Abstract
The gut mycobiome exhibits major influence on the gastrointestinal health and disease but received less attention due to low abundance. This study characterizes the fungal community and compares the microbial diversity between indoor and outdoor cats. Genomic DNA was extracted and sequenced by targeting the Internal Transcribed Spacer 1 (ITS1) region using Flongle flow cell on MinION™ sequencing platform. Results show the phylum Ascomycota and genus Peniophorella were numerous in indoor cats, whereas the Basidiomycota and Pichia were abundant in outdoor cats. Peniophorella formed the core mycobiome in both feline populations. Furthermore, alpha (p value = 0.0207) and beta diversities (p value = 0.009) results showed significant differences between the two groups. Overall, indoor cats have greater amounts of Peniophorella, whereas outdoor cats have higher Trichosporon and unclassified Sordariaceae. The study also suggests that keeping a cat indoors or left as a stray will affect their respective gut mycobiome.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Data availability
The sequencing data were deposited in the National Center for Biotechnology Information (NCBI) database and registered as BioProject PRJNA813732, Sequence Read Archive (SRA) was deposited as SRR18272551-SRR18272542, and BioSample with accession numbers SAMN26521149–SAMN26521158. Additional data regarding results can be found in supplementary file.
References
Ahmad HF, Mejia JLC, Krych L, Khakimov B, Kot W, Bechshøft RL, Reitelseder S, Højfeldt GW, Engelsen SB, Holm L, Faust K, Nielsen DS (2020) Gut mycobiome dysbiosis is linked to hypertriglyceridemia among home dwelling elderly danes. BioRxiv 2020.04.16.044693. https://doi.org/10.1101/2020.04.16.044693
Ahmad Kendong SM, Raja Ali RA, Nawawi KNM, Ahmad HF, Mokhtar NM (2021) Gut dysbiosis and intestinal barrier dysfunction: potential explanation for early-onset colorectal cancer. Front Cell Infect Microbiol 11(December):1–18. https://doi.org/10.3389/fcimb.2021.744606
Ariyawansa HA, Tanaka K, Thambugala KM, Phookamsak R, Tian Q, Camporesi E, Hongsanan S, Monkai J, Wanasinghe DN, Mapook A, Chukeatirote E, Kang JC, Xu JC, McKenzie EHC, Jones EBG, Hyde KD (2014) A molecular phylogenetic reappraisal of the Didymosphaeriaceae (= Montagnulaceae). Fungal Divers 68(1):69–104. https://doi.org/10.1007/s13225-014-0305-6
Banos S, Lentendu G, Kopf A, Wubet T, Glöckner FO, Reich M (2018) A comprehensive fungi-specific 18S rRNA gene sequence primer toolkit suited for diverse research issues and sequencing platforms. BMC Microbiol 18(1):1–15. https://doi.org/10.1186/s12866-018-1331-4
Bengtsson-Palme J, Ryberg M, Hartmann M, Branco S, Wang Z, Godhe A, De Wit P, Sánchez-García M, Ebersberger I, de Sousa F, Amend A, Jumpponen A, Unterseher M, Kristiansson E, Abarenkov K, Bertrand YJK, Sanli K, Eriksson KM, Vik U, Nilsson RH et al (2013) Improved software detection and extraction of ITS1 and ITS2 from ribosomal ITS sequences of fungi and other eukaryotes for analysis of environmental sequencing data. Methods Ecol Evol 4(10):914–919. https://doi.org/10.1111/2041-210X.12073
Benjamin D (1996) Mushrooms: poisons and panaceas: a handbook for naturalists, mycologists, and physicians. Choice Rev Online 33(06):33-3303. https://doi.org/10.5860/choice.33-3303
Bernhardt A, von Bomhard W, Antweiler E, Tintelnot K (2015) Molecular identification of fungal pathogens in nodular skin lesions of cats. Med Mycol 53(2):132–144. https://doi.org/10.1093/mmy/myu082
Biegańska MJ, Rzewuska M, Dąbrowska I, Malewska-Biel B, Ostrzeszewicz M, Dworecka-Kaszak B (2018) Mixed infection of respiratory tract in a dog caused by Rhodotorula mucilaginosa and Trichosporon jirovecii: a case report. Mycopathologia 183(3):637–644. https://doi.org/10.1007/s11046-017-0227-4
