PONE-D-22-22212Prevalence of different trypanosome species in naturally infected cattle of three sleeping sickness foci of the south of ChadPLOS ONE

Dear Dr. Simo,

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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

Reviewer #3: Yes

Reviewer #4: Yes

Reviewer #5: Yes

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2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: I Don't Know

Reviewer #3: Yes

Reviewer #4: No

Reviewer #5: No

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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: No

Reviewer #3: Yes

Reviewer #4: No

Reviewer #5: Yes

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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

Reviewer #3: Yes

Reviewer #4: Yes

Reviewer #5: Yes

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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: This is a valuable addition to the literature and research effort on animal trypanosomiasis in endemic countries. The MS is well written and presented for the most part. I found the discussion overlong for my taste at 4+ pages, but on the other hand this gave space for thorough discussion and all material was relevant. I have the following suggestions for improvement, many just tweaks to improve readability:

Title: could be more informative "Prevalence of pathogenic trypanosome species in..."

Abstract line 22: cattle is a plural word, use animal instead. Body condition (not plural); ditto line 32. Line 28 no cattle were found...Line 36 expand abbreviation AAT - not previously used?

Intro: Line 69 constitute (livestock is a plural word).

Methods: Line 105, 110, 116 replace peasant with "subsistence". Line 109 replace precipitations with "annual rainfall". Line 123/4 rewrite to clarify "In each village, only cattle that had been exposed to tsetse fly bites by spending at least 3 months in the study zone were included and randomly selected for sampling. From each selected animal,...". Line 128 Delete "Moreover" = unnecessary. Line 141 replace associated to with "supplemented with". Line 143 Clarify whether this procedure was done with all 1466 samples or just the CTC positives. Line 225 P value was <0.05.

Results: Line 230 rewrite "were aged 2 years or over". Line 231/2 Delete "about" = unnecessary; body condition (not plural). Line 307 Moissala spelling. Line 344, 349 "one triple infection", "the last double infection" (not plural).

Table 1, footnote line 250 trypanosomosis.

Table 2 - age groups "<2 and >2 years" leaves out cattle of 2 years exactly - which category includes them?

Discussion: Line 389 delete "too" - not necessary. Line 390 "no cattle were found...". Line 404 "this test is unable to..." - there are certainly ways to distinguish T.evansi from T. brucei, and here you have used a specific PCR test for T.b.gambiense. Line 413 T.equiperdum?? not very likely considering this is an equine tryp transmitted via coitus? or do you envisage some strange transmission scenarios in Chad? Line 420 discrepancies also could be due to susceptibility of different livestock to T.brucei infection? Similarly for T.congolense forest and savannah line 444-449. Line 428 indicate (not plural). Line 432 italics submorsitans. Line 469-470 female and male cattle - no plural needed. Line 483 surely poor body condition can be attributed to trypanosome infection - it's a sign of the disease. Line 501 "for a bovine..." Line 543 smallholder is one word.

General: Might be useful to include a map of Chad with 3 foci locations. Throughout - subgenus does not need a hyphen.

Reviewer #2: The ms entitled « Prevalence of different trypanosome species in naturally infected cattle of three

sleeping sickness foci of the south of Chad » aims to provide information on the prevalence of trypanosomes in randomly selected cattle within sleeping sickness foci in Chad. Although the idea is very good and interesting, and the ms is clearly presented, there are severe lacks that need to be addressed before this can be published. In particular it is very difficult to understand why the authors do not mention and discuss the importance of a vector control campaign in the Mandoul that has been published and the likely impact on their results. Another limit, linked to this, is that it is difficult to draw any conclusion on the epidemiology without knowing the sedentary or transhumant status of the cattle, which have unfortunately not been taken into account in their study design.

Mahamat et al (2017) published a paper entitled « Adding tsetse control to medical activities contributes to decreasing transmission of sleeping sickness in the Mandoul focus (Chad) » where they show a likely disappearance of tsetse flies from 2016 onwards as a consequence of a vector control campaign using tiny targets (https://journals.plos.org/plosntds/article/figure?id=10.1371/journal.pntd.0005792.g003).

Even more, Rock et al., 2022 (Rock et al. Infectious Diseases of Poverty (2022) 11:11

https://doi.org/10.1186/s40249-022-00934-8) , using a modelling approach, concluded that transmission of T. b. gambiense had probably been interrupted since 2015, so well before the sampling described for this present ms (2019). Again, I hardly understand why the authors do not even cite these references, and do not discuss the likely impact of this on their results, at least for the Mandoul g-HAT focus.

