Control of Unexpected Mucor lusitanicus in Litchi Fruit by Hydrocooling with Hypochlorous Acid and Cold Storage
Round 1
Reviewer 1 Report
Comments and Suggestions for AuthorsThe manuscript focuses on a control study of Mucor lusitanicus in litchis from Taiwan and examines pre-cooling methods incorporating HClO. The overall research closely relates to practical production issues and develops techniques that are simple and feasible. However, the study still requires further improvement by the authors.
1. Lack of Consistency in the Study Design:
The manuscript primarily focuses on controlling Mucor lusitanicus on litchi fruits, based on its title. While the first half of the study investigates measures to control the pathogen, including hydrocooling integrated with HClO, it does not address the critical period between harvesting and pre-cooling where the pathogen might be affected. The second half of the manuscript shifts focus to the impact of pre-cooling with HClO on the quality of litchi fruits (although it mentions “disease incidence,” it does not specifically target Mucor lusitanicus). The lack of connection between the two parts of the study highlights a design flaw the authors currently may not be able to resolve experimentally. Therefore, it is essential that the authors acknowledge and discuss these limitations in the "Discussion" section of the paper.
2. Insufficient Introduction of Study Material:
It is inappropriate that the “2. Materials and Methods” section does not provide a detailed introduction to the litchi material used in the study—namely, the ‘Yu-Her-Pao’ variety. Additionally, the susceptibility of this litchi variety to Mucor lusitanicus should be briefly discussed in the “Introduction.” If the ‘Yu-Her-Pao’ litchi is rarely infected by the pathogen, then this study would lose its significance.
3.Mismatch Between Title and Content:
The title primarily focuses on Mucor lusitanicus in litchi fruits; however, this issue is discussed only in the first half of the manuscript, with the second half addressing litchi fruit pre-cooling methods and quality, largely unrelated to Mucor lusitanicus. Therefore, the title needs to be revised to better reflect the content of the manuscript.
4. Deficiencies in the “Abstract” and “Conclusion” Sections:
The “Abstract” does not adequately address the issue of pre-cooling time, which forms a key focus of the latter part of the manuscript. On the other hand, the “Conclusion” completely neglects the core issue of this study—the control of Mucor lusitanicus. Both sections need to be revised and improved to better align with the manuscript's key findings.
5. Formatting Issues in Tables:
In all tables, the markers for group comparisons (e.g., a, b, c) should be placed as superscripts above the respective results. The journal where the manuscript is submitted has specific formatting requirements that must be followed.
Author Response
Comments 1: The manuscript primarily focuses on controlling Mucor lusitanicus on litchi fruits, based on its title. While the first half of the study investigates measures to control the pathogen, including hydrocooling integrated with HClO, it does not address the critical period between harvesting and pre-cooling where the pathogen might be affected. The second half of the manuscript shifts focus to the impact of pre-cooling with HClO on the quality of litchi fruits (although it mentions “disease incidence,” it does not specifically target Mucor lusitanicus). The lack of connection between the two parts of the study highlights a design flaw the authors currently may not be able to resolve experimentally. Therefore, it is essential that the authors acknowledge and discuss these limitations in the "Discussion" section of the paper.
Response 1: We agree with this comment. Therefore, we revised the article to establish a clearer link between Mucor lusitanicus and hydrocooling. Please refer to the updated version, where the revisions have been highlighted in red.
Comments 2: It is inappropriate that the “2. Materials and Methods” section does not provide a detailed introduction to the litchi material used in the study—namely, the ‘Yu-Her-Pao’ variety. Additionally, the susceptibility of this litchi variety to Mucor lusitanicus should be briefly discussed in the “Introduction.” If the ‘Yu-Her-Pao’ litchi is rarely infected by the pathogen, then this study would lose its significance.
Response 2: Thank you for pointing this out. We added the basic information about the disease. Because this disease was first found in litchi fruit in Taiwan, we used DNA sequences to identify (Line100-104). Pathogenicity tests showed M. lusitanicus was able to cause decay on the wound-inoculated litchi fruit (Line291-292)
Comments 3: The title primarily focuses on Mucor lusitanicus in litchi fruits; however, this issue is discussed only in the first half of the manuscript, with the second half addressing litchi fruit pre-cooling methods and quality, largely unrelated to Mucor lusitanicus. Therefore, the title needs to be revised to better reflect the content of the manuscript.
