Astrovirus-Associated Polioencephalomyelitis in an Alpaca
, Michel C. Koch 3, Melanie M. Hierweger 3, Ronja V. Kauer 3, Céline L. Boujon 3, Monika Hilbe 4, Anna Oevermann 3, Patrik Zanolari 2, Torsten Seuberlich 3 and Corinne Gurtner 1
Round 1
Reviewer 1 Report
Dear authors,
well done and presented research work.
The manuscript is well written, the key information is presented in a suitable form.
The main message of this manuscript is the first detection of Astrovirus in alpaca with polioencephalomyelitis in Switzerland. The manuscript is relevant and interesting for publication.
The manuscript is well written, the originality of the topic is the first description of astrovirus in alpaca with polioencephalomyelitis. The subject was well compared with other published material.
The manuscript is well written. The conclusions are well presented, however further studies are needed to predict zoonotic potential of detected virus, infection dynamics and the shedding and transmission of virus within and between this species.
I have no remarks.
Author Response
We thank the reviewers for the detailed comments.
Reviewer 2 Report
Küchler et al describe the first case report of astrovirus encephalitis in an alpaca. This is a well-written case report that adds an additional mammalian species in which astroviruses can cause neurological disease. The authors are comprehensive and provide bona fide data demonstrating that an astrovirus was the causative pathogen. This manuscript would be of interest to the readers of Viruses.
Overall, the authors can enhance their manuscript by removing some extraneous information and creating a more concise story.
Specific comments:
The introduction is long and reads as if it was a review for astrovirus biology. The authors should consider deletion of lines 28-39 as these details add very little to the overall manuscript, especially since the focus is not on astrovirus biology.
Line 32: The authors state that the astrovirus genomes range in size from “6.8-7.9 kb.” However, some reference human astroviruses are smaller, including astrovirus MLB1 (NC_011400.1: 6171 nt in length) or VA1 (NC_013060.1: 6586 nt in length). If the authors keep this statement, they need to revise their discussion regarding genome size.
Lines 37-38: “Several studies suggest that astroviruses are the second most common pathogen after rotavirus to cause juvenile gastroenteritis.” This statement is based on a reference that is now almost 24 years old (see the citation used in reference 1). Most other studies have astroviruses as the 3rd to 5th most common cause of gastroenteritis (see more recent publications like: PMID 26202075). The authors should use a more up-to-date reference if they wish to keep this statement.
What was the timeframe from the timing of the necropsy to analysis by nucleic acid testing like PCR? If this timeframe was several years, it would also help to explain the fragmentation issue the authors discuss on lines 294-299.
Did the authors use the unmapped reads from their NGS analysis and map them to a viral database to determine if other viruses were present? If the astrovirus species was the only pathological virus detected, this would further strengthen the authors’ findings. If no additional pathological viruses were detected, the authors should report this in their discussion on lines 222-226.
Lines 230-235, and 268-270: On these lines, the authors state the virus localizes to neurons: “This led to unambiguous immunostaining within neuronal cells…” However, I disagree that the authors have clearly demonstrated that neurons are infected. While morphologically the positive cells for BoAstV-CH13/NeuroS1 appear to be neurons, the authors have not corroborated these results by staining the cells with a specific marker that these cells are indeed neurons. As discussed later, there is a publication demonstrating the capacity of a different astrovirus (astrovirus VA1) to infect astrocytes but not neurons (PMID 31289185). The authors should qualify their statements in these lines that the cells appear to be morphologically similar to neurons, or present co-staining results that unambiguously demonstrate the viral tropism for BoAstV-CH13/NeuroS1 is truly neurons.
Lines 253-265: These lines describe the pathology of rabies and Borna disease in alpacas. This information is unnecessary since testing was negative, so it can be deleted. I have no doubt this is a case of astrovirus encephalitis and not rabies or Borna disease based on the abundance of data presented by the authors.
Lines 266-272 and 279-290: Much of these sentences are rehashing of information already provided in the results section. This data can be further summarized and/or deleted.
Lines 306-307: The authors state: “Although many studies on these agents have been performed, virus isolation has still been impossible.” I disagree with this statement. Astrovirus VA1 causes encephalitis in humans, and a different group has been able to culture it in primary human astrocytes, but not neurons, resulting in production of inflammatory cytokines. Human astrovirus 4 also causes abortive infection in primary human astrocytes, see PMID 31289185. The authors should edit their statement as some preliminary studies into the neuropathogenesis of astrovirus infection has indeed been previously reported.
Line 321: Family name Astroviridae needs to follow ICTV naming conventions and be written in italics.
Figure 1: Will this figure be made in higher resolution? In the PDF that this image was reviewed, the image is blurry at all zoom levels.
Author Response
Dear Reviewer
Please find enclosed our revised manuscript, entitled „ Astrovirus-associated polioencephalomyelitis in an Alpaca”, which is intended for publication in Viruses, Section Animal Viruses, Special Issue “New Insights into Astrovirus Research”. We have carefully reviewed the comments and have revised the manuscript accordingly. Our responses are given in a point-by point manner below.
