*3.4. Effects of AMF on Uptake of Plant P*

AMF both in aerobic and anaerobic rice cultivation increases nutrient concentration in the rice plant tissue; the bioavailability of nutrients increased in the soil solution due to mycorrhizae inoculation [52]. As shown in Figure 5, the P concentration in plants was higher in the rice variety CR Dhan 207 (14.796 mg. pot<sup>−</sup>1), followed by Kasalath IC459373 (14.186 mg. pot<sup>−</sup>1) and CR Dhan 204 (14.156 mg. pot−1). Additionally, all the rice varieties inoculated with *Rhizophagus* sp., showed maximum P uptake, followed by *Funneliformis* sp., and *Glomus* sp. inoculation. The results deciphered 16.60–28.50% higher P uptake with AMF inoculation in all the rice varieties, compared to the uninoculated control.

#### *3.5. Responses of AMF on Soil Enzyme and Microbial Properties*

Among the several AMF treatments, *Rhizophagus* sp. (56.59 g p-nitrophenol released h−<sup>1</sup> g−<sup>1</sup> soil) and *Funneliformis* sp. (31.99 g p-nitrophenol released h−<sup>1</sup> g−<sup>1</sup> soil) showed the highest levels of both acid (Figure 6a) and alkaline (Figure 6b) phosphatase activity in CR Dhan 207. Irrespective of the treatments, all rice varieties showed significantly higher acid and alkaline phosphatase activity in AMF-inoculated treatments as compared to the uninoculated control.

**Figure 4.** Enhancement of plant growth parameters due to AMF inoculation in different aerobic rice varieties. Abbreviations: (**a**) root length in cm. (RL); (**b**) shoot length in cm. (SL); (**c**) leaf area m2 (LA); (**d**) chlorophyll SPAD (Chl); (**e**) fresh biomass in gm. (fBioM); (**f**) dry biomass in gm. (dBioM).

**Figure 5.** AMF inoculation on uptake of plant P in different aerobic rice varieties.

**Figure 6.** Enhancement of acid and alkaline phosphatase activities in different aerobic rice varieties. Abbreviations: (**a**) acid phosphatase (AcP) [μg p-nitrophenol released h−<sup>1</sup> g−<sup>1</sup> soil]; (**b**) alkaline phosphatase (AkP) [μg p-nitrophenol released h−<sup>1</sup> g−<sup>1</sup> soil].

In terms of microbial properties, *Funneliformis* sp., *Rhizophagus* sp., *Glomus* sp., *Acaulospora* sp. and *Claroideoglomus* sp. treatments significantly increased MBC in CR Dhan 204 and CR Dhan 207 (706.8 and 688.4 μg g−<sup>1</sup> soil) (Figure 7a). A similar trend was also noticed in DHA (29.43 and 31.82 μgTPF h−1g−<sup>1</sup> soil) (Figure 7b) and FDA (15.37 and 16.13 μg fluorescein h<sup>−</sup>1g−<sup>1</sup> soil) (Figure 7c).

**Figure 7.** AMF and its influence on enhancement of microbial properties in different aerobic rice varieties. Abbreviations: (**a**) microbial biomass carbon (MBC) [μg g−<sup>1</sup> soil]; (**b**) dehydrogenase activity (DHA) [μgTPF h<sup>−</sup>1g−<sup>1</sup> soil]; (**c**) fluorescein diacetate assay (FDA) [μg fluorescein h<sup>−</sup>1g−<sup>1</sup> soil].

Through increasing microbial activity in the soil or by the exudation of enzymes by plants, AMF can also have an impact on soil enzyme activity as well as plant growth promotion [53–55]. Several studies have described how AMF intervention could stimulate soil enzyme activity through soil microorganisms [20,27,56,57]. Generally, soil enzymes are primarily produced by microorganisms; others, such as phosphatase [58], urease, and peroxidases, are also secreted by plant roots. Reports [59–61] have shown that the effects of AMF on various soil enzyme activities and growth-promoting compounds, which release the more biologically accessible nutrients from complex materials, were positively

correlated with increasing ratios of soil-available P and plant biomass as well as strongly abiotic context-dependent factors, with beneficial implications for plant growth. All of the aforementioned data made it very evident that AMF will increase soil enzyme activity, which could improve nutrient cycling.

#### *3.6. Assessing the Mycorrhizal Responsiveness in Different Aerobic Rice Varieties*

Out of the selected rice varieties, mycorrhizal responsiveness was found highest in CR Dhan 207 followed by CR Dhan 204, CR Dhan 205 and Kasalath IC459373 with the application of *Funneliformis* sp. and *Rhizophagus* sp. under P-deficient conditions (Figure 8); however, the AMF responsiveness varies with different rice varieties.

**Figure 8.** Mycorrhizal responsiveness in six aerobic rice varieties with five AMF inoculum effects.

*3.7. Correlation of AMF Colonization with Soil and Plant Properties Using Linear Models*

The linear model was used to select the important parameters linked to AMF colonization and to calculate the correlation of the important variables (Table 2).


**Table 2.** Identification of important parameters using step regression model.

C(p): Mallows' Cp constant; AIC: Akaike information criterion; RMSE: root mean square error.

