Proteomics and immunocharacterization of Asian mountain pit viper (Ovophis monticola) venom

Siravit Sitprija; Lawan Chanhome; Onrapak Reamtong; Tipparat Thiangtrongjit; Taksa Vasaruchapong; Orawan Khow; Jureeporn Noiphrom; Panithi Laoungbua; Arissara Tubtimyoy; Narongsak Chaiyabutr; Supeecha Kumkate

doi:10.1371/journal.pone.0260496

Peer Review History

Original SubmissionJune 7, 2021
13 Jul 2021 Decision Letter - Rafael Ximenes, Editor PONE-D-21-18686 Proteomics and immunocharacterization of Asian Mountain pit viper (Ovophis monticola) venoms PLOS ONE Dear Dr. Kumkate, Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. Please submit your revised manuscript by Aug 27 2021 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file. 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The PLOS ONE style templates can be found at https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf 2. In your Methods section, please provide additional location information, including geographic coordinates for the data set if available. 3. In your Methods section, please provide additional information regarding the permits you obtained for the work. Please ensure you have included the full name of the authority that approved the field site access and, if no permits were required, a brief statement explaining why. 4. We understand that you extracted venom from Ovophis monticola for this study. In your Methods section, please provide additional details regarding the source of the animal (Asian Mountain pit viper). 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If applicable, please specify in the figure caption text when a figure is similar but not identical to the original image and is therefore for illustrative purposes only. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes Reviewer #2: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: N/A Reviewer #2: N/A ****** 3. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes Reviewer #2: Yes ****** 4. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes Reviewer #2: Yes ****** 5. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: Referee comments: In this work, the authors demonstrate by venomics approaches, a comprehensive characterization of the venom proteome and immunocharacterization of Asian Mountain pit viper (Ovophis monticola) venoms, a medically important snake from Thailand, found at high altitude areas of several mountain ranges of East, South and Southeast Asia. Moreover the authors analyzed the immunoreactivity of monospecific (monovalent) and polyspecific (polyvalent) antivenoms against green pit viper Trimeresurus albolabris and hemotoxic venoms, respectively. The authors present a study with satisfactory quality in methodology and data generation. However the manuscript has an inventory of toxins and immunoreactive proteins without coherence with the scientific literature and poor consistency with some cited references. Furthermore, in many topics of the manuscript the authors present a lot of information without any reference. For example, the authors don't compare the results regarding 2D immunoblotting with any previously study. I suggest that this manuscript should be reword to improve the quality about the data presentation. I'm sending some notes - Abstract Line 33 ..... venoms was investigated by indirect ELISA..... Line 37.....were subjected to 2D immunoblotting These related methodologies would be more appropriate if they were introduced together. - Introduction . Line 46-50 - The geographical distribution data deserve references. The same for the sentences introducing biological data about pit viper from lines 51-58.... length of 110 cm. Lines 59: Incidence of pit vipers bites, including those from mountain pit vipers, have been documented in their known ranges???. The lack of references should be revised in all manuscript... -Materials and methods Line 92 : Snake venom, - Are there more information about the venom's sample used in this study? Number of specimens? Localities? size/age... Line 121: ...a non-linear immobilized pH gradient (IPG) strip (pH 3-10 Amersham Bioscience, USA).... - Why the authors choose the non-linear gradient instead of linear gradient, since the propose were excised immunoreactive spots for mass spectrometric analysis? Just for curious - Lack references in almost all methods. The references for 2D immunoblotting is required Results Line 208: Change Ovophis / Ovophis sp or genera Ovophis - Are there any published information about neutralize activity of monovalent and polyvalent antivenoms against O. monticola venom? This information should be provided or I suggest that an experiment to determine the lethal dose 50% should be done. Table 2. List of identified proteins ... immunologically reacted ... - If the MW of the spots content reacted/ non-reacted proteins could be provided, this information can improve the quality table. Discussion The Authors should revise the references from the first paragraph.... For example: the statement about hematotoxic potential of SVMP, SVSP and PLA2 is already a scientific consensus. Therefore, there are many sources specially from the seminal studies that corroborate with this question. However, the Authors present references from venom proteomic analysis of other snakes to support this question. On the other hand, the authors present coherency in the literature proposed as reference to address the mechanism of action of P-III metalloproteinase. I suggest that his paragraph must be reworded. Line 208: The presence of class P-II metalloproteinase, which possesses metalloproteinase and disintegrin-like domians was also observed.???? Line 303: The combination of SVMP and SVSP observed in O. monticola venom is responsible for prey attack, incapacitation, and digestion.??? Reviewer #2: It is an interesting work to be published in PLOS. This