The role of ACC deaminase producing bacteria in improving sweet corn (*Zea mays* L. var *saccharata*) productivity under limited availability of irrigation water

Drought is a major environmental stress limiting crop productivity globally, particularly in arid and semi-arid regions. Water stress causes economic losses in agriculture by impairing plant growth, physiology, and nutrient uptake. Sweet corn (Zea mays L. var saccharata) is cultivated for its high sugar content and plays important nutritional and industrial roles, including potential as a bioethanol feedstock. PGPR such as Pseudomonas can mitigate drought effects via multiple mechanisms, notably ACC deaminase activity that lowers stress ethylene, improving plant stress tolerance. Despite substantial research on PGPR and drought tolerance in maize, fewer studies have focused on physiological responses and nutrient uptake in sweet corn inoculated with Pseudomonas fluorescens under water stress. This study aimed to evaluate drought tolerance induced by four P. fluorescens strains (P1, P3, P5/P8, P14) and their combination under varying irrigation levels, with the hypothesis that combined inoculation would be more effective than single strains in mitigating water limitation effects in sweet corn.

Previous studies report that PGPR alleviate drought stress by enhancing antioxidant balance, photosynthetic pigments, osmolyte accumulation, and nutrient uptake. In pea and maize, inoculation with ACC deaminase-producing bacteria reduced stress-induced antioxidant enzyme activities and increased chlorophyll, proteins, and carbohydrates, indicating improved water status. PGPR with ACC deaminase can lower plant ethylene by degrading its precursor ACC, promoting root growth and nutrient acquisition. Reports also show enhanced N, P, K uptake in maize under water stress due to reduced ethylene accumulation and improved root architecture. However, detailed assessments of sweet corn physiological traits (chlorophylls, fluorescence parameters, osmolytes), antioxidant enzymes, and leaf macro- and micronutrients in response to P. fluorescens strains under defined irrigation deficits are limited, motivating this study.

Bacterial strains and inoculation: Four drought-tolerant Pseudomonas fluorescens strains (P1 = MT949838, P3 = MT949840, P5 = MT949845, P14 = MT949851) were obtained from Vali-e-Asr University of Rafsanjan. ACC deaminase activity, auxin synthesis, phosphate solubilization, and siderophore production were characterized using standard protocols. Sweet corn seeds (Chase hybrid) were surface-sterilized (1.5% NaOCl, 20 min), rinsed, and soaked 24 h in bacterial suspensions adjusted to OD600 = 0.5 (~1×10^8 CFU ml−1). Culture conditions included growth on nutrient agar at 27–28 °C for 48 h, transfer to TSB, shaking incubation, centrifugation, and resuspension. Experimental site and design: Field trials were conducted in Marvdasht, Fars Province, Iran (29°56′N, 52°47′E) during 2016 and 2017 (semi-arid climate; mean annual temperature 28.5 °C; rainfall 365 mm). Soil was clay-loam (Typic Calcixerpts). Pre-experiment soil properties (0–30 cm) were measured for texture, pH, EC, N, P, K. The experiment used a randomized complete block design with three replications. Main plots: irrigation regimes at 100% (I100), 80% (I80), and 60% (I60) of crop water requirement. Subplots: bacterial inoculation treatments—control (no bacteria), P1, P3, P5/P8, P14, and a combination of all four strains. Plots were 3×3 m; row and plant spacing 75 and 20 cm (66,000 plants ha−1). Fertilization: 110 kg P ha−1 (triple superphosphate) and 80 kg K ha−1 (potassium sulfate) at sowing; 200 kg N ha−1 as urea (two-thirds at sowing, one-third at tasseling). Irrigation scheduling: Tape irrigation. Irrigation applied uniformly until seedling establishment; treatments imposed from 5-leaf stage to end. Reference evapotranspiration (ET0) calculated via FAO Penman–Monteith; crop evapotranspiration (ETc = Kc × ET0; Kc 1.15 mid-season, 1.05 maturity). Irrigation depth determined by cumulative ETc, with plot water volume measured by water counters. Measurements: At milk grain stage, the upper three leaves were sampled, immediately frozen in liquid nitrogen, and stored at −80 °C. Pigments (chlorophyll a, b, total) measured spectrophotometrically. Chlorophyll fluorescence parameters (Fo, Fm, Fv/Fm) recorded with a fluorimeter. Free proline quantified (acid-ninhydrin method). Total soluble sugars measured by colorimetry. Antioxidant enzymes (catalase, peroxidase) assayed spectrophotometrically. Leaf N (Kjeldahl), P (vanadomolybdate colorimetry), K (flame photometry); micronutrients (Fe, Zn, Cu, Mn) by dry ash digestion and atomic absorption. Yields: ear yield and canned seed yield measured from 3 m2 per plot at milk stage. Statistics: SAS v9.1; ANOVA for main and interaction effects; means separated by LSD at p ≤ 0.05; three replicates.

