Frequency, Resistance Patterns, and Serotypes of Salmonella Identified in Samples from Pigs of Colombia Collected from 2022 to 2023

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The scope of the study is interesting and provides information to take health measures in the departments/country. The effort to collect the samples is high, more than 5000 samples, the authors even say that with some departments it was easier than with others. But there is an important bias, 199 positive samples were detected, but only 105 samples were subjected to antiobigram and 61 to serotyping, the rest of the samples because they were discarded? It is not indicated in the study, nor how this selection was made. Another bias is the selection of antibiotics, even they themselves indicate it in the final part of the article, this should be corrected and select the same set of antibiotics and by taking conclusions of multi-resistance.

Line 62 - The European Union's One Health 2023 annual report has already been published, the data could be updated.

Line 89 - The authors used two diagnostic methods, one method from serum samples and the other from tissue or feces, but did not indicate whether the samples were from the same animals or from different animals. Were these samples related to the others? Did they come from the same animals?

Line -172 Be careful with the results, for example, although they indicate the sample size, they indicate that 100% of the samples were positive for erythromycin, but only one sample was tested. Regardless of the client, wouldn't it be better to use the same set of antibiotics for all samples in the study? 

Line 190 - only 61 out of 199 positive samples have been serotyped, this only corresponds to 30%, the rest of the samples because they were not serotyped? Those 61 were randomly selected? Because this is another bias.

Line 296 - be careful with this conclusion, some antibiotics have only been tested on a single sample.

Bibliography - Please review the bibliography, enter full citations and Salmonella always in italics.

Author Response

 

1. The scope of the study is interesting and provides information to take health measures in the departments/country. The effort to collect the samples is high, more than 5000 samples, the authors even say that with some departments it was easier than with others. But there is an important bias, 199 positive samples were detected, but only 105 samples were subjected to antiobigram and 61 to serotyping, the rest of the samples because they were discarded? It is not indicated in the study, nor how this selection was made. Another bias is the selection of antibiotics, even they themselves indicate it in the final part of the article, this should be corrected and select the same set of antibiotics and by taking conclusions of multi-resistance.

An additional clarification was incorporated into the study methodology to provide the reader with more comprehensive information:

"This cross-sectional study utilized clinical samples referred for isolation and identification, primarily feces and tissues from pigs suspected of having salmonellosis. For enzyme-linked immunosorbent assay (ELISA) tests, serum samples were submitted both for swine production monitoring and suspected disease cases, although serology is predominantly used for surveillance. The exact number of sick animals is unknown, and it is unclear how many samples were collected specifically for monitoring purposes.

The selection of antibiotics for testing, the decision to perform antibiograms, and the choice to conduct serotyping were entirely client-driven. The data analyzed in this study were obtained from the database of the Diagnostic Unit of the Faculty of Agricultural Sciences at the University of Antioquia. As this unit provides diagnostic services to the public, the analyses performed were based on client requests and needs rather than a standardized research protocol. Consequently, variability in testing reflects real-world diagnostic practices rather than methodological bias. Furthermore, client-driven anti-biotic selection, as well as the proportions of serotyping and antibiogram requests, were included in the analysis, as they provide valuable insights into veterinary diagnostic decision-making and antimicrobial resistance monitoring trends".

 

1. Line 62 - The European Union's One Health 2023 annual report has already been published, the data could be updated.

We appreciate this suggestion and accept the change in the manuscript. The data will be updated accordingly to reflect the European Union's One Health 2023 annual report.

2. Line 89 - The authors used two diagnostic methods, one method from serum samples and the other from tissue or feces, but did not indicate whether the samples were from the same animals or from different animals. Were these samples related to the others? Did they come from the same animals?

 

We appreciate this observation and have included the following explanation in the manuscript:

"The total number of samples with complete records was 5,820 serum samples tested with ELISA. Additionally, 848 different samples (feces and tissues, most of them from the intestine) were submitted for culture to isolate and identify Salmonella. The serum samples were used for the ELISA test, while the tissue and fecal samples were used for microbiological culture. However, these samples did not always come from the same animals. Although all samples were from pigs, the individuals selected for ELISA testing were not necessarily the same as those selected for microbiological culture."

 

3. Line -172 Be careful with the results, for example, although they indicate the sample size, they indicate that 100% of the samples were positive for erythromycin, but only one sample was tested. Regardless of the client, wouldn't it be better to use the same set of antibiotics for all samples in the study? 