We would like to thank you and the reviewers for the positive and constructive comments regarding our manuscript. Specifically, we made the following changes:
Reviewer 2
- Comments and suggestions to Authors:
Küchler et al describe the first case report of astrovirus encephalitis in an alpaca. This is a well written case report that adds an additional mammalian species in which astroviruses can cause neurological disease. The authors are comprehensive and provide bona fide data demonstrating that an astrovirus was the causative pathogen. This manuscript would be of interest to the readers of Viruses.
Overall, the authors can enhance their manuscript by removing some extraneous information and creating a more concise story.
- Specific comments
a. The introduction is long and reads as if it was a review for astrovirus biology. The authors should consider deletion of lines 28-39 as these details add very little to the overall manuscript, especially since the focus is not on astrovirus biology.
Response: Revised accordingly (lines 29-32)
b. Line 32: The authors state that the astrovirus genomes range in size from “6.8-7.9 kb.” However, some reference human astroviruses are smaller, including astrovirus MLB1 (NC_011400.1: 6171 nt in length) or VA1 (NC_013060.1: 6586 nt in length). If the authors keep this statement, they need to revise their discussion regarding genome size.
Response: Revised accordingly.
c. Lines 37-38: “Several studies suggest that astroviruses are the second most common pathogen after rotavirus to cause juvenile gastroenteritis.” This statement is based on a reference that is now almost 24 years old (see the citation used in reference 1). Most other studies have astroviruses as the 3rd to 5th most common cause of gastroenteritis (see more recent publications like: PMID 26202075). The authors should use a more up-to-date reference if they wish to keep this statement.
Response: Revised accordingly with updated reference (line 32).
d. What was the timeframe from the timing of the necropsy to analysis by nucleic acid testing like PCR? If this timeframe was several years, it would also help to explain the fragmentation issue the authors discuss on lines 294-299.
Response: The analysis by nucleic acid testing was performed three months after necropsy. The authors added information about the timeframe to the text (lines 54 and 157).
e. Did the authors use the unmapped reads from their NGS analysis and map them to a viral database to determine if other viruses were present? If the astrovirus species was the only pathological virus detected, this would further strengthen the authors’ findings. If no additional pathological viruses were detected, the authors should report this in their discussion on lines 222-226.
Response: No additional pathological viruses were detected and the text was revised accordingly (line 220).
f. Lines 230-235, and 268-270: On these lines, the authors state the virus localizes to neurons: “This led to unambiguous immunostaining within neuronal cells…” However, I disagree that the authors have clearly demonstrated that neurons are infected. While morphologically the positive cells for BoAstV-CH13/NeuroS1 appear to be neurons, the authors have not corroborated these results by staining the cells with a specific marker that these cells are indeed neurons. As discussed later, there is a publication demonstrating the capacity of a different astrovirus (astrovirus VA1) to infect astrocytes but not neurons (PMID 31289185). The authors should qualify their statements in these lines that the cells appear to be morphologically similar to neurons, or present co-staining results that unambiguously demonstrate the viral tropism for BoAstV-CH13/NeuroS1 is truly neurons.
Response: Revised accordingly (lines 224-230).
g. Lines 253-265: These lines describe the pathology of rabies and Borna disease in alpacas. This information is unnecessary since testing was negative, so it can be deleted. I have no doubt this is a case of astrovirus encephalitis and not rabies or Borna disease based on the abundance of data presented by the authors.
Response: Revised accordingly
h. Lines 266-272 and 279-290: Much of these sentences are rehashing of information already provided in the results section. This data can be further summarized and/or deleted.
Response: Revised accordingly (lines 251-260 and 271-274).
i. Lines 306-307: The authors state: “Although many studies on these agents have been performed, virus isolation has still been impossible.” I disagree with this statement. Astrovirus VA1 causes encephalitis in humans, and a different group has been able to culture it in primary human astrocytes, but not neurons, resulting in production of inflammatory cytokines. Human astrovirus 4 also causes abortive infection in primary human astrocytes, see PMID 31289185. The authors should edit their statement as some preliminary studies into the neuropathogenesis of astrovirus infection has indeed been previously reported.
Response: The authors thank the Reviewer for his/her comment and edited their statement accordingly (lines 290-293)
j. Line 321: Family name Astroviridae needs to follow ICTV naming conventions and be written in italics.
Response: Revised accordingly (line 307)
k. Figure 1: Will this figure be made in higher resolution? In the PDF that this image was reviewed, the image is blurry at all zoom levels.
Response: The figure was updated in a higher resolution (line 186).
- Additional comment from the authors
The availability of video material in supplementary materials, which shows the animals gait abnormalities, was noted in brackets in line 62/63.
If you have any further questions regarding our manuscript, please do not hesitate to contact me at any time. We are looking forward to hearing your decision whether the revised manuscript can now be published in Viruses, Special Issue “New Insights into Astrovirus Research”.
Yours sincerely,
Leonore Küchler