Bolyen E, Rideout JR, Dillon MR, Bokulich NA, Abnet CC, Al-Ghalith GA, Alexander H, Alm EJ, Arumugam M, Asnicar F, Bai Y, Bisanz JE, Bittinger K, Brejnrod A, Brislawn CJ, Brown CT, Callahan BJ, Caraballo-Rodríguez AM, Chase J, Caporaso JG et al (2019) Reproducible, interactive, scalable and extensible microbiome data science using QIIME 2. Nat Biotechnol 37(8):852–857. https://doi.org/10.1038/s41587-019-0209-9
Cai L, Jeewon R, Hyde KD (2006) Phylogenetic investigations of Sordariaceae based on multiple gene sequences and morphology. Mycol Res 110(2):137–150. https://doi.org/10.1016/j.mycres.2005.09.014
Celi P, Cowieson AJ, Fru-Nji F, Steinert RE, Kluenter AM, Verlhac V (2017) Gastrointestinal functionality in animal nutrition and health: new opportunities for sustainable animal production. Anim Feed Sci Technol 234(September):88–100. https://doi.org/10.1016/j.anifeedsci.2017.09.012
Chong J, Liu P, Zhou G, Xia J (2020) Using MicrobiomeAnalyst for comprehensive statistical, functional, and meta-analysis of microbiome data. Nat Protoc 15(3):799–821. https://doi.org/10.1038/s41596-019-0264-1
Colombo AL, Padovan ACB, Chaves GM (2011) Current knowledge of trichosporon spp. and trichosporonosis. Clin Microbiol Rev 24(4):682–700. https://doi.org/10.1128/CMR.00003-11
Dhariwal A, Chong J, Habib S, King IL, Agellon LB, Xia J (2017) MicrobiomeAnalyst: a web-based tool for comprehensive statistical, visual and meta-analysis of microbiome data. Nucleic Acids Res 45(W1):W180–W188. https://doi.org/10.1093/nar/gkx295
Edward MJ (2000) Dictionary of toxicology second edition. J Toxicol Cutan Ocul Toxicol 19(2–3):169–170. https://doi.org/10.3109/15569520009051513
Egidi E, Delgado-Baquerizo M, Plett JM, Wang J, Eldridge DJ, Bardgett RD, Maestre FT, Singh BK (2019) A few Ascomycota taxa dominate soil fungal communities worldwide. Nat Commun. https://doi.org/10.1038/s41467-019-10373-z
Garcia-Mazcorro JF, Minamoto Y (2013) Gastrointestinal microorganisms in cats and dogs: a brief review. Arch Med Vet 45(2):111–124. https://doi.org/10.4067/S0301-732X2013000200002
Gouba N, Hien YE, Guissou ML, Fonkou MDM, Traoré Y, Tarnagda Z (2019) Digestive tract mycobiota and microbiota and the effects on the immune system. Hum Microbiome J 12(May):100056. https://doi.org/10.1016/j.humic.2019.100056
Guan QX, Zhao TJ, Zhao CL (2020) Morphological characters and phylogenetic analyses reveal two new species of Peniophorella from southern China. Mycol Prog 19(4):397–404. https://doi.org/10.1007/s11557-020-01568-6
Guinane CM, Cotter PD (2013) Role of the gut microbiota in health and chronic gastrointestinal disease: Understanding a hidden metabolic organ. Ther Adv Gastroenterol 6(4):295–308. https://doi.org/10.1177/1756283X13482996
Haleem Khan AA, Mohan Karuppayil S (2012) Fungal pollution of indoor environments and its management. Saudi J Biol Sci 19(4):405–426. https://doi.org/10.1016/j.sjbs.2012.06.002
Hallenberg N, Nilsson RH, Antonelli A, Wu SH, Maekawa N, Nordén B (2007) The Peniophorella praetermissa species complex (Basidiomycota). Mycol Res 111(12):1366–1376. https://doi.org/10.1016/j.mycres.2007.10.001
Huffnagle GB, Noverr MC (2013) The emerging world of the fungal microbiome. Trends Microbiol 21(7):334–341. https://doi.org/10.1016/j.tim.2013.04.002
Ikmal Hisham FB (2015) A comparison of gastrointestinal microbial communities between indoor cats and outdoor cats Farhana Ikmal Hisham. Honors Theses., p 2629. https://scholarworks.wmich.edu/honors_theses/2629
Jensen T, Bechshoeft RL, Giacalone D, Otto MH, Castro-Mejía J, Bin Ahmad HF, Reitelseder S, Jespersen AP (2016) Whey protein stories—an experiment in writing a multidisciplinary biography. Appetite 107:285–294. https://doi.org/10.1016/j.appet.2016.08.010
Lyu Y, Su C, Verbrugghe A, Van de Wiele T, Martos Martinez-Caja A, Hesta M (2020) Past, present, and future of gastrointestinal microbiota research in cats. Front Microbiol. https://doi.org/10.3389/fmicb.2020.01661
Mith O, Benhamdi A, Castillo T, Bergé M, Macdiarmid CW, Steffen J, Eide DJ, Perrier V, Subileau M, Gosti F, Berthomieu P