Response 3: We agree with this comment. Due to inconsistence, we have revised the content to establish a stronger relationship between Mucor lusitanicus and hydrocooling. Please see the new version.
Comments 4: The “Abstract” does not adequately address the issue of pre-cooling time, which forms a key focus of the latter part of the manuscript. On the other hand, the “Conclusion” completely neglects the core issue of this study—the control of Mucor lusitanicus. Both sections need to be revised and improved to better align with the manuscript's key findings.
Response 4: We appreciate this comment. We added some sentences to the abstract and conclusion sections to enhance clarity. Lin20-23“The browning index and disease incidence of litchi fruit hydrocooled with an 8-hour hydrocooling delay were significantly lower than those with a 12-hour hydrocooling delay after 21 days at 5 °C, followed by 1 day at 26 °C. Therefore, hydrocooling within 8 hours of harvest is recommended for commercial scales.” Lin356-360 “To prevent M. lusitanicus, critical HClO concentrations and optimal temperatures were identified and integrated into the hydrocooling stage of the postharvest process. Taking into account the operational capacity and workflow of packinghouse, determining the maximum allowable delay for precooling while preserving product value may present challenges in practical applications.”
Comments 5: In all tables, the markers for group comparisons (e.g., a, b, c) should be placed as superscripts above the respective results. The journal where the manuscript is submitted has specific formatting requirements that must be followed.
Response 5: Agree. We have done according to the specific formatting requirements. Please see the new version.
Author Response File: 
Author Response.pdf
Reviewer 2 Report
Comments and Suggestions for Authors
The current study has proposed a method to extend the storage ability of early-maturing ‘Yu He Pao’ litchis through hydrocooling and temperature management. The paper's topic is interesting and the aim is to help maintain the shelf life of fresh fruit and vegetables. The paper is well-organized and the material and methods were described comprehensively. However, the introduction section should highlight the current gap and describe directly how the current paper can address the existing issue. In addition, the discussion should revise deeply and recent literature should considered for discussion. In fact, lack of enough discussion and comparison of the achieved output with the literature is tangible. Overall, the paper is appropriate to publish in the journal after addressing the aforementioned and following comments.
L35- The author mentioned ‘’…exacerbate pericarp browning and fruit infection during postharvest processes. This statement needs reference.
L38- describe more about the literature that has linked browning to the degradation of anthocyanins and the activity of polyphenol oxidase.
L48- Instead of ‘’ modified atmosphere (MA) by packaging’’ , you should use ‘’ modified atmosphere packaging (MAP). But I think active packaging also can be useful.
L58- mention some references after ‘’ numerous studies have documented postharvest 57 diseases in litchis..’’
L68- Although the introduction is straightforward and the authors avoid using redundant information, mentioning some literature and highlighting the current gap is recommended.
L92- Why the plates were incubated in darkness at various temperatures (1, 5, 10, 15, 20, 25, 91 30, and 35 °C). The strategy was considered based on the literature?
L127- Please clarify why the samples were transported to the lab within 1.5 h. Overall, some scenarios were considered for the trial without any reason. Please underscore why the utilized strategies were performed.
L153- here it needs to mention the juice extraction device.
L169- Table 1 presents the effect of different temperatures on the mycelial growth of Mucor lusitanicus for 7 days, but the description is not enough of why and for what reason temperature was between 5 and 0 °C. Thus, describe more.
L214- Table 5 shows the respiration rates and ethylene production of 'Yu-Her-Pao' litchis at different temperatures. This information can be useful for description of shelf life, hence, I suggest to add some information about shelf life. Also, the information about ethylene produced is strange, why the ethylene produced by the 15th d is zero!?
