Elsevier

Scientia Horticulturae

Volume 193, 22 September 2015, Pages 47-54
Scientia Horticulturae

Morphology, photosynthesis, and internal structure alterations in field apple leaves under hidden and acute zinc deficiency

https://doi.org/10.1016/j.scienta.2015.06.016Get rights and content

Highlights

  • We elucidated a possible damage mechanism of Zn deficiency in field apple leaves.

  • Leaves experiencing hidden Zn deficiency were used as a deficient transition state.

  • The reductions in PS II activity were first detected under hidden Zn deficiency.

  • Chloroplast disintegration and PS II damage occurred as Zn deficiency aggravated.

  • Some chlorophyll fluorescence parameters may be used to identify Zn deficiency.

Abstract

In the field, apple trees (Malus domestica L.) do not exhibit visible deficient symptoms when affected by hidden zinc (Zn) deficiency, while internal injury in the leaves may prior occurred. We named the leaves on Zn deficient trees without and with visible rosette symptoms as ‘hidden rosette leaves’ and ‘rosette leaves’, respectively. The healthy leaves of normal trees were named ‘normal leaves’ and selected as the control. The changes in leaf photosynthesis, physiology and internal structure were analyzed to elucidate the damage mechanism of Zn deficiency in apple leaves. We found that the reduction in photosynthesis under Zn deficiency was primarily due to non-stomatal limitation, in particular the changes in photosystem II (PS II). Zinc deficiency significantly reduced chlorophyll content in the hidden rosette leaves without obvious changes to chloroplasts and grana number. The intrinsic efficiency of PS II (Fv′/Fm′), actual photochemical efficiency of PS II (ϕPS II) and electron transport rate (ETR) were all significantly decreased in both hidden rosette leaves and rosette leaves, while a further significant increase in the minimal fluorescence (Fo) along with reductions in maximum photochemical efficiency of PSII (Fv/Fm) and photochemical quenching (qP) occurred in rosette leaves. These results indicate that the decrease in chlorophyll content and inhibition of PS II light-harvesting activity may account for the photosynthesis reduction during the initial stage of Zn deficiency. As Zn deficiency aggravated, cellular deficient characteristics like abnormal cell arrangements and disorganization of thylakoids became more exaggerated; visible external symptoms of Zn deficiency appeared with internal alterations, including photosynthetic apparatus damage accompanied by subsequent chloroplast and grana disintegration. We concluded that the PS II activity was relatively sensitive to Zn deficiency, hence the chlorophyll fluorescence parameters like Fv′/Fm′ and ϕPS II may be used as predictive indictors for hidden Zn deficiency on apple trees.

Introduction

Zinc deficiency is the most widespread micronutrient deficiency in soil and crops (Alloway, 2008). It is estimated that approximately 30% of the agricultural soil in the world are Zn deficient (Cakmak et al., 1996). In China, Zn deficiency affects more than 48.6 Mha of soil, primarily calcareous soil distributed in Northern China (Zou et al., 2008). Apple plants, as the main economic fruit trees planted in China, are considered to be very sensitive to Zn deficiency (Rashid and Ryan, 2004). It was recorded that China is the largest producer of apple in the world and Shandong Province accounts for over half of China’s apple exports (Miyata et al., 2009). Zinc deficiency is now very common in apple orchards in China, especially in Shandong Province, resulting in severe losses in yield and deterioration in fruit quality.

In the field, Zn deficiency occurs naturally, and the symptoms of deficiency develop depending on the degree of stress (Brennan et al., 1993). Visual symptoms of Zn deficiency only occur in cases of relatively severe deficiency (Alloway, 2008) and are described as stunted growth, small leaves, chlorosis, rosette and even shoot die back (Swietlik, 2002, Sharma et al., 2013). Rosette and little leaf at the shoot tip are the most characteristic symptoms of Zn deficiency in fruit trees (Swietlik, 2002). In marginally Zn deficient soils, yield and quality can be affected without the obvious appearance of symptoms; this is called ‘hidden’ (or ‘latent’) deficiency (Alloway, 2008, Alloway, 2009). Some studies have reported that dry matter production is reduced by 40% or more by hidden Zn deficiency (Carroll and Loneragan, 1968). Given these circumstances, the understanding to the intrinsic changes in the physiological processes of leaves affected by hidden Zn deficiency may be helpful to determine the initial damages caused by Zn deficiency along with the final development of deficiency symptoms.

