Seroepidemiology of Crimean-Congo Hemorrhagic Fever Virus (CCHFV) in Cattle across Three Livestock Pastoral Regions in Kenya

Round 1

Reviewer 1 Report

Authors present in this manuscript new data from Crimean-congo hemorrhagic fever virus prevalence in Kenya cattle. Results are interesting, but conclusions are not well uphold by sampling size.

Things to be consider:

1. Lines 114-115:  "Samples were collected during the low rain (2014 and 2019) and drought season (2016)." This means that two factors (year - season) are reduced to one
2. "A confidence interval of 95%, a margin of error of 5%, an infinite population, an assumed true overall prevalence of 50% to obtain maximum sample size, and the sensitivity and specificity of the analytical test were used in calculations of the sample size." For me this equations means 384 animals, but for authors it means 148 serum samples; why? And there we have the main problem in the manuscript: 148 samples, 46 positives, five factor to analyze, below a limit of calculation.
3. Abstract: multiple logistic mixed model... household as random effect. I understand household as Sites (table 1), with 18 categories, and only 148 cases, no posible mixed analysis to be done. Authors say they have joined categories but, which of them? This is not clear in text
4. Season, significant in table 4, should have been influenced by year, as dry was sampled in 2016 and Low rain in 2014 and 2019, and we assume (by this paper and others) that CCHFV is an increasing infection in Kenya from Uganda and other countries.
5. It is very difficult to be sure about which factor is really important with only 148 samples

On the other hand, I find simple results very important in the area and country.I think that results are important, and I recommend a reduction to a short communication maintaining prevalence and basic results.

Author Response

We would like to thank both the two reviewers for their comments, which have helped to make big clarity in the manuscript. We kindly ask the Editors to consider the manuscript for short communication (following the suggestion of Reviewer 1). We hope that this modified version will meet the expectations of the two reviewers and the editorial team.

Reviewer 1

Authors present in this manuscript new data from Crimean-Congo hemorrhagic fever virus prevalence in Kenya cattle. Results are interesting, but conclusions are not well uphold by sampling size.

AU: Dear anonymous Reviewer 1,

Thanks very much for your valuable comments and for reviewing our manuscript in such detail. We have taken great care to acknowledge all your comments and to consider your suggestions for improvements the manuscript. Please find below our responses to each of the points raised. We hope that this modified version will meet your expectations.

Regarding the limitations of the sample, we have tried to give more context to the study and circumscribe the outcomes to the sample. Therefore, the conclusion has been modified.

We had limited time and budget to perform more samples on small ruminants. The authors gave priority to the geographical coverage of contrasting areas with different husbandry practices. Nevertheless, we agree that representativeness is low. We have incorporated new sentences at the end of the discussion (lines 292-294 and lines 298-299).

Things to be consider:

1. Lines 114-115:  "Samples were collected during the low rain (2014 and 2019) and drought season (2016)." This means that two factors (year - season) are reduced to one

AU: The reviewer is totally right. We show a merged variable in the analytical part of the manuscript (Table 4) that now has been renamed (seasonality on sampling year) since it was misleading. We would like to explain that we first purposed to show the two single variables in the descriptive part of the manuscript so the reader could check the tendency by years (in Table 1 and Figure 3).

We have, in addition, inserted a rephrased sentence in Material and Methods (line: 164-166).

1. "A confidence interval of 95%, a margin of error of 5%, an infinite population, an assumed true overall prevalence of 50% to obtain maximum sample size, and the sensitivity and specificity of the analytical test were used in calculations of the sample size." For me this equations means 384 animals, but for authors it means 148 serum samples; why? And there we have the main problem in the manuscript: 148 samples, 46 positives, five factor to analyze, below a limit of calculation.

AU:  We understand the comment of the reviewer. We planned to sample 148 sheep and goats to reach the target but we have faced different challenges that forced us to end the field study without reaching the goal. Please notice that we have 4 factors in the logistic regression model.

1. Abstract: multiple logistic mixed model... household as random effect. I understand household as Sites (table 1), with 18 categories, and only 148 cases, no posible mixed analysis to be done. Authors say they have joined categories but, which of them? This is not clear in text

AU: We thank the reviewer for this comment. We have tried to improve the clarity of the Abstract. We listed some descriptors in the abstract but the logistic regression model was performed with four predictors: age, sex, season of time year and ecosystem.

1. Season, significant in table 4, should have been influenced by year, as dry was sampled in 2016 and Low rain in 2014 and 2019, and we assume (by this paper and others) that CCHFV is an increasing infection in Kenya from Uganda and other countries.

AU: We have renamed the variable since it is a merged variable combining dry season of different years. We understand the argument provided by the Reviewer and we apologize for the distraction.

1. It is very difficult to be sure about which factor is really important with only 148 samples

AU: We understand the comments of the reviewer and we have expanded more the study limitations at the end of the Discussion section.

On the other hand, I find simple results very important in the area and country. I think that results are important, and I recommend a reduction to a short communication maintaining prevalence and basic results.

AU: We have followed the suggestion of the reviewer. We have converted the manuscript into the requested guidance of the Journal for short communication. We thank the reviewer for this suggestion.