As an example, in the Mandoul, given the likely disappearance of tsetse and of Tbg, the expectation is to find zero tsetse transmitted trypanosomes in sedentary animals. The occurrence of trypanosomes may be possible in animals that are transhumant. Unfortunately the authors did not design their study in a way allowing testing and discussing this. Alternatively, it then becomes quite tricky to explain the author’s results describing so many tsetse transmitted trypanosomes in an area where there is no longer any possibility of cyclical transmission, based on these papers.

As an example, how to reconcile, line 306 « The highest prevalence (8.3%) of trypanosomes of the sub-genus Trypanozoon was obtained in cattle from the Mandoul HAT focus » ? Would there be any possibility of false positives in their results ? what does mean a positive PCR, does this mean an active trypanosome infection ? not so sure, by the way…there may be other hypotheses to explain this result (transhumant cattle/Trypanozoon trypanosomes that are not T. b. gambiense/etc.), but ignoring these references is misleading for this study, at least for the situation in the Mandoul.

Reviewer #3: The article which discusses the prevalence of trypanosomosis in the HAT foci of Southern Chad primarily allayed the fears of cattle serving as reservoir of infection to humans. The manuscript was written in simple, coherent and easy to read language. The methods used were quite clearly explained and helps to understand the investigation carried out. i commend the authors for this.

introduction

Line 45: delete..."parasitic diseases" and replace with ..."parasites"

Line 137: replace entire subheading with "Parasitological tests"

Line 223: How did you compare the prevalence of different trypanosome species with Chi-square? Could you be talking about the associations of the variables with positive trypanosome results from the different study areas?

Line 230: replace ..."had" with "were"

Line 249-253: Describe "CT"

Line 259: The percentage age prevalence of positive RDTs was repeated. Delete the repetition.

Line 320: replace "having" with "being"

Line 391-397: What is the author's view on sensitivity and specificity of the RDT in excluding Trypanosoma brucei gambiense?

Line 474-479: I would agree more with the fact that the older cattle are more exposed than the younger cattle a reason for the high prevalence. it might be interesting to note that being immunologically immature connote a higher susceptibility in the younger than the older cattle.

Line 480-488: Many factors may predispose cattle to poor body conditions namely; other concurrent parasitic infections particularly helminth infections in addition to their nutritional status. i had also expected the authors to correlate the body condition of the cattle to the occurrence of anaemia. This is a common feature of trypanosomosis and should have been investigated in line with the body condition scores of these cattle. This could have helped in the discussion of the element of body condition in this study

Reviewer #4: This is a good study. As stated by the authors, identification of trypanosome species and determining their prevalence in animals from different settings are essential for the understanding of AAT epidemiology and the development of control strategies. Much as it explored AAT mainly, it had the potential contribute information on the role of animal reservoirs in the epidemiology of HAT. Current knowledge of T. b. gambiense infections in animals is limited and fragmented.

Main comments/questions

Study design not well described. Was sampling just opportunistic? There is no formula to calculate a representative sample size and therefore we cannot infer if this sample size is reasonable vis -a-vis the cattle population in the region.

The approach to analyse the data is correct but inadequate. Bivariate analyses should be followed by multiple logistic regressions to assess the magnitude of association of the factors with prevalence of infection!

The results for RDT designed for the screening of T.b.gambiense seems interesting. For T. b. gambiense, despite early data generated on its infectivity and transmissibility in animals, the epidemiological significance of any animal reservoir remain at best not well understood. The discussion on this according to me has not been done with sufficient rigour.

The authors opine in line 386-393 that the presence of +ve RDTs but lack of Tbg positive DNAs in cattle could be due to past exposure! Can they cite their sources of this information? How frequent has this phenomenon been observed in cattle? What is the role of cattle as reservoirs of T.b.gambiense overall?

The write up would benefit from edits to make sentences more precise and less wordy. As an example, look at line 372-375. There are three sentences all essentially talking about the same thing! This is rather common throughout the manuscript!

Minor comments

Line 46: Replace “induce” with “cause”

Line 60-61: Should read “the pathogenic animal trypanosomes species…”

Line 64: Use “constrain” instead of “restrain”….

Line 88-90: revise sentence; doesn't read really well!

Line 101: Maro focus be described first, followed by Mandoul and then Moissala

Line 173: check spelling of “congolense”; it’s written as “congolence”. And it occurs in other instances in the manuscript.

Line 426-433: I think G. fuscipes fuscipes in particular has been shown to have a higher transmission index for T. vivax than other species like pallidipes. Moreover T. vivax is known to have the shortest life cycle among all the trypanosome species. So the least predominance of T. vivax seems surprising. Could there be other explanations for this finding in the study area other than vector competence?

Line 450-457: The explanation advanced here can be true for the difference in overall prevalence but may not suffice for the difference in species of trypanosomes observed in the different foci!