article describes describe proteomics and immuno characterization of snake Ovophis monticola venom. I have the following comments that the authors may like to consider: Line 95 - Provide the total number of individuals, age, sex, locality (adults or young people, males, females) - It is important to provide some functional analysis Discussion: Despite little published literature on Ovophis monticola: Please, include in the discussion this reference: Pandey, D. P., Chaudhary, B., & Ram Shrestha, B. (2021). Documentation of a proven Mountain Pitviper (Ovophis monticola) envenomation in Kathmandu, Nepal, with its distribution ranges: implications for prevention and control of pitviper bites in Asia. Journal of venom research, 11, 1–6. Mainly about clinical characteristic of poisoning by this specie and lack of specific treatment. Provide conclusion or future perspectives in the last paragraphy about immunocharacterization results/ envenomation treatment. Line 527 Exclude the number 75. ****** 6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review?** For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: No Reviewer #2: No [NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.] While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. 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Revision 1
20 Aug 2021 Author Response Responses to reviewers Reviewer #1: In this work, the authors demonstrate by venomics approaches, a comprehensive characterization of the venom proteome and immunocharacterization of Asian Mountain pit viper (Ovophis monticola) venoms, a medically important snake from Thailand, found at high altitude areas of several mountain ranges of East, South and Southeast Asia. Moreover the authors analyzed the immunoreactivity of monospecific (monovalent) and polyspecific (polyvalent) antivenoms against green pit viper Trimeresurus albolabris and hemotoxic venoms, respectively. The authors present a study with satisfactory quality in methodology and data generation. However the manuscript has an inventory of toxins and immunoreactive proteins without coherence with the scientific literature and poor consistency with some cited references. Furthermore, in many topics of the manuscript the authors present a lot of information without any reference. For example, the authors don't compare the results regarding 2D immunoblotting with any previously study. I suggest that this manuscript should be reword to improve the quality about the data presentation. Response: We thank the reviewer for suggestion. We rewrote introduction and discussion to improve data presentation. Relevant references were cited in every part to enhance literature coherence. Biological and geographical details regarding all snakes were also provided in materials and methods (Table 1; page 5, line 105). - Abstract Line 33 ..... venoms was investigated by indirect ELISA..... Line 37.....were subjected to 2D immunoblotting These related methodologies would be more appropriate if they were introduced together. Response: We edited an abstract according to reviewer’s suggestion. - Introduction . Line 46-50 - The geographical distribution data deserve references. The same for the sentences introducing biological data about pit viper from lines 51-58.... length of 110 cm. Lines 59: Incidence of pit vipers bites, including those from mountain pit vipers, have been documented in their known ranges???. The lack of references should be revised in all manuscript... Response : We rewrote introduction with relevant references cited in a revised version of manuscript; ref no.6 and 8). -Materials and methods Line 92 : Snake venom, - Are there more information about the venom's sample used in this study? Number of specimens? Localities? size/age... Response: Biological and geographical information regarding all snakes is provided as a table in materials and methods (Table 1; page 5 line 105). Line 121: ...a non-linear immobilized pH gradient (IPG) strip (pH 3-10 Amersham Bioscience, USA).... - Why the authors choose the non-linear gradient instead of linear gradient, since the propose were excised immunoreactive spots for mass spectrometric analysis? Just for curious Response: According to our previous experiments, we found that the non-linear immobilized pH gradient (IPG) could provide the well separation of protein in venoms. Therefore, we chose the NL strip for this research. - Lack references in almost all methods. The references for 2D immunoblotting is required. Response: In a revised materials and methods of manuscripts, we provided relevant references for all methods used. For 2DE immunoblotting, a reference is doi: 10.3390/toxins6051526 as reference page 7, line 137 (ref no.14) Results Line 208: Change Ovophis / Ovophis sp or genera Ovophis - Are there any published information about neutralize activity of monovalent and polyvalent antivenoms against O. monticola venom? This information should be provided or I suggest that an experiment to determine the lethal dose 50% should be done. Table 2. List of identified proteins ... immunologically reacted ... - If the MW of the spots content reacted/ non-reacted proteins could be provided, this information can improve the quality table. Response: We added a column of MW of proteins in a revised Table 2. In this particular study, we focused mainly on providing the inventory of all protein constituents of Asian mountain pit viper O. monticola venom. Those with antigenic properties reacted with monovalent and hematotoxic polyvalent antivenom were given. However, we did not perform neutralizing experiments. Discussion The Authors should revise the references from the first paragraph.... For example: the statement about hematotoxic potential of SVMP, SVSP and PLA2 is already a scientific consensus. Therefore, there are many sources specially from the seminal studies