indicated that pumpkin root nutrition had a significant effect on the production of allelochemicals.

Farming practices influence wild pollinator populations on squash and pumpkin.

Recent declines in managed honey bee, Apis mellifera L., colonies have increased interest in the current and potential contribution of wild bee populations to the pollination of agricultural crops. Because wild bees often live in agricultural fields, their population density and contribution to crop pollination may be influenced by farming practices, especially those used to reduce the populations of other insects. We took a census of pollinators of squash and pumpkin at 25 farms in Virginia, West Virginia, and Maryland to see whether pollinator abundance was related to farming practices. The main pollinators were Peponapis pruinosa Say; honey bees, and bumble bees (Bombus spp.). The squash bee was the most abundant pollinator on squash and pumpkin, occurring at 23 of 25 farms in population densities that were commonly several times higher than that of other pollinators. Squash bee density was related to tillage practices: no-tillage farms hosted three times as great a density of squash bees as tilled farms. Pollinator density was not related to pesticide use. Honey bee density on squash and pumpkin was not related to the presence of managed honey bee colonies on farms. Farms with colonies did not have more honey bees per flower than farms that did not keep honey bees, probably reflecting the lack of affinity of honey bees for these crops. Future research should examine the economic impacts of managing farms in ways that promote pollinators, particularly pollinators of crops that are not well served by managed honey bee colonies.

Influence of nutrient amendments on the phytoextraction of weathered 2,2-bis(p-chlorophenyl)-1,1-dichloroethylene by cucurbits.

Field experiments were conducted to determine the impact of nutrient amendments on the phytoextraction of weathered 2,2-bis(p-chlorophenyl)-1,1-dichloroethylene (p,p '-DDE) by eight cultivars of cucurbits over a single growing season. Four cultivars of Cucurbita pepo ssp pepo are accumulators and extract percent level quantities of persistent organic pollutants (POPs), whereas C. pepo ssp ovifera and Cucumis sativus are nonaccumulators. The nonamended accumulators phytoextracted 1.0% of the p,p'-DDE and had a translocation factor of 0.44; however, the nonaccumulators removed 0.16% of the contaminant and had a translocation factor value of 0.09. The accumulators also had 3.8 times greater inorganic element content than the nonaccumulators. Duplicate mounds of each cultivar also received weekly nutrient amendments of phosphorus (400 mg/L K2HPO4), nitrogen (200 mg/L KNO3), or nitrogen/phosphorus (400 mg/L K2HPO4, 200 mg/L KNO3); a minus phosphorus treatment involved a 1-L addition of 1 g/L AlSO4 to the soil before planting. When normalized to respective control values (unamended vegetation), the root and stem p,p'-DDE bioconcentration factors (BCF) of the accumulator cultivars were significantly greater than those of the nonaccumulator cultivars under most nutrient regimes. The biomass of accumulator cultivars decreased by up to 61% under certain nutrient regimes, resulting in mixed effects on the amount of p,p'-DDE extracted. Treatment with N and P increased nonaccumulator biomass by 40 to 100%, and increased p,p'-DDE extraction from soil by 75%. Although generally assumed that fertilizer amendments will enhance phytoremediation, as evidenced here by the nonaccumulators, additions of macronutrients may reduce the phytoextraction of weathered POPs by C. pepo ssp pepo. These findings support our hypothesis that the ability of C. pepo ssp pepo to remove sequestered organic contaminants is governed by unique nutrient-acquisition mechanisms.

Plant growth, phosphorus nutrition, and root morphological responses to arbuscular mycorrhizas, phosphorus fertilization, and intraspecific density.

We examined the effects of arbuscular mycorrhizas (AM), phosphorus fertilization, intraspecific density, and their interaction, on the growth, phosphorus uptake, and root morphology of three facultative mycotrophic crops (Capsicum annuum, Zea mays, and Cucurbita pepo). Plants were grown in pots with or without AM at three densities and four phosphorus availabilities for 10 weeks. AM colonization, plant weight, and shoot phosphorus concentration were measured at harvest. Root morphology was assessed for C. annuum and Z. mays. Phosphorus fertilization reduced but did not eliminate AM colonization of all species. AM, phosphorus, and density interacted significantly to modify growth of C. annuum and C. pepo such that increased density and phosphorus diminished beneficial effects of AM. Increased density reduced positive effects of AM on C. annuum and C. pepo shoot phosphorus concentrations. AM altered both Z. mays and C. annuum root morphology in ways that complemented potential phosphorus uptake by mycorrhizas, but increased density and phosphorus diminished these effects. We infer that increased density predominantly influenced plant responses by affecting whether or not carbon (photosynthate) or phosphorus limited plant growth. By exacerbating carbon limitation, high density reduced the benefit/cost ratio of mycorrhizas and minimized their effects.