Line 474-475: How does “immature immunity of younger cattle” explain lower disease prevalence in younger cattle than older ones? Shouldn’t it be the contrary???

Reviewer #5: Joel et al. submitted an article on "prevalence of different trypanosome species in naturally infected cattle of three sleeping sickness foci of the south of Chad". The authors did a great job which will be an interesting read among the scientific community. However, there are few points that need to be addressed which have been highlighted in the pdf. The major limitation in the work is the failure to speciate the "Trypanozoon" group of trypanosomes identified. This is a major setback in understanding the full epidemiology and best control strategies. Otherwise, the authors have done justice to most part of the work.

Important points

Line 23: Is there any correlation between the HAT foci (location) and AAT infections observed? Or it's there any impact of this location on the AAT species identified?

Line 293: Did the authors differentiate the Trypanozoon group at all? The TBR primers will identify the T. brucei brucei species. The T. evansi primer is also available. The high Trypanozoon group could be dominated by a trypanosome species, which could give us the idea of the predominant tsetse species responsible for transmission in a bid to control the disease.

Line 306: You'd observe that the distribution is even. Knowing the species composition would add to the knowledge if it can be done.

Line 378: The HAT foci considered in Chad, is it a tsetse-free belt area or otherwise? In the literature, you didn't report the distribution of tsetse flies caught in that area before.

Line 440-441: Comparing with equine result from another study doesn't add anything to this explanation. That comparison should be deleted

The 95% confidence interval should be included in all percentages.

The discussion is too long and contains so many unnecessary information.

There's no proper reporting of statistical inferential analysis in the manuscript (%95 CI, P-value)

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Reviewer #1: No

Reviewer #2: No

Reviewer #3: No

Reviewer #4: Yes: Dr Robert Opiro

Department of Biology

Gulu University, Uganda

Reviewer #5: Yes: Dr Paul Odeniran

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Submitted filename: PONE-D-22-22212_reviewer.pdf

PONE-D-22-22212R1Prevalence of pathogenic trypanosome species in naturally infected cattle of three sleeping sickness foci of the south of ChadPLOS ONE

Dear Dr. Simo,

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.

One of the reviewers recommends that you make minor revisions to the manuscript as outlined in their attached coments. Please attend to all their comments and return the revised manuscript as adivised in this letter. 

Please submit your revised manuscript by Dec 16 2022 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 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,

Martin Chtolongo Simuunza, PhD

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.

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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 #2: (No Response)

Reviewer #4: All comments have been addressed

Reviewer #5: 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: (No Response)

Reviewer #2: Partly

Reviewer #4: Yes

Reviewer #5: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: (No Response)

Reviewer #2: Yes

Reviewer #4: Yes

Reviewer #5: 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: (No Response)

Reviewer #2: Yes

Reviewer #4: Yes

Reviewer #5: 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 Response)

Reviewer #2: Yes

Reviewer #4: Yes

Reviewer #5: 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 have addressed all points. The altered spelling of Moissala needs to be consistently applied throughout, not only in Results but also Abstract and Methods.

Reviewer #2: The authors have correctly answered most of the comments made. A couple of points still need clarification/better formulation. In the discusssion, the authors should spend more time discussing the limits of their results and the tools they use, whereas the discussion is very long on sometimes very speculative results. E.g. are they sure that a PCR positive means an active infection ? I am not. Can’t their be any false positive or false negative results ? the authors use RDTs for gambiense in cattle, but what is the performance on this test designed for humans in animals ? do you have any idea ? any reference ? I think these are exemples of points that need to be discussed before interpreting what they may mean epidemiologically…

In their answer, the authors state

« However, from 2016 to 2020, HAT cases have been reported in all Chadian HAT foci including the Mandoul HAT focus where the number of HAT cases decreased while a slight increase was observed in the Maro HAT focus (Franco et al., 2022). This continuous detection of HAT cases in the Mandoul HAT focus indicates that a slight cyclical transmission of trypanosomes may still occurs despite vector control operations. ».

May I clarify here : if Rock et al., 2022 are correct in saying that g-HAT transmission has been interrupted in the Mandoul since 2015, this does not preclude from still finding HAT cases, that would have been infected before the interruption of transmission (g-HAT is a chronic, long lasting disease), or that would come from other locations where there is g-HAT…but it does not indicate that cyclical transmission occurs, because this would be in total contradiction with the beginning of the sentence. So please just remove this sentence otherwise you also to need to discuss this point into details, the rest is ok.