L264- Figure 3 illustrates the effects of hydrocooling with 40 mg L–1 HClO delay on the appearance of 'Yu- Her-Pao' litchis stored at 5 °C for 21 d followed by 26 °C for 1 d. The figure tries to show the appearance and dividing procedure, but, the figure is confusing. Please arrange each section in a better vision for more understanding. If the main aim of the figure is to divide the sample, so please remove it and explain the required information in the text.
L268-276 the content is not similar to the discussion, therefore, please remove it.
L277-280 The statement represented the observed results and the lack of appropriate discussion is tangible. The conclusion needs deep revision. Authors should mention the output of the observed results and how the output addresses the current gap considering the subject of the paper. In addition, I suggest mentioning briefly the encountered challenges in the study and suggesting some ideas to cope with the challenges and limitations for further research.
Author Response
Comments 1: L35- The author mentioned ‘’…exacerbate pericarp browning and fruit infection during postharvest processes. This statement needs reference.
Response 1: We appreciate this comment. We added three references (Line 38)
Comments 2: L38- describe more about the literature that has linked browning to the degradation of anthocyanins and the activity of polyphenol oxidase.
Response 2: We appreciate this comment. We added some sentences “Anthocyanins are hydrolyzed into anthocyanidins by anthocyanase and then oxidized by PPO, leading to the formation of brown pigments and browning of the pericarp” (Line 41-43)
Comments 3: "L48- Instead of ‘’ modified atmosphere (MA) by packaging’’ , you should use ‘’ modified atmosphere packaging (MAP). But I think active packaging also can be useful.
Response 3: Correction: modified atmosphere packaging (MAP) (Line 50)
Comments 4: L58- mention some references after ‘’ numerous studies have documented postharvest 57 diseases in litchis.’’
Response 4: We agree with this comment. We added three references (Line 73-74)
Comments 5: L68- Although the introduction is straightforward and the authors avoid using redundant information, mentioning some literature and highlighting the current gap is recommended.
Response 5: Thank you for pointing this out. We added some literature and the current gap. Several significant postharvest fungal pathogens have been identified in litchi fruit, including, Gloeosporium, and Colletotrichum, Penicillium, Botrytis, Galactomyces, Peronophythora, Gloeosporium, and Colletotrichum and Mucor spp. were repentantly isolated from 17 litchi packinghouses [5,17,18]. Recently, an unknown postharvest disease affecting litchi fruit was observed in Taiwan, particularly in fruit stored for several weeks at low temperatures during 2023 and 2024. The disease symptoms were highly similar to the mucor rot so that we tried to identify this postharvest disease and support disease control methods. In recent years, mucor rot has emerged as the dominant postharvest disease in mandarin fruit in California [19] and Mucor contribute to substantial losses in fresh agricultural products [20]. To combat this issue, natamycin has been recommended for controlling Mucor spp. in citrus, and combination with salts and heat treatments has proven effective [21]. However, no official chemical treatments are currently recommended for Mucor spp. litchi fruit during postharvest process in Taiwan. Once the pathogen infects the fruit, either in the field or during the postharvest process, it can proliferate rapidly under favorable conditions. Precooling is a critical step to inhibit pathogen growth and preserve fruit quality, as low temperatures not only suppress fungal growth but also reduce the respiration rate of horticultural produce [15,22]. (Line57-73)
Comments 6: L92- Why the plates were incubated in darkness at various temperatures (1, 5, 10, 15, 20, 25, 91 30, and 35 °C). The strategy was considered based on the literature?
Response 6: Due to the storage of the fruit under dark and cold conditions, low temperatures can inhibit the growth of pathogens. Conversely, higher temperatures, such as those applied during heat treatments, may reduce pathogen populations. Therefore, the temperature range considered in this study was 1 to 35 °C. Please see this literature: Saito, S.; Michailides, T.J.; Xiao, C.L. Mucor rot—an emerging postharvest disease of mandarin fruit caused by Mucor piriformis and other Mucor spp. in California. Plant Dis. 2016, 100, 1054-1063
Comments 7: L127- Please clarify why the samples were transported to the lab within 1.5 h. Overall, some scenarios were considered for the trial without any reason. Please underscore why the utilized strategies were performed.
Response 7: We agree with this comment. As the production areas are located in mountainous regions, it requires approximately 1.5 hours to transport the samples to the packinghouses.
