Response of Nagpur mandarin (Citrus reticulata Blanco) to high density planting systems

High density planting system i.e. accommodating a higher number of plants than routine in a given area is an innovative agro-technology to increase yield and thereby early net returns. Due to conventional wide spacing plantation in Nagpur mandarin (Citrus reticulata Blanco), the land remains unutilized as the plant canopy gradually increases over the years. In the present study, Nagpur mandarin (Citrus reticulata Blanco) budded on Rangpur lime rootstock was evaluated under six different planting spacings. It was observed that the organic carbon (1.10–1.82%) and major nutrients viz. N (309–430 kg ha−1), P (20–54 kg ha−1) and K (291–810 kg ha−1) increased vis-à-vis plant density and was highest under 2 × 2 m spacing. Plants were tallest at 2 × 2 m spacing with the higher PAR interception (88.2) and the lowest leaf area index (1.09). Fruit yield on area basis, under 2 × 2 m spacing was 26, 7.1, 4.6 times more as compared to conventional plantation during the first, second and third year, respectively. At fifth year of crop harvest, the highest B:C ratio (6.36) was recorded in 6 × 3 m followed by 4 × 2 m and 2 × 2 m.

. Effect of planting density on soil fertility status. Means with at least one letter common are not statistically significant using TUKEY's Honest Significant Difference at 5% Control: Planting at 6 m (row to row) × 6 m (plant to plant) spacing (tree density-277 plants ha −  Irrespective of the planting densities, the gradual increase in all the growth parameters was observed. Plant height ranged between 1.77 and 4.16 m during the year 2015-16 to 2019-20 and was significantly high in all the treatments over the years (Fig. 1). Plant height decreased with increase in plant spacing. Under conventional spacing, plant height increased gradually. It was found that for want of ample sun light, the plants in close distance grew taller than plants of wide distance plantings. The columnar growth of plants in close spacing was due to poor light interception while plants in wider spacing had lateral and balanced growth due to sufficient space for light interception 15,16 . Plant canopy volume is a result of photosynthetically active radiations (PAR), nutrient uptake and irrigation in combination with pests and disease infestation. After third year, large variations were observed in plant canopy volume (1.05-29.18 cu m) (Fig. 2). During fourth year (2015-16), canopy of plants in UHDP-2 system   In Kinnow mandarin also the higher growth of plants was observed in conventional planting system (6 × 6 m) than in the high density i.e. 6 × 3 m planting 17 .
In various treatments the average stock girth ranged between 18.34 to 46.18 cm (Fig. 3). However, during initial year the results were non-significant. Plants grown under UHDP-1 (4 × 2 m spacing) recorded maximum stock girth (21. . Results revealed no direct relationship between stock girth and canopy volume of the plants. The trees in the conventional planting systems were found to acquire incremental trend of higher stock girth than plants of high and ultra-high density planting. The increased stock girth was to support higher leaf biomass and plant spread of conventional system 16,18 . Physiological parameters (Intensity of PAR light, leaf area index and chlorophyll content). Photosynthetically active radiations (PAR) in terms of photosynthetic photon flux density (PPFD) recorded highly significant variation in all East (97-160), West (96-173), North (78-174) and South (77-136) directions of the plants ( Table 2). The highest PAR in all the directions (136-174) was observed in plants grown under control (6 × 6 m spacing). It revealed that there was the lowest solar radiation utilization per unit of land in conventional spacing. Whereas, significantly low intensity of PAR in all directions of plants was found in UHDP-1 (77-101) and UHDP-2 (96-99) spacing treatments. Optimum light interception can be defined as a level of light intercepted by an orchard system above or below which, the economic yield will be reduced 19 . It indicated that there was the highest sunlight utilization per unit area in ultra high density plantings, which could be due to higher coverage of leaf area than control. Plants grown under these treatments facilitated better utilization of solar radiation and increased photosynthetic efficiency of the plant 6 .