 

The following explanation has been added to the manuscript:

"The selection of antibiotics varied across samples, as it was determined according to the client's specific preferences, making it impossible to use the same set of antibiotics for all samples in the study. Among the antibiograms performed, resistance was observed to ampicillin (3/3), tiamulin (1/1), penicillin (2/2), tylosin (2/2), and erythromycin (1/1), followed by florfenicol (44/54), doxycycline (4/5), spectinomycin (18/25), amoxicillin (32/46), chloramphenicol (2/3), tetracycline (2/3), and enrofloxacin (34/64), with lower resistance observed for fosfomycin (2/38) and ceftiofur (5/35), as shown in Table 4."

 

 

4. Line 190 - only 61 out of 199 positive samples have been serotyped, this only corresponds to 30%, the rest of the samples because they were not serotyped? Those 61 were randomly selected? Because this is another bias.

 

We appreciate your comment. The following explanation has been added to the manuscript:

"Out of the 199 isolated Salmonella samples, only 61 were serotyped due to the client's specific preferences and testing priorities. The selection of samples for serotyping was based on the client's request rather than a standardized approach across all isolates."

 

5. Line 296 - be careful with this conclusion, some antibiotics have only been tested on a single sample.

 

The following explanation has been added to the manuscript:

"However, caution must be exercised when interpreting these results, as some antibiotics were tested on a very limited number of samples due to client preferences. Since the data were sourced from a public diagnostic service institution, the selection of antibiotics tested was not standardized but rather determined by the specific needs of each client."

 

 

6. Bibliography - Please review the bibliography, enter full citations and Salmonella always in italics.

We appreciate your comment. The requested changes have been made: the bibliography has been reviewed, full citations have been entered, and Salmonella is now consistently formatted in italics.

Author Response File: Author Response.docx

Reviewer 2 Report

Comments and Suggestions for Authors

Dear Authors,

After reviewing the manuscript entitled "Frequency, resistance patterns, and serotypes of Salmonella identified in samples from pigs of Colombia collected from 2022 to 2023"

I see that the article is very important from a 

I have the several comments regarding this work:

01- I suggest to change the title from "Frequency, resistance patterns, and serotypes of Salmonella identified in samples from pigs of Colombia collected from 2022 to 2023" to "Prevalence, resistance patterns, and serotypes of Salmonella identified in samples from pigs of Colombia collected from 2022 to 2023".

02- You are invited to add a section in the Introduction related to the virulence factors that are present in Salmonella species, to mention the presence of pathogenicity island, presence of genes coding for metallophores (which are considered as virulence factors), etc. I suggest to use the following papers as reference for this point: 

The Different Types of Metallophores Produced by Salmonella enterica: A Review

Salmonella pathogenicity islands: big virulence in small packages

03- One major point regarding the present work, appears in Table 3, when you mentioned that 17 samples are of unknown type!! How can you explain this point, if you are working on a project how can you define a sample of unknown type?!!

04- Another major point related to the present study is the results of antibiogram, in this part the problem is related to the number of times the antibiotic was used, some antibiotics were used just for one time, another for 2 or 3 times!! Do you think that the results of these tests are meaningful !!!! I think that is a big issue of the work. Let's take the example of Tiamulin which was used for 1 time, you mentioned that the resistance to Tiamulin is 100% based on one sample!! I do not see that is meaningful, And I suggest to analyze just antibiotics used several times  (15 times or more for example).

05- In the Results, the follow "Out of the 199 isolated Salmonella samples, only 61 were serotyped". Why just these 61 samples were serotyped?? Why did you select these samples?

06- In the Results, You mentioned the follow "Of the 105 strain samples subjected to antibiogram, 45 underwent serotyping". Same question as above, Why just 105 were subjected to antibiogram and why 45 underwent serotyping, the high number of missing data in the present paper is a big Issue.

07- Noting the high pathogenicity of Salmonella, taking into consideration its high resistance to antibiotics in the present study, I suggest to go deeply in the discussion and talk about the importance of finding new families of antibiotics with new strategies to fight AMR in different pathogenic bacteria. I suggest to talk about new strategies such as:

• The Trojan Horse Strategy
• Fighting bacteria by targeting metallophores synthesis, export and import systems
• Other strategies.

You can use the following manuscripts as references for this point:

Towards new antibiotics classes targeting bacterial metallophores

Advances in ‘Trojan horse’ Strategies in Antibiotic Delivery Systems

Best Regards,

Author Response

After reviewing the manuscript entitled "Frequency, resistance patterns, and serotypes of Salmonella identified in samples from pigs of Colombia collected from 2022 to 2023"

I see that the article is very important from a 

I have the several comments regarding this work:

1. I suggest to change the title from "Frequency, resistance patterns, and serotypes of Salmonella identified in samples from pigs of Colombia collected from 2022 to 2023" to "Prevalence, resistance patterns, and serotypes of Salmonella identified in samples from pigs of Colombia collected from 2022 to 2023".