Hence this paragraph should now be (l.446 of the revised version) :

« The highest prevalence of trypanosomes of the subgenus Trypanozoon in cattle from the Mandoul HAT focus is surprising because the “tiny targets” deployed in this focus for vector control during three consecutive years (2014 to 2016) before our sampling were expected to stop trypanosomes transmission (Mahamat et al., 2017). This hypothesis was strengthened by results of mechanistic transmission model suggesting that HAT transmission would have been interrupted in 2015 due to intensified interventions in the Mandoul HAT focus (Rock et al., 2022). However, from 2016 to 2020, HAT cases have been reported in all Chadian HAT foci including the Mandoul HAT focus where the number of HAT cases decreased while a slight increase was observed in the Maro HAT focus (Franco et al., 2022). This continuous detection of HAT cases in the Mandoul HAT focus indicates that a slight cyclical transmission of trypanosomes may still occurs despite vector control operations. Nonetheless, the probability of mechanical transmission of trypanosomes of the subgenus Trypanozoon such as T. evansi cannot be excluded. In addition to that, the movement of herders with some infected cattle in the search of pastures during transhumance phenomenon is another factor that could explain the highest prevalence of trypanosomes of the subgenus Trypanozoon in cattle from the Mandoul HAT focus. »

In additon, the fact that there has been a vector control campaign in the Mandoul must be clarified very early in the ms for the reader. This may be stated in M&M section, line 115 for instance.

Please avoid this kind of arbitrary sentence, especially in your conclusion « No infection of T. b. gambiense was recorded in cattle despite the positivity of RDT used to screen HAT. Cattle cannot be considered as potential reservoir for human-infective trypanosomes in Chad. » please avoid this ! say something like « based on our results, we do not think , bla bla… »

You also spend a very long time discussion your results, but very little time discussing the limits of your results and tools used.

Reviewer #4: The authors have have tried and addressed all my concerns in the initial review! I therefore have no further comments to make except to offer my congratulations to the authors on a job well done

Reviewer #5: The authors have addressed the queries, and provided scientific limitation to their study. The statistical analysis have been performed thoroughly after being raised as a concern in the initial version. This new version is a better one and can be published.

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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.

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Reviewer #1: No

Reviewer #2: No

Reviewer #4: Yes: Robert Opiro

Department of Biology

Gulu University

Reviewer #5: Yes: Dr Paul Odeniran

**********

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Submitted filename: PLOS_Reviewer 5.docx

PONE-D-22-22212R2Prevalence of pathogenic trypanosome species in naturally infected cattle of three sleeping sickness foci of the south of ChadPLOS ONE

Dear Dr. Simo,

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 Jan 22 2023 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 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,

Martin Chtolongo Simuunza, PhD

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:

One of the reviewers is still of the view that you have not adequately attended to their comments. I am therefore requesting you to either incoperate their concerns in the manuscript or give a reason why that should not be the case. Then you can resubmit the revised manuscript as advised in this letter.

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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 #2: (No Response)

**********

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: (No Response)

Reviewer #2: (No Response)

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Reviewer #1: (No Response)

Reviewer #2: (No Response)

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Reviewer #1: (No Response)

Reviewer #2: (No Response)

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Reviewer #1: (No Response)

Reviewer #2: (No Response)

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Reviewer #1: (No Response)

Reviewer #2: everything ok now thank you, only one of my earlier comment has not been taken into account, not because of the authors, but because of a typo in earlier comments. Please this paragraph starting l 448 should appear like that :

« The highest prevalence of trypanosomes of the subgenus Trypanozoon in cattle from

the Mandoul HAT focus is surprising because the “tiny targets” deployed in this focus

for vector control during three consecutive years (2014 to 2016) before our sampling

were expected to stop trypanosomes transmission (Mahamat et al., 2017). This

hypothesis was strengthened by results of mechanistic transmission model suggesting

that HAT transmission would have been interrupted in 2015 due to intensified

interventions in the Mandoul HAT focus (Rock et al., 2022). However, from 2016 to

2020, HAT cases have been reported in all Chadian HAT foci including the Mandoul

HAT focus where the number of HAT cases decreased while a slight increase was

observed in the Maro HAT focus (Franco et al., 2022). This continuous detection of

HAT cases in the Mandoul HAT focus indicates that a probability of mechanical transmission of trypanosomes of the subgenus Trypanozoon such as T. evansi cannot be excluded. In addition to that, the movement of herders

with some infected cattle in the search of pastures during transhumance phenomenon

is another factor that could explain the highest prevalence of trypanosomes of the

subgenus Trypanozoon in cattle from the Mandoul HAT focus.

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Reviewer #1: No

Reviewer #2: No

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Prevalence of pathogenic trypanosome species in naturally infected cattle of three sleeping sickness foci of the south of Chad

PONE-D-22-22212R3

Dear Dr. Simo,

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