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Tan, Guang Heng (2025) Volatile profiling and microbiome analysis of papaya fruit rot caused by Colletotrichum gloeosporioides, Lasiodiplodia theobromae, and Fusarium solani and postharvest control strategies. PhD thesis, University of Nottingham.

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Abstract

Papaya (Carica papaya) is a vital crop, particularly in tropical countries like Malaysia, due to its rich nutritional content, economic value, excellent flavour, and applications in drug development and industrial processing. The export of fresh papaya and processed papaya products has become a significant source of income for papaya-growing countries, especially in regions where the demand for tropical fruit is high. However, despite these benefits, papaya's climacteric nature makes it highly susceptible to rapid deterioration, primarily due to severe infections by various pathogens that develop both epiphytically and endophytically during postharvest handling. These irreversible decays lead to substantial changes in overall fruit quality and contribute to significant postharvest losses in papaya-producing countries, including Malaysia. To mitigate these losses, understanding the causal agents of these infections is crucial. Early diagnosis and identification methods have been introduced and implemented to detect the spread of diseases, enabling timely interventions. Additionally, various control strategies are applied to inhibit and delay disease establishment through different modes of action. This thesis aims to elucidate the volatile profile of ‘Sekaki’ papaya during ripening and its changes when infected with Lasiodiplodia theobromae, Fusarium solani, and Colletotrichum gloeosporioides. Furthermore, the microbiome of the whole papaya fruit is analysed using next-generation sequencing (NGS) to understand the impact of fruit ripening and rotting on microbial shifts within the fruit. In addition, the potential of an underutilized crop, Garcinia atroviridis, is explored for its ability to inhibit the growth of L. theobromae, F. solani, and C. gloeosporioides at different concentrations. An in-vivo analysis of G. atroviridis fruit extract against C. gloeosporioides is also conducted, along with an assessment of its impact on the physico-chemical qualities of papaya when applied at varying concentrations.

Solid-phase microextraction coupled with gas chromatography- mass spectrometry (SPME-GC-MS) analysis was conducted on ripe and unripe papaya fruit revealing significant differences in their volatile profiles at different ripening stages. Principal component analysis (PCA) accounted for 20.0% and 31.0% of the variance in the first two components, respectively. The identified volatiles, including alkanes, alcohols, and esters, could serve as potential biomarkers for fruit ripening. Specifically, 4-methyl-1-hexanol and 2,4-dimethylheptane were detected exclusively in unripe fruit, while oxalic acid, butyl propyl ester and pentyl propionate were found in ripe fruit.

Further GC-MS analysis was conducted on papaya fruits infected with C. gloeosporioides, L. theobromae, and F. solani to identify potential biomarkers associated with each pathogen. Based on disease incidence, severity, and PCA, days 1, 3, and 5 were determined to be optimal for volatile profiling. Significant differences in the volatile profiles were observed across these time points. A total of 61, 70, and 68 volatile compounds were detected in samples infected with C. gloeosporioides, L. theobromae, and F. solani, respectively. Methylglyoxal and 1-butanol emerged as potential biomarkers for C. gloeosporioides infection, while 1-pentanol, 1-heptanol, and L-lactic acid were identified for L. theobromae infection, and 1-octanol and β-Phellandrene for F. solani infection.

Additionally, Next- generation sequencing (NGS) was employed for metagenome analysis to detect microbial communities in papaya fruits at different ripening stages and elucidate potential interaction with the host. Alpha diversity analysis revealed a significantly higher bacterial diversity in unripe samples, while the fungal microbiome showed no significant difference between the stages. Beta diversity analysis indicated a significant difference in microbial abundance between unripe and ripe samples, which was further supported by relative abundance data. Specifically, bacterial families Acetobacteraceae and Pseudomonadaceae were more abundant in unripe samples, while Erwiniaceae and Caulobacteraceae were predominant in ripe samples. For the fungal microbiome, Cladosporiaceae, Pichiaceae, and Saccharomycetaceae were significantly more abundant in unripe samples, while pathogenic families Botryosphaeriaceae, Glomerellaceae, and Nectriaceae were more prevalent in ripe samples. Overall, unripe and ripe papaya samples exhibited distinct bacterial and fungal microbiomes in terms of composition, diversity, and abundance.

Metagenome analysis using next-generation sequencing was conducted to investigate microbial shifts during papaya fruit rotting and to understand the roles of each microbe involved. The results revealed that diseased fruits exhibited higher diversity in both bacterial and fungal microbiota compared to healthy samples. Taxonomic characterization identified several pathogenic bacteria and fungi in healthy samples, including Erwinia sp., Enterobacter cloacae, Fusarium solani, Colletotrichum okinawense, and Lasiodiplodia sp. In contrast, the microbiota of diseased samples included various antagonistic microbes with different modes of action against pathogens, such as Pichia kluyveri, Issatchenkia orientalis, Rhodotorula mucilaginosa, and Enterobacter hormaechei. These findings provide valuable insights into the microbial dynamics during fruit rot and highlight the potential of developing antagonistic microbes as biocontrol agents to combat postharvest diseases.

The in-vitro and in-vivo effects of Garcinia atroviridis fruit extract on postharvest diseases were also investigated over an 8-day storage period. The results demonstrated that a concentration of 15 mg/mL of G. atroviridis fruit extract exhibited the highest inhibition capacity against the mycelial growth of Colletotrichum gloeosporioides, Lasiodiplodia theobromae, and Fusarium solani in-vitro. Similarly, the same concentration was able to completely inhibit the development of anthracnose on fruit caused by C. gloeosporioides until day 6, based on assessments of disease incidence and severity.

When evaluating postharvest quality across different treatments, no significant differences were observed in any of the parameters measured, although changes were noted as the fruit ripened from day 0 to day 8. A similar trend was observed in antioxidant analysis, confirming that the fruit extract did not interfere with the normal ripening process while effectively inhibiting the growth and development of phytopathogens. These findings suggest that G. atroviridis fruit extract has the potential to serve as an alternative postharvest strategy for managing postharvest diseases, offering a solution to the limitations of existing technologies.

Item Type:	Thesis (University of Nottingham only) (PhD)
Supervisors:	Ali, Asgar Siddiqui, Yasmeen Supramaniam, Christina Vimala
Keywords:	papaya, postharvest loss; fungus, gas chromatography-mass spectrometry (GC-MS); next-generation sequencing (NGS); Garcinia atroviridis; postharvest quality
Subjects:	S Agriculture > SB Plant culture
Faculties/Schools:	University of Nottingham, Malaysia > Faculty of Science and Engineering — Science > School of Biosciences
Item ID:	81447
Depositing User:	Tan, Guang Heng
Date Deposited:	26 Jul 2025 04:40
Last Modified:	26 Jul 2025 04:40
URI:	https://eprints.nottingham.ac.uk/id/eprint/81447

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