 

At some point, we considered using that same title. However, given that a prevalence study requires specific methodological characteristics, we decided that "Frequency" was a more appropriate term for our study. Prevalence studies typically require a well-defined sampling framework that ensures representativeness, which was not the case in our study, as the data were obtained from a public diagnostic service and were dependent on client-requested analyses.

 

2. One major point regarding the present work, appears in Table 3, when you mentioned that 17 samples are of unknown type!! How can you explain this point, if you are working on a project how can you define a sample of unknown type?!!

 

The data used in our study were sourced from the database of the Diagnostic Unit of the Faculty of Agricultural Sciences at the University of Antioquia, covering the years 2022 and 2023. This unit provides diagnostic services to the public, meaning that the information reflects real-world veterinary diagnostic practices. In many cases, clients do not provide complete or accurate sample identification, which can result in some samples being classified as "unknown type." This situation is inherent to routine diagnostic services, where sample submission depends entirely on the information provided by the clients rather than a controlled research setting.

 

An additional clarification was incorporated into the study methodology to provide the reader with more comprehensive information.

 

3. Another major point related to the present study is the results of antibiogram, in this part the problem is related to the number of times the antibiotic was used, some antibiotics were used just for one time, another for 2 or 3 times!! Do you think that the results of these tests are meaningful !!!! I think that is a big issue of the work. Let’s take the example of Tiamulin which was used for 1 time, you mentioned that the resistance to Tiamulin is 100% based on one sample!! I do not see that is meaningful, And I suggest to analyze just antibiotics used several times  (15 times or more for example).

The data used in our study were sourced from the database of the Diagnostic Unit of the Faculty of Agricultural Sciences at the University of Antioquia, covering the years 2022 and 2023. This unit provides public diagnostic services, where clients decide which antibiotics to test and whether or not to request an antibiogram or serotyping. As a result, the selection of antibiotics was not standardized but rather reflected the specific needs and preferences of each client. This study represents real-world veterinary diagnostic practices rather than a controlled research setting. The inclusion of all tested antibiotics, regardless of the number of times they were analyzed, is considered important to illustrate which antibiotics are most and least requested by clients. This information is valuable for understanding antibiotic use trends in veterinary diagnostics and guiding future research on antimicrobial resistance.

An additional clarification was incorporated into the study methodology to provide the reader with more comprehensive information.

 

4. In the Results, the follow "Out of the 199 isolated Salmonella samples, only 61 were serotyped". Why just these 61 samples were serotyped?? Why did you select these samples?

The selection of samples for serotyping was entirely based on the client's request. Since the data were obtained from the database of the Diagnostic Unit of the Faculty of Agricultural Sciences at the University of Antioquia, which provides public diagnostic services, the decision to perform serotyping was made by the clients according to their specific needs and priorities. To clarify this point, additional explanations have been incorporated into the manuscript.

5. In the Results, You mentioned the follow "Of the 105 strain samples subjected to antibiogram, 45 underwent serotyping". Same question as above, Why just 105 were subjected to antibiogram and why 45 underwent serotyping, the high number of missing data in the present paper is a big Issue.

 

The selection of samples for serotyping was entirely based on the client's request. Since the data were obtained from the database of the Diagnostic Unit of the Faculty of Agricultural Sciences at the University of Antioquia, which provides public diagnostic services, the decision to perform serotyping was made by the clients according to their specific needs and priorities.

 

6. Noting the high pathogenicity of Salmonella, taking into consideration its high resistance to antibiotics in the present study, I suggest to go deeply in the discussion and talk about the importance of finding new families of antibiotics with new strategies to fight AMR in different pathogenic bacteria. I suggest to talk about new strategies such as:

• The Trojan Horse Strategy
• Fighting bacteria by targeting metallophores synthesis, export and import systems
• Other strategies.

You can use the following manuscripts as references for this point:

Towards new antibiotics classes targeting bacterial metallophores

Advances in ‘Trojan horse’ Strategies in Antibiotic Delivery Systems

We appreciate the suggestion and find the proposed references very interesting. For this reason, we have incorporated the following section into the manuscript:

"Given the high pathogenicity of Salmonella and the significant antimicrobial resistance observed in this study, it is crucial to explore new strategies for combating antimicrobial resistance in pathogenic bacteria. The implementation of innovative therapeutic strategies [27,28] could contribute to mitigating the spread of resistant Salmonella strains and improving treatment options in both veterinary and human medicine."

 

Author Response File: Author Response.docx

Round 2

Reviewer 2 Report

Comments and Suggestions for Authors

Dear Authors,

The revised version of your article has been reviewed,

Thanks to the modifications you made, the article is suitable for publication in this new version.

Best Regards,