• Water stress effects: I60 versus I100 reduced leaf nutrients (except K), chlorophylls, and Fv/Fm, and increased Fo, Fm, proline, total soluble sugars (TSS), and CAT and POX activities. Specifically, I60 reduced leaf N, Zn, and Mn by 36.0%, 32.0%, and 39.0%, respectively, and increased K by 21.98%. P and Fe contents decreased by 37.0% and 24.5%. The lowest Fv/Fm (0.648) occurred at I60. • Bacterial inoculation effects: All strains improved physiological traits and nutrient accumulation relative to control, with the four-strain combination generally most effective and P1 often the best single strain. The four-strain combination increased leaf N, K, Zn, and Mn by 46.04%, 43.0%, 34.50%, and 24.0% over control. P1 yielded the highest P and Fe across irrigation levels. • Antioxidant enzymes: CAT and POX activities rose under water deficit (by 53.0% and 65.5% at I80 and I60 for CAT; 55.0% and 64.0% for POX). Inoculation reduced CAT and POX compared with control at all irrigation levels, with the lowest activities under the four-strain combination. • Photosynthetic pigments: Drought reduced chlorophyll a, b, and total chlorophyll. Inoculation with P1 and P5 significantly increased chlorophyll a under stress; P1 produced the highest chlorophyll b and total chlorophyll across irrigation regimes. • Chlorophyll fluorescence: The four-strain combination reduced Fo and Fm compared with control and produced the highest Fv/Fm (0.685). Control had the highest Fo (77.40), while the four-strain combination had the lowest (72.84). • Osmolytes: Inoculation increased proline and TSS, with strain- and irrigation-dependent responses. Under I60, P1 increased proline by 111% and TSS by 28% over control; under I100 and I80, P1 increased TSS by 54% and 64%. • Yield: Water stress (I60) reduced ear yield and canned seed yield by 22.2% and 21.7%. Inoculation improved yields at all irrigation levels. Maximum ear yield (1651.6 g m−2) and canned seed yield (616.1 g m−2) occurred with the four-strain combination at I100. Yield losses at I60 were mitigated from 25.0% (ear) and 25.0% (canned seed) in control to 17.5% and 21.0% with the four-strain combination. Overall, the four-strain combination increased ear yield by 44% and canned seed yield by 27% over control.

Findings demonstrate that limited irrigation impairs sweet corn physiology by reducing chlorophyll content and photosystem II efficiency (lower Fv/Fm), and diminishing macro- and micronutrient uptake, while provoking oxidative stress (elevated CAT, POX) and osmotic adjustment (higher proline, sugars). Inoculation with ACC deaminase-producing Pseudomonas fluorescens mitigated these effects via multiple complementary mechanisms: lowering stress ethylene through ACC deaminase, enhancing root growth via auxin production, solubilizing mineral phosphates, and producing siderophores to mobilize iron. These mechanisms improved water and nutrient acquisition (notably N, P, K, Fe, Zn, Mn), maintained photosynthetic pigment levels, optimized chlorophyll fluorescence parameters (higher Fv/Fm, lower Fo), and moderated oxidative enzyme activities, collectively supporting assimilate production and yield components. The four-strain combination delivered synergistic benefits exceeding single strains, indicating that stacking functional traits (ACC deaminase, IAA synthesis, phosphate solubilization, siderophore production) enhances drought resilience and yield stability under irrigation deficits.

Water scarcity alters sweet corn physiological characteristics and nutrient uptake, reducing yields. Inoculation with P. fluorescens strains improved physiological attributes and yields under all irrigation regimes. The combined four-strain inoculation, leveraging synergistic ACC deaminase activity, auxin synthesis, phosphate solubilization, and siderophore production, most effectively reduced stress ethylene impacts and enhanced water and nutrient absorption, significantly improving ear and canned seed yields under both optimal and limited irrigation. Future work could explore root system architecture, rhizosphere processes, and multi-environment validation to optimize consortia composition and application strategies.