that corroborate with this question. However, the Authors present references from venom proteomic analysis of other snakes to support this question. On the other hand, the authors present coherency in the literature proposed as reference to address the mechanism of action of P-III metalloproteinase. I suggest that his paragraph must be reworded. Line 208: The presence of class P-II metalloproteinase, which possesses metalloproteinase and disintegrin-like domians was also observed.???? Line 303: The combination of SVMP and SVSP observed in O. monticola venom is responsible for prey attack, incapacitation, and digestion.??? Response: We thank the reviewer for comments on discussion. We rewrote discussion by updating and improving literature coherence. Reviewer #2: It is an interesting work to be published in PLOS. This article describes describe proteomics and immuno characterization of snake Ovophis monticola venom. I have the following comments that the authors may like to consider: Line 95 - Provide the total number of individuals, age, sex, locality (adults or young people, males, females) - It is important to provide some functional analysis Response: Biological and geographical information regarding all snakes is provided as a table in materials and methods (Table 1; page 5 line 105). In this particular study, we focused mainly on providing the inventory of all protein constituents of Asian mountain pit viper O. monticola venom. Those with antigenic properties reacted with monovalent and hematotoxic polyvalent antivenom were given. However, we did not perform the experiments to analyse the functions. Discussion: Despite little published literature on Ovophis monticola: Please, include in the discussion this reference: Pandey, D. P., Chaudhary, B., & Ram Shrestha, B. (2021). Documentation of a proven Mountain Pitviper (Ovophis monticola) envenomation in Kathmandu, Nepal, with its distribution ranges: implications for prevention and control of pitviper bites in Asia. Journal of venom research, 11, 1–6. Mainly about clinical characteristic of poisoning by this specie and lack of specific treatment. Provide conclusion or future perspectives in the last paragraphy about immunocharacterization results/ envenomation treatment. Line 527 Exclude the number 75. Response: We cited the above literature as a reference in a revised version of manuscript (ref no.5). Attachments Attachment Submitted filename: Response to reviewers.docx https://doi.org/10.1371/journal.pone.0260496.r002
6 Oct 2021 Decision Letter - Rafael Ximenes, Editor PONE-D-21-18686R1Proteomics and immunocharacterization of Asian Mountain pit viper (Ovophis monticola) venomsPLOS ONE Dear Dr. Kumkate, Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. Please submit your revised manuscript by Nov 20 2021 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file. Please include the following items when submitting your revised manuscript: A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'. A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'. An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'. If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter. If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols. We look forward to receiving your revised manuscript. Kind regards, Rafael Ximenes Academic Editor PLOS ONE Journal Requirements: Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice. Additional Editor Comments (if provided): Based on the new round of peer-round some minor concerns have arisen. I recommend a minor revision before acceptance of the manuscript. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation. Reviewer #1: All comments have been addressed Reviewer #3: All comments have been addressed ******** 2. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes Reviewer #3: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #3: Yes ****** 4. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes Reviewer #3: Yes ****** 5. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: No Reviewer #3: Yes ****** 6. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: The authors rewrote the manuscript and provide relevant references and endeavored to comply with all referee requests.I just emphasize that only Ovophis don't belong to a taxonomic system. This task dont were revide for authors. However. The authors present a study with high quality in methodology, data generation and data analysis, producing significant contribution for toxinology field. Thus, this work is suitable for publication on PLOS ONE. Reviewer #3: The article highlights the spectrometry-based proteomics and the immunoreactivity of the venom of the Ovophis monticola viper. It is a work based on an excellent premise and the methodologies are very well defined. I recommend the work for publication. However, some minor revisions are needed. 1. In the session "Two-dimensional polyacrylamide gel electrophoresis (2DE)" (Material and Methods), the authors state that a 100 µg sample of venom was used. However, since the protein concentrations were measured using the Lowry method, please review whether these 100 µg are venom or protein. 2. In the session “Proteomic analysis of O. monticola venom” (Results), the first paragraph deals with a methodological aspect. 3. The same observation applies to the following topics. Instead of repeating the concepts of the methodology, the objective could be presented if evaluating that result, since this information is not present in the text. 4. In “Immunoreactivity of protein antigens in O. monticola venom to monovalent and polyvalent antivenoms by indirect ELISA”, the information “Since there is no homospecific antivenom to Ovophis venoms currently available, all pit viper envenoming victims usually receive either monovalent antivenom (raised against T. albolabris venom) or hematotoxic polyvalent antivenom (produced against venoms of C. rhodostoma, D. siamensis and T. albolabris) to alleviate symptoms” is applicable to the discussion, requiring references. 