The 6 × 3 m spacing occupied the available space between plant to plant direction (3 m) by the dense and vigorous plant canopy and recorded maximum (1.35) leaf area index ( Table 2). The space in the row to row direction (6 m) facilitated light interception that in turn, could have resulted in the higher number of well-developed leaves. Harnessing water, fertilizers, sunlight in this treatment might have translated into increased photosynthesis and thereby higher chlorophyll contents (0.71) than control ( Leaf nutrient contents. Pooled data showed that leaf N, P and K content significantly varied from 2.32-2.44%, 0.09-0.10% and 1.64-1.79%, respectively and was maximum in the plants spaced at 6 × 3 m (Table 3). www.nature.com/scientificreports/ Micronutrient contents in the leaves also varied significantly by different treatments of planting. All the nutrients were above optimum range in all the treatments. Pooled data also revealed that leaf Cu, Fe, Mn and Zn contents varied from 11.95-14.3 ppm, 116.7-133.5 ppm, 67.2-76.9 ppm and 15.5-20.51 ppm, respectively (Table 3). Maximum leaf contents of Cu and Mn was observed in 6 × 3 m spacing while Fe and Zn were observed in 4 × 4 m spacing. Improved physical as well as biological health of the soil due to increased organic carbon could have resulted in mobilization of immobile nutrients and helped in increasing their uptake by the plants. Increased nutrient uptake with the increased organic carbon was reported in several fruit crops viz. mandarin 20 , guava 21 , sweet orange 22 and pomegranate 23 . Almost all the nutrient contents in ultra-high density and high density plantings were low. It was in spite of higher organic carbon contents in these treatments. Periodical pruning and training of the threes could have removed considerable amounts of nutrients in various plant parts. Moreover, substantial amount of nutrients could be required to produce nearly three times more fruit yield per unit area as compared to the plants grown under conventional system of plantation, thereby, reducing nutrient contents in leaf in long run 6 .
Incidence of insect-pests. Citrus leaf miner, citrus psylla, aphids and mites were observed to be the most important pests of Nagpur mandarin during the different growth phases from 2013 to 2020 (Table 4). The data were collected coinciding with different periods of new flushes during pre-bearing period so as to get exact picture of pest incidence, without interference of fruiting seasons. It was found that citrus leaf miner remained active during all the flushing seasons irrespective of spacing. Maximum incidence of leaf miner was observed in closer spacing of UHDP-2 with mean value of 40.07% over a period of six years while wider spacing of 6 × 6 m recorded significantly lower infestation (18.46%). The psylla population also followed similar trend during the seasons and varied from 27.59 per 5 cm shoot in UHDP-2 to 6.44 per 5 cm shoot in control (Table 4). Pooled mean data indicated that the closer spacing had significantly highest aphid population (30.58 shoot −1 ). Similar trend was also observed in the case of mites (Table 4). There was significant increase in sucking pests' infestation and was inversely related to planting distance. It was evident that high density spacings proportionately favored the insect infestation. Narrow planting distance   24,25 . The pruning operation in high density systems accelerates the new growth that attracts leaf miner and the intensity of infestation remains in ratio of area and intensity of pruning 26,27 . It is now established that in citrus, pruning is needed for the high density planting and therefore, it was felt that the tree structure of a hedge of maximum 2.5 m height along with 0.7 m in the base of the trunk and 1 m width that is free of lateral canopy would be needed 28 . Also, to have such desired canopy, customized mechanization should be contemplated on priority. Ultra high density planting system (4.2 × 0.9 m, 4.2 × 1.2 m, 4.2 × 1.5 m, 4.2 × 1.5 and 2.5 × 2.5 m) and high density planting system (5.0 × 2.5 m, 5.0 × 5.0 m and 7.5 × 5.0 m) in mango recorded higher incidence of sucking insect pests like leaf hopper 24,25 . Also, fruit fly damage was comparatively less in conventional planting system 10 × 10 m 24 . Similarly, incidence of thrips, mealybug, aphids were observed significantly high in ultrahigh density plantation of pomegranate 29 .