Comments 8: L153- here it needs to mention the juice extraction device.
Response 8: Thank you for pointing this out. The total soluble solids (TSS) of the fruit juice were extracted by hands and determined by a digital refractometer (Line 171)
Comments 9: L169- Table 1 presents the effect of different temperatures on the mycelial growth of Mucor lusitanicus for 7 days, but the description is not enough of why and for what reason temperature was between 5 and 0 °C. Thus, describe more.
Response 9: Thank you for pointing this out. We added some sentences. Therefore, the critical temperature for inhibiting the growth of M. lusitanicus was between 5 and 1 °C. These two temperatures (1 and 5°C) were chosen for precooling and storage, respectively because the farther can remove the heat and prevent pathogen growth fast and the later would be used for transportation and storage of litchi fruit in retail stores to avoid chilling injure. (Line 185-189)
Comments 10: L214- Table 5 shows the respiration rates and ethylene production of 'Yu-Her-Pao' litchis at different temperatures. This information can be useful for description of shelf life, hence, I suggest to add some information about shelf life. Also, the information about ethylene produced is strange, why the ethylene produced by the 15°C is zero!?
Response 10: The rate of ethylene production in litchi is less than 0.5 µL/kg·hr at 20 °C, as litchi is classified as a non-climacteric fruit (University of California, Davis, Postharvest Technology Center). ( https://postharvest.ucdavis.edu/produce-facts-sheets/lychee)
Comments 11: L264- Figure 3 illustrates the effects of hydrocooling with 40 mg L–1 HClO delay on the appearance of 'Yu- Her-Pao' litchis stored at 5 °C for 21 d followed by 26 °C for 1 d. The figure tries to show the appearance and dividing procedure, but, the figure is confusing. Please arrange each section in a better vision for more understanding. If the main aim of the figure is to divide the sample, so please remove it and explain the required information in the text.
Response 11: We agree with this comment. The figures have been rearranged for clarity and improved presentation. Please refer to the updated version.
Comments 12: L268-276 the content is not similar to the discussion, therefore, please remove it.
Response 12: We agreed with this comment and this part was removed.
Comments 13: L277-280 The statement represented the observed results and the lack of appropriate discussion is tangible. The conclusion needs deep revision. Authors should mention the output of the observed results and how the output addresses the current gap considering the subject of the paper. In addition, I suggest mentioning briefly the encountered challenges in the study and suggesting some ideas to cope with the challenges and limitations for further research.
Response 13: We agree with this comment. Therefore, we revised this part to establish a clearer link between observed results and challenges. “This disease was first observed in litchi fruit from farmers in Taiwan that had been stored at cold temperatures (from 5 to 10 °C) for a period in 2023. The decayed litchi fruit was collected and the pathogen was identified as M. lusitanicus 2023 to 2004. Although the Mucor spp. were isolated from 17 litchi packinghouses, pathogenicity tests was not be confirmed [18]. Pathogenicity tests showed M. lusitanicus was able to cause decay on the wound-inoculated litchi fruit (Table 4). After storage, the mucor rot has become the dominant postharvest disease in mandarin fruit in California where no official chemical treatments are currently recommended for cultivation [19]. Similarly, it is highly likely that commonly used fungicides for litchi production target major diseases in fields such as litchi sour rot, anthracnose, peronophythora downy blight, and fruit rot, which may render these chemicals ineffective against decay caused by Muco spp. Further research is needed to investigate the infection pathways of litchi fruits from farms to retail stores in Taiwan and to determine whether Mucor spp. becomes the dominant pathogen during long-term cold storage. The optimal growth temperature for M. lusitanicus was identified as 25 °C, and complete growth suppression occurred at 1 °C, with partial inhibition at 5 °C (Table 1). This finding could be applied to control the disease and evaluate the effect of delayed precooling on the litchi fruit. “(Line 287-303)
Round 2
Reviewer 1 Report
Comments and Suggestions for AuthorsThe author has made revisions; acceptance is recommended.
Reviewer 2 Report
Comments and Suggestions for AuthorsAll comments were considered and the current version is appropriate for publication.