Author Response File: Author Response.pdf

Reviewer 2 Report

This work focused to Crime-Congo hemorrhagic fever virus in Kenya livestock. Crime-Congo hemorrhagic fever virus is high danger disease. This zoonosis has relationship with tick (tick-vertebrate-tick cycle). Authors analysed samles in ecosystems in 2014, 2016 and 2019. Each work is very important for better understanding epidemiology situation on the site.
Reviewer has follw comments and sugestions:
a) Time of collection of samples was 2014, 2016 and 2019. Why authors selected this intervals? 
b) Authors analysed cattle, sheep and goats samples. Can you make table results form each species year by year. I underestand that count of samples are not so high but It could be compared. Sheep (tab.3) have very low seropositive. Do you have any idea why?
c) Table 1 should be divaided for specific parameters.
d) Analysis of tick could be important too. Did you do any experiments?
e) Do you any information about gentic type of your virus. We could by expected UGU3010(DR Congo); PLOS https://doi.org/10.1371/journal.pntd.0009452
f) New information from published paper could by add to your Introduction or Discussion part. I finded some important
PLOS NEGLECTED TROPICAL DISEASES  Volume: ‏ 15   Issue: ‏ 4     Article Number: e0009299   Published: ‏ APR 2021
PATHOGENS  Volume: ‏ 9   Issue: ‏ 4     Article Number: 301   Published: ‏ APR 2020
VECTOR-BORNE AND ZOONOTIC DISEASES  Volume: ‏ 20   Issue: ‏ 5   Pages: ‏ 348-357   Published: ‏ MAY 1 2020
g) Did you collect any information about health of animals. All animals were without clinical symptoms.
h) Do you have any information about incidenci of zoonosis in human population?

This is my curious report.

Author Response

We would like to thank both the two reviewers for their comments, which have helped to make big clarity in the manuscript. We kindly ask the Editors to consider the manuscript for short communication (following the suggestion of Reviewer 1). We hope that this modified version will meet the expectations of the two reviewers and the editorial team.

Reviewer 2

This work focused to Crime-Congo hemorrhagic fever virus in Kenya livestock. Crime-Congo hemorrhagic fever virus is high danger disease. This zoonosis has relationship with tick (tick-vertebrate-tick cycle). Authors analysed samles in ecosystems in 2014, 2016 and 2019. Each work is very important for better understanding epidemiology situation on the site.
Reviewer has follw comments and sugestions:

AU: Dear Reviewer 2, Thanks very much for your useful comments – We have taken great care to acknowledge all your comments and to consider your suggestions for improvements to the manuscript. Please find below our responses to each of the points raised. We hope that this modified version will meet your expectations.

a) Time of collection of samples was 2014, 2016 and 2019. Why authors selected this intervals? 

AU: The sampling coincided with routine schedules for serosurveillance of multiple endemic diseases e.g FMD, RVF, CBPP, PPR, bTB. The rationale behind this corresponds with the study area experiencing scarce and unreliable rainfall patterns. So, the prolonged dry seasons trigger the movement of people and livestock to the River areas where water and pasture are abundant long after the rains have gone. This movement pattern facilitates the movement of potentially infected ticks across great distances, presenting an opportunity for the ex-change of diverse tick species and livestock populations.

We have rephrased this rationality in material and methods (line 108-118).

1. b) Authors analysed cattle, sheep and goats samples. Can you make table results form each species year by year. I underestand that count of samples are not so high but It could be compared. Sheep (tab.3) have very low seropositive. Do you have any idea why?

AU: This is an excellent suggestion however, we have not digitalized the sheep and goat data yet. So, we are not able to distribute small ruminants by time series along with the cattle population. Samples from goats and sheep were collected from the same periods as cattle unfortunately we are not able to provide this detailed information.

It might indicate that they are at a lesser risk of getting infected but under the discrete population of our study we are not in the position to make any speculation.

1. c) Table 1 should be divaided for specific parameters.
   AU: We have amended Table 1 improving the readability of it, thanks.

1. d) Analysis of tick could be important too. Did you do any experiments?
   AU: The reviewer is perfectly right notifying it. We are planning to perform some analysis with samples that have been recently collected. We will integrate this note of the reviewer into further studies in the conclusion of the manuscript.

10. e) Do you any information about gentic type of your virus. We could by expected UGU3010(DR Congo); PLOS https://doi.org/10.1371/journal.pntd.0009452
    AU: Many Thanks for providing this comment. The reference has been inserted in the manuscript.

Global representative of CCHFV genotypes must be studied and virus evolution, and phylogeography understood for preventative measures.

1. f) New information from published paper could by add to your Introduction or Discussion part. I finded some important
   PLOS NEGLECTED TROPICAL DISEASES  Volume: ‏ 15   Issue: ‏ 4     Article Number: e0009299   Published: ‏ APR 2021
   PATHOGENS  Volume: ‏ 9   Issue: ‏ 4     Article Number: 301   Published: ‏ APR 2020
   VECTOR-BORNE AND ZOONOTIC DISEASES  Volume: ‏ 20   Issue: ‏ 5   Pages: ‏ 348-357   Published: ‏ MAY 1 2020
   AU: Many Thanks for providing these references that help to keep the manuscript fully updated. Both articles have been inserted in the manuscript as suggested and they support previous statements. We are not able to develop it more due to word limitations (since we kindly request the editorial team to consider the manuscript as a short communication).

1. g) Did you collect any information about health of animals. All animals were without clinical symptoms.
   AU: There were no apparent clinical symptoms except swellings in some herds that were thought to be lumpy skin disease. We have inserted a clarification in the text (line: 123).
2. h) Do you have any information about incidenci of zoonosis in human population?

AU: Many Thanks for formulating this question. In Kenya, a human case of CCHFV was isolated from a farm with livestock heavily infested by ticks in the former western province. The study describing it is number [5] of the manuscript.

This is my curious report.

AU: Many thanks for taking the time and contribute to improving this manuscript.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Now we can accept the new version

Author Response

Many Thanks for helping to improve the manuscript. Very much appreciated. We have sent the manuscript for English proof editing. Best Regards

Reviewer 2 Report

Paper could be accept to publication.

Author Response

Many Thanks for helping to improve the manuscript. Very much appreciated. We have sent the manuscript for English proof editing. Best Regards