Fruit yield and quality. Early fruiting (2015-16) was observed under UHDP-2 (2500 plants ha −1 (Fig. 4). Afterward, the highest yield per plant was observed in conventional 6 × 6 m spacing treatment. The canopy volume of the plants under this treatment was increasing, while the growth restricted high density systems as plants occupied almost all the allotted space. As such, significantly lowest per plant yield was obtained in close spacing plants as compared to conventional spacing treatments. However, on area basis, there was vast difference in fruit yield (1.2-85.5 t ha −1 ) amongst the treatments (Fig. 4). Under UHDP-2 treatment, fruit yield (36.2 t ha −1 ) was more than six folds as compared to conventional 6 × 6 m (5.1 t ha −1 ) during 2016-17. The difference in yield on area basis amongst treatments became narrow over the years and during 2019-20 the yield was just two folds more in the UHDP-2 than conventional treatment.   www.nature.com/scientificreports/ The increase in plant density increased the yield per unit area. Better utilization of solar radiation, increased nutrients, water and photosynthetic efficiency facilitated the increase in yield. As such under UHD, the largest amount of fruit bearing area per unit volume could be attained if the trees are planted as narrow hedges at about 1.8 m spacing with tree height and distance between rows adjusted to allow sufficient light 4,6 . Higher yields in close spacing confirms the findings of Nawaz et al. 16 , Wheaton et al. 30 and Azevedo et al. 31 found that there is an inverse relationship between tree spacing and yields. Closely spaced trees were reported to harness higher amount of solar radiations to synthesize photosynthates, which directly contribute to the growth, higher number of leaves, branches that proved beneficial towards increased sink strength resulting in higher yield in Kinnow 32 . However, moderate densities than higher density of fewer than 1000 trees per ha were preferred for Florida conditions 33 . The fruit quality parameters showed that various spacing treatments significantly influenced quality of fruits in terms of fruit breadth (72.58-75.61 cm), fruit length (65.35-69.69 cm), peel thickness (2.65-2.90 mm), juice content (43.08-45.71%), total soluble solids (TSS) contents (9.15-9.57 0 Brix) and juice acidity (0.75-0.81%) while, TSS:acidity ratio in fruit juice showed non-significant variation (Table 5). Fruit length and fruit breadth were significantly higher in the fruits of the plants grown at normal spacing of 6 × 6 m (control); however, fruit weight was non-significant. Fruit weight is a function of number polygenic characters viz. of juice content, peel thickness, number of seeds etc. Hence, the size of fruits could not only alter the fruit weight. Similar results were obtained in Kinnow where bigger size fruits with higher juice contents were recorded at 5 × 4 and 5 × 5 m spacings 34 . Fruits with thin peel were produced under UHDP-2. Fruit TSS and juice acidity were higher in HDP-1 and HDP-2, respectively. Light canopy thinning every year promoted penetration of sunlight in UHDP-2 resulting in leaves, shoots and good quality fruit development. Klerk 35 observed that higher rate of transpiration and respiration produce more carbohydrates when leaves are exposed to sunlight. Increased fruit yield without compromising quality under high density planting of fruit crops were reported by Mishra and Goswami 36 . Albeit, Gaikwad et al. 37 could not find any improvement in fruit quality of mango by reducing planting distance. After harvest, skirting and light hedging was recommended as correctly skirted trees appear to "reallocate" flowering and fruiting to remaining parts of the canopy 38,39 . Findings indicate that plant density and prunning practices for canopy management have certain effect on fruiting and fruit quality that warrants detailed further study. B:C ratio. Though fruit yield was obtained during fourth year of plantation (2015-16), monetary returns were less than the cumulative expenditure incurred towards initial plantation and yearly maintenance cost. B:C ratio was below one (0.18-0.93) in various treatments but it was close to 1 (0.93) at UHDP-2 at the first harvest itself (Table 6). At UHDP-2 yield was 15 times more than the conventional system (6 × 6 m) and benefit with respect to cost was equal. During next year (2016-17), except conventional method of plantation, profits were obtained under all HDP and UHDP treatments as indicated by B:C ratio, which was above one and the highest (3.14) was under UHDP-2. During sixth year after plantation (2017-18) the lowest profit (B:C ratio 2.38) was recorded under control and highest (B:C ratio 5.26) in UHDP-2. At the end of the experiment (2019-20), the highest B:C ratio was recorded in 6 × 3 m (B:C ratio 6.36) followed by 4 × 2 m (B:C ratio 6.09) and 2 × 2 m (B:C ratio 6.04) spacing treatment. It was well evident that the increased cost of cultivation in high density planting systems than control was due to additional cost incurred for field preparation to accommodate higher number of plants per unit area and increased plant population. Therefore, although there were higher gross returns in high density plantings during initial harvests than control, it could not be translated in net returns. Wheaton et al. 33 was of opinion that there was little advantage in high tree density (2020 trees ha −1 ) than moderate densities of fewer than 1000 trees ha −1 in Hamlin and Valencia oranges, 'Murcott' tangor and 'Redblush' planted on 15 rootstocks and own-rooted cuttings under high density planting. In Afourer mandarin and Navel orange, high density planting (952 plants ha −1 or 600 plants ha −1 ) had higher investment initially but they produced higher yields in the early years and could pay-off the debt resulting in quicker cash flow breakeven, two years earlier than traditional (440 plants ha −1 ) density 40 .