5. Figures and tables should be self-explanatory. In the case of table 2, a legend stating the meaning of MW, PI, emPI must be provided. 6. Discussion: What is the toxinological consequence related to the fact that the venom has more acidic than basic protein spots? How does this interfere with symptoms related to accident? Is this positive or negative regarding the immunoreactivity of the venom, especially with the polyvalent serum? Present references. 7. Why are some references in numerical format (ex (22) (23)) and others are in nominal format (ex Damm et al, 2021)? ****** 7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: Yes:** Carlos Correa-Netto Reviewer #3: No [NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.] While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0260496.r003
Revision 2
9 Nov 2021 Author Response Additional Editor Comments (if provided): Based on the new round of peer-round some minor concerns have arisen. I recommend a minor revision before acceptance of the manuscript. Reviewers' comments: Reviewer #1: The authors rewrote the manuscript and provide relevant references and endeavored to comply with all referee requests. I just emphasize that only Ovophis don't belong to a taxonomic system. This task dont were revide for authors. However. The authors present a study with high quality in methodology, data generation and data analysis, producing significant contribution for toxinology field. Thus, this work is suitable for publication on PLOS ONE. Response: We corrected Ovophis to Ovophis spp. as suggested by the reviewer (e.g. lines 68, 83, 91, 288, 305, 313, 396) Reviewer #3: The article highlights the spectrometry-based proteomics and the immunoreactivity of the venom of the Ovophis monticola viper. It is a work based on an excellent premise and the methodologies are very well defined. I recommend the work for publication. However, some minor revisions are needed. 1. In the session "Two-dimensional polyacrylamide gel electrophoresis (2DE)" (Material and Methods), the authors state that a 100 µg sample of venom was used. However, since the protein concentrations were measured using the Lowry method, please review whether these 100 µg are venom or protein. Response: : It was 100 �g of protein. We corrected the sentence in line 135. 2. In the session “Proteomic analysis of O. monticola venom” (Results), the first paragraph deals with a methodological aspect. Response: We rewrote this paragraph according to the reviewer’s suggestion. 3. The same observation applies to the following topics. Instead of repeating the concepts of the methodology, the objective could be presented if evaluating that result, since this information is not present in the text. Response: We revised result session and eliminated repetition particularly on methodology. 4. In “Immunoreactivity of protein antigens in O. monticola venom to monovalent and polyvalent antivenoms by indirect ELISA”, the information “Since there is no homospecific antivenom to Ovophis venoms currently available, all pit viper envenoming victims usually receive either monovalent antivenom (raised against T. albolabris venom) or hematotoxic polyvalent antivenom (produced against venoms of C. rhodostoma, D. siamensis and T. albolabris) to alleviate symptoms” is applicable to the discussion, requiring references. Response: We clarified this point by providing a reference (line 232, ref no. 16 ) 5. Figures and tables should be self-explanatory. In the case of table 2, a legend stating the meaning of MW, PI, emPI must be provided. Response: We added the explanation in the table 2 legend (lines 238-241, 244-246) and the definition of abbreviation is provided as in a legend as suggested. 6. Discussion: What is the toxinological consequence related to the fact that the venom has more acidic than basic protein spots? How does this interfere with symptoms related to accident? Is this positive or negative regarding the immunoreactivity of the venom, especially with the polyvalent serum? Present references. Response: In this revised version of manuscript, we discussed the above points raised by the reviewer with relevant references, lines 323-327, 391-395. In addition, to make table 4 more readable, we added a column of MW of non-immunoreactive peptides. 7. Why are some references in numerical format (ex (22) (23)) and others are in nominal format (ex Damm et al, 2021)? Response: We corrected all references appeared in text in numerical format according to Plos guideline. Attachments Attachment Submitted filename: Response to reviewers.docx https://doi.org/10.1371/journal.pone.0260496.r004
11 Nov 2021 Decision Letter - Rafael Ximenes, Editor Proteomics and immunocharacterization of Asian Mountain pit viper (Ovophis monticola) venom PONE-D-21-18686R2 Dear Dr. Kumkate, We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements. Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication. An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org. If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org. Kind regards, Rafael Ximenes Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: https://doi.org/10.1371/journal.pone.0260496.r005
Formally Accepted
19 Nov 2021 Acceptance Letter - Rafael Ximenes, Editor PONE-D-21-18686R2 Proteomics and immunocharacterization of Asian Mountain pit viper (Ovophis monticola) venom Dear Dr. Kumkate: I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org. If we can help with anything else, please email us at plosone@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Dr. Rafael Ximenes Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0260496.r006

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