The exploration of factors associated with citrus fruit non-chilling rind pitting : the case study of highly prone "Benny" valencia variety

Abstract

The rind physiological disorders incidence such as rind pitting is a challenge to the citrus industry as it affects appearance; and ultimately, acceptability and purchase in both local and international markets. Although the internal quality is not directly affected by rind pitting, fruit damaged by this disorder are rejected in the fresh fruit market. The susceptibility to this disorder varies among citrus fruit cultivars. Other factors impacting rind physiological disorders include; pre-harvest environmental conditions and postharvest storage conditions. However, the main cause of this disorder is still unknown. In South Africa, ‘Benny’ valencias are the most prone orange cultivars to rind pitting disorder within the sweet-orange-type. Therefore, the aim of this study was to investigate production site and postharvest treatments’ effect on physico-chemical, biochemical properties, antioxidants, rind soluble sugars and gene expression in relation to rind pitting development of ‘Benny’ valencia citrus fruit. During 2016 and 2017 seasons, the study was conducted, whereby, ‘Benny’ valencia citrus fruit were harvested from Tzaneen, Groblersdal and Musina in South Africa. After harvesting, the fruits were transported to the Agricultural Research Council- Tropical and Subtropical Crops (ARC-TSC) postharvest laboratory in Nelspruit for sorting, grading treatment, cold storage and post-storage quality evaluation. After sorting and grading, fruits were subjected to the following treatments: T1 = no wax plus dehydration, T2 = wax plus dehydration and T3 = wax plus no dehydration. Dehydrated treatments were applied for 3 days at relative humidity of ±45%, thereafter, fruit were stored at -0.6 and 4.5°C for 28 days plus 7 days shelf-life. After removal from cold storage plus 7 days shelf-life, fruit were analysed for rind pitting incidence (RPI), weight loss percentage (WL), firmness, total electrolyte leakage (TEL), total soluble solids (TSS), titratable acidity (TA) and TSS: TA ratio. xxvi

Afterwards, fruit were peeled to remove flavedo, thereafter; the flavedo peels were freeze-dried, milled and stored at -21°C for further physiological analysis. Freeze dried flavedo peel was analysed for total flavonoids, total phenolics, vitamin C, soluble sugars (glucose, fructose and sucrose), antioxidant assays (FRAP, DPPH, ABST and ORAC) and genes. The results showed that rind pitting incidence was high on fruit subjected to wax plus no dehydration across all storage temperatures and production sites. Furthermore, results showed that fruit harvested from Musina exposed to T1 had higher incidence of rind pitting than those from Groblersdal and Tzaneen, irrespective of storage temperature. Meanwhile, fruit harvested from Musina had the highest TEL when compared with Groblersdal and Tzaneen irrespective of treatments and storage temperatures. A significantly higher (P<0.05) WL was observed in Musina fruit harvested from Tzaneen exposed to T3 at both storage temperatures. Moreover, increased TSS was observed after storage across all production site and postharvest treatments. The study showed that production site and postharvest treatments had a significant influence on rind pitting and total electrolyte leakage. Additionally, fruit treated with no wax + dehydration was found to be more susceptible to rind pitting. However, fruit sourced from Tzaneen had significantly (P<0.0001) high TPC and TFC, irrespective of postharvest treatments therefore, low rind pitting incidence. While rind vitamin C was higher in fruit from Groblersdal when compared with Tzaneen and Musina. However, low RPI was also observed in fruit sourced from Groblersdal. Fruit from Musina subjected to wax plus dehydration had higher RSA and low RPI at both temperatures when compared with fruit sourced from Groblersdal and Tzaneen. Therefore, wax plus dehydration resulted in low rind pitting with an increased accumulation of rind biochemical concentrations, xxvii

irrespective of cold storage temperatures. These results suggested that there is a link between rind pitting and rind biochemical concentrations in the ‘Benny’ valencia citrus fruit. Moreover, fruit from Musina subjected to wax plus dehydration had higher antioxidant measured by DPPH and low RPI at both low storage temperatures when compared with fruit harvested from Groblersdal and Tzaneen. Furthermore, fruit harvested from Musina and treated with no wax plus dehydration, thereafter, stored at -0.6°C had low RPI with high antioxidant activity measured by FRAP than Tzaneen and Groblersdal regions in both seasons. With respect to sugars, fructose was not significantly (P<0.05) affected by production sites, postharvest treatments and cold storage temperature, hence, the low pitting incidence. The highest glucose was observed in fruit harvested from Groblersdal, irrespective of treatments and cold storage temperatures when compared with those from Tzaneen and Musina, low RPI was also observed in fruit harvested from Groblersdal. However, fruit harvested from Groblersdal treated with wax plus dehydration and stored at 4.5°C had higher sucrose and low RPI when compared with Tzaneen and Musina. Moreover, this study suggested that soluble sugars in ‘Benny’ valencia flavedo during cold storage is involved in rind pitting tolerance mediated by wax plus dehydration treatment. Three homologic genes: CsCP gene; CsNAC-domain protein gene; CsCP-F gene; were chosen to examine the relationship between their expression and citrus rind pitting through quantitative RT-PCR analysis in pitting and no-pitting fruits. Results showed that the expression of CsCP, CsNAC and CsCP-F genes were all higher in the pitting rind fruit harvested from Tzaneen and low in fruit with low pitting. Groblersdal and Musina fruit had low expression of genes and low rind pitting was observed. Therefore, findings suggested that CsCP, CsNAC and CsCP-F genes may xxviii

be linked to non-chilling rind pitting and could serve as targets for future investigation. Generally, the overall results obtained in this study provided an understanding into the previous unknown complexities of citrus non-chilling rind pitting. Moreover, the study revealed that the studied factors had an influence on non-chilling rind pitting and physico-chemical properties of ‘Benny’ valencia citrus fruit. In addition, postharvest treatments resulted in low non-chilling rind pitting with an increased accumulation of rind biochemical concentrations. The fruit with high antioxidant capacity were found to be tolerant to rind pitting, whereas, fruit with low antioxidant capacity were found to be susceptible to rind pitting. Furthermore, soluble sugars are believed to be involved in the defence mechanisms against non-chilling rind pitting in the fruit. Gene expression changes also provided clues about the possible mechanisms involved in non-chilling rind pitting development.