Conclusions
The present study attempted to ascertain effects of various plant densities on horticultural, physiological and fruit quality parameters and yield of Nagpur mandarin along with B:C ratio. It was possible to manage the canopy of plants planted under extremely close spacing of 2 × 2 m (2500 plant ha −1 ) and translate it into getting early and high yields. The yield obtained under 2 × 2 m spacing treatment was twenty-six times higher than control from the very first fruiting. Albeit, monetary returns were not translated into net profitability during first year due to very high initial cost incurred on this treatment. Higher returns with increasing B:C ratio was obtained under this treatment from 2nd to 4th years of fruiting. In 5th year 6 × 3 m spacing treatment (555 plants ha −1 ) was found economically superior. The management practices helped to retain the quality of fruits and yield which could not be adversely affected by increased insect-pest and disease infestation by virtue of close planting. It is important to observe performance of these plantations further for next 4-5 years from commercial point of view. This study has established practical feasibility and economic superiority of high and ultra-high density planting of Nagpur mandarin over conventional system of plantation. In high density plantation technique, canopy management is equally important in bearing citrus trees to have better interception of PAR and aeration, strong sap flow for fruiting and supporting branches for convenient harvesting. Last but not the least, effective alternative to harness available space for increasing net return for first few years of fruiting is high density planting. However, the synchronization of spacings vis-à-vis growth pattern needs to be standardized and mechanization for canopy management would be an area for future work to ascertain optimum economic returns.  www.nature.com/scientificreports/ vermicompost, 150 g N, 50 g P and 25 g K per plant which was doubled for second year and tripled for third year. Nutrient application was fixed at 2 kg vermicompost, 600 g N, 200 g P and 200 g K per plant from fourth year onwards. Nutrient dose was applied through fertigation using diammonium phosphate, urea, muriate of potash and phosphoric acid fertilizers. For application of micronutrients, foliar sprays were applied on fully expanded leaves during new flush seasons i.e. July-August and February-April. Field was irrigated on alternate days using drip irrigation at cumulative irrigation as equivalent to 0.70 E-Pan with 90% irrigation efficiency of drip. Balance sheet method was used to calculate the effective rainfall from the actual rainfall received. Accordingly, daily water requirement was met out.

Materials and methods
Standard package of practices was adopted for the control of various insect-pests and diseases 41 . During initial years, plants were raised under various treatments to provide proper shape. While training, branches were oriented in such a manner that the plant canopy at central place would facilitate better light penetration and distribution. Up to 75 cm from ground no shoots were allowed to grow. Thereafter 3-4 main scaffold branches were grown followed by secondary and tertiary branches. At 2 × 2 m spacing (UHDP-2), between two rows, duct space of 60 cm was created for proper access after 3rd year with light pruning. This light canopy thinning was required in this treatment every year after third year. Dried twigs were removed after harvesting every year followed by fungicidal sprays. Leaf nutrient contents. Leaf samples (5-7 months old leaves) were collected every year from non-bearing shoots of each selected plant in each treatment 46 and analyzed for different macro-and micro-nutrient status. Samples were washed thoroughly in water, liquid soap, acidic water and glass redistilled water sequentially, air dried followed by oven drying at 70 °C till samples maintained constant weight. Composite leaf samples were used for analyzing total macro (N, P and K) and micronutrients (Fe, Zn, Mn and Cu) using standard methods described in Chapman and Pratt 47 .

Incidence of insect-pests. Incidence of insect pests of citrus viz. citrus leaf miner (Phyllocnistis citrella
Stainton), Asian Citrus Psylla (Diaphorina citri Kuwayama), aphids (Aphis sp., Toxoptera sp.) and red spider mites (Eutetranychus orientalis Klein) were recorded during flushing seasons i.e. Ambia (January-February), Mrig (June-July) and Hasta (September-October) at fortnightly intervals. Four twigs of 15-20 cm length from North, South, East and West direction of each tree were observed and likewise 10 replications were taken to calculate infestation (in case of citrus leaf minor and mites). Population count per 5-10 cm twig for psylla and aphids and counts per tapping for thrips was taken. Similarly trap catch of citrus leaf miner adult males using sex pheromone lure in a delta trap 48 were also installed for monitoring the field population during flushing season.
Fruit quality and yield. The fruit yield data from 2015-16 to 2019-20 was recorded on tree basis (kg tree −1 ) and area basis (t ha −1 ). Also, physico-biochemical properties of fruit viz. Total soluble solids (TSS), acidity, juice content and peel thickness were recorded with standard procedures 49 . B:C ratio. Benefit:Cost (B:C) ratio was calculated out to unravel economic viability of the technology. The cost incurred towards land preparation, pit digging, irrigation and budling (budded seedlings with selected scion bud by shield budding) during first year was Rs. 55,942, 73,984, 85,281, 1,29,812 and 2,10,875 in planting at 6 × 6 m, 6 × 3 m. 4 × 4 m, 4 × 2 m and 2 × 2 m spacing. It cumulatively increased to Rs. 1,63,875, 2,34,751, 2,45,331, 4,05,662 and 8,33,325, respectively until fruit bearing stage (fourth year). B:C ratio was calculated for treatments over conventional planting system of 6 × 6 m. spacing. The B:C ratio greater than one (> 1) represents profit, as the benefit will be more than the cost incurred.
Statistical analysis. The present study which had five treatments replicated four times, was conducted in randomized block design. Significant differences among various treatments during the years of experimentation were pooled for statistical analysis. Tukey's HSD test was done by using SAS software for shortest significant range tests 50 .