The correlation analysis (Figure 9) showed that AMF colonization had a significant (*p* < 0.001) positive correlation with FDA (R<sup>2</sup> = 0.911), MBC (R2 = 0.707) and plant-available P (R2 = 0.743). The correlation between AMF colonization and FDA, the *Claroideoglomus* sp. (R<sup>2</sup> = 0.797) and *Acaulospora* sp. (R<sup>2</sup> = 0.700) treatments, showed a higher coefficient than other treatments. Similarly, with AMF colonization and MBC correlation, the higher coefficients were recorded in the treatment *Funneliformis* sp. (R<sup>2</sup> = 0.880) followed by *Glomus* sp. (R2 = 0.850), *Acaulospora* sp. (R2 = 0.845), —*Rhizophagus* sp. (R2 = 0.804) and *Claroideoglomus* sp. (R<sup>2</sup> = 0.744) at *p* < 0.011 levels of significance. The correlation coefficient between AMF colonization and plant P was significantly (*p* < 0.01) at par for microbial treatments *Acaulospora* sp. (R2 = 0.919), *Glomus* sp. (R2 = 0.919), *Funneliformis* sp. (R2 = 0.908), *Rhizophagus* sp. (R<sup>2</sup> = 0.705), and *Claroideoglomus* sp. (R<sup>2</sup> = 0.632). Similarly, many scientific

reports have well documented that AMF plays a crucial role in soils for improving microbial activity, nutrient cycling, soil structure and plant–soil microbe interactions [62–66].

**Figure 9.** Correlation of AMF treatments in different aerobic rice varieties on plant P uptake and soil microbial properties. \* *p* < 0.05, \*\* *p* < 0.01, \*\*\* *p* < 0.001.

Further correlation studies among varieties given in Figure 10 show that CR Dhan 207 (R<sup>2</sup> = 0.972), CR Dhan 204 (R<sup>2</sup> = 0.969), and Kasalath IC459373 (R2 = 0.969) had the maximum coefficients between AMF colonization and FDA (R2 = 0.911) among the different aerobic varieties. However, the correlation between AMF colonization and MBC (R2 = 0.707) indicated that, among the varieties, IR 36 (R<sup>2</sup> = 0.884) and CR Dhan 201 (R2 = 0.856) had the highest coefficient values, whereas CR Dhan 207 (R2 = 0.560) and Kasalath IC459373 (R2 = 0.653) registered the lowest coefficient among other varieties. Regarding the correlation between varieties and plant P uptake (R<sup>2</sup> = 0.743), the highest coefficient was found in CR Dhan 207 (R<sup>2</sup> = 0.927), at *p* < 0.001 significance. This finding clearly indicates that the response of AMF differs based on the type of variety. Thus, the selection of the right type of AMF is essential for exploring the maximum benefit from AMF symbiosis. Das et al. [67] reported that the application of *Glomus* spp. inoculation improved rice crop yields with better P availability in the rhizosphere under alternate wetting and drying irrigation.

**Figure 10.** Response of aerobic rice varieties in AMF colonization correlation with plant P and soil microbial properties using Pearson correlation. \* *p* < 0.05, \*\* *p* < 0.01, \*\*\* *p* < 0.001.

#### **4. Conclusions**

Soil phosphorus deficiency is one of the major problems in aerobic rice cultivation. The fixation of this element in the soil makes it unavailable for plant uptake. The present study revealed that AMF intervention could significantly increase the plant growth and enhance P uptake by 16.60–28.50% compared to the control. Among the four different aerobic rice varieties, the mycorrhizal responsiveness was found to be superior in CR Dhan 207, followed by CR Dhan 204, CR Dhan 205, and CR Dhan 201. The linear modelling approach found that the AMF colonization in all the rice varieties had significant (*p* < 0.001) positive correlation with FDA, MBC, and P uptake, deciphering the importance of AMF association in rice for the improvement of phosphate availability to plants. The present findings require further field validation. However, results suggest that the external application of suitable AMF is essential for improving the plant growth and enhancing the uptake of P in aerobic rice in P-deficient soil.

**Author Contributions:** Conceptualization, P.P., P.K.D.M. and D.M.; methodology, P.P., P.K.D.M. and D.M.; software, A.S. and D.M.; validation, P.P. and P.C.; formal analysis, D.M.; investigation, P.P., P.C., A.K.N. and P.K.D.M.; data curation, D.M.; writing—original draft preparation, D.M., P.P. and A.S.; visualization, D.M. and A.S.; supervision, P.P. and P.K.D.M.; project administration, P.P. and A.K.N. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research received no external funding.

**Institutional Review Board Statement:** Not applicable.

**Informed Consent Statement:** Not applicable.

**Data Availability Statement:** The data used to support the findings of this study are available from the corresponding author upon request.

**Acknowledgments:** The authors are grateful to the Honourable Director, ICAR—the National Rice Research Institute, India; the Honourable Vice-Chancellor, Raiganj University, India; the Department of Biotechnology, Government of India (BT/PR36476/ NNT/28/1723/2020) and Project no 2.7 (ICAR-NRRI, Cuttack) for support. DM is grateful to the Government of West Bengal, India, for a Swami Vivekananda Merit Cum Means Ph.D. Scholarship (WBP191584588825). The authors wish to extend special thanks to A. Anandan, Principal Scientist (Genetics and Plant Breeding), ICAR-NRRI, Cuttack, for providing seeds and support for this experiment. The research article is a part of Debasis Mitra's Ph.D. research programme, which was supervised by P. Panneerselvam, Principal Scientist, CPD, ICAR-NRRI, Cuttack, and Pradeep K. Das Mohapatra, Associate Professor and Head, Department of Microbiology, Raiganj University, India.

**Conflicts of Interest:** The authors declare no conflict of interest.

#### **References**


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