Zinc deficiency depresses plant growth especially shoot growth and chlorophyll concentrations (Balakrishnan et al., 2001), leading to chlorosis in leaves along with abnormalities in leaf structure and ultimately affecting chloroplast structure (Kim and Wetzstein, 2003). A decline in the net photosynthetic rate (Pn) induced by Zn deficiency is often observed. The reasons for this decline may be associated with a decrease in stoma conductance (Hu and Sparks, 1991), a decrease in carbonic anhydrase activity (Sasaki et al., 1998), a reduction in carbon fixation (Sharma et al., 1995) or damage to the photosynthetic apparatus (Balakrishnan et al., 2001, Chen et al., 2008). However, the mechanism by which Zn deficiency damages photosynthetic activity, especially in field-grown apple leaves, is not yet known.

Although Zn deficiency can alter a variety of physiological processes, the primary site of its effect is probably the cell membrane systems (Candan and Tarhan, 2003). Severe Zn deficiency can induce a general disorganization of chloroplast thylakoids (Henriques, 2001) and extensive degeneration of chloroplast membranes (Chen et al., 2008). The loss of chloroplast membrane integrity and thylakoid disorganization can affect the photochemical processes on thylakoids, thus inhibiting the biophysical processes of photosynthesis (Cakmak, 2000, Daub et al., 2013). Zinc deficiency also reduces the actual photochemical efficiency of PS II (ϕPS II) and affects the activity of some related enzymes (Tavallali et al., 2009, Billard et al., 2015). However, most of the above mentioned reports were conducted under severe Zn deficiency or after the formation of typical deficiency symptoms, when noticeable damage to chloroplasts and photosynthetic centers in affected leaves has already occurred. Little information is available about the structural and functional alterations of leaves at the transitional stage of hidden Zn deficiency.

The aim of this study is to elucidate the mechanism by which Zn deficiency affects the physiological function and internal structure of apple leaves at various Zn levels under field conditions. With this aim, we use pigment and photosynthesis analyses as well as microstructure and ultrastructure observation to characterize the physiological and structural changes induced in apple leaves by Zn deficiency at latent and deficient levels. Chlorophyll fluorescence was also investigated to better understand alterations of the photosynthetic center on chloroplast membranes under different Zn deficiency conditions. This work presents detailed microscopic evaluation of Zn deficiency in apple leaves and elucidates how progressive Zn deficiency affects leaf structure and function.

Section snippets

Field site and plant materials

Experiments were conducted in 2012 and 2013 using 13-year-old ‘Fuji’ apple trees in an orchard in Chaoquan (latitude 36°14′N; longitude 116°50′E), Feicheng County, Tai’an, Shandong Province, China. In field-grown apple trees, leaves at the tops of shoots on Zn-deficient trees are more likely to appear tufted or in a rosette formation. Other leaves on the same branch or on other shoots of the same tree exhibit no visible deficiency symptoms; this is referred to as ‘hidden’ deficiency. Both

General leaf characteristics under zinc deficiency

The first characteristic reaction of plants to zinc deficiency was the reduction in shoot elongation and leaf size (Fig. 1, Table 1). The visible symptoms induced by Zn deficiency were observed in field apple trees as short internodes and small and narrow leaves with chlorosis in a rosette formation on the top of the shoot (Fig. 1). Under Zn-deficiency, HRL and RL had lower Zn concentrations and leaf areas than the NL. The reductions in Zn concentration were 31.2% and 51.6% for HRL and RL, and

Discussion

In most fruit and other crops, the critical Zn deficiency level is considered to be below 15–20 mg kg−1 dry weight of leaves (Swietlik, 2002). The HRL in this study showed no visible rosette symptoms at a Zn concentration of 14.92 mg kg−1. This indicated that the content of Zn in HRL was at the latent level; thus, the physiological differences between HRL and RL is more helpful for us to understand the progression of damage induced by Zn deficiency in apple leaves under field conditions.

In the

Conclusions

The damage of Zn deficiency on apple leaves were mainly reflected in the reduction of leaf area, chlorophyll content and net photosynthesis rate (Pn), and the alterations in internal leaf structure. Among these deficient characters, Pn showed to be very sensitively regulated by Zn. Under hidden Zn deficiency, the decrease of photosynthesis was related to the inhibition of PS II intrinsic efficiency (Fv′/Fm′), the actual PS II photochemical efficiency (ϕPS II) and electron transport rate (ETR).

Acknowledgements

This work was financially supported by the Natural Science Foundation of China (Grant No. 31372015 and 31401683), the Special Fund for Agro-scientific Research in the Public Interest (Grant No. 201103003) and the Natural Science Foundation of Shandong Province, China (Grant No. ZR2013CM022). The authors are grateful to Prof. Huiyuan Gao for help in review our English composition and to Mr. Chengjun Yin for the management of apple trees in field, and to Mr. Yuankui Guo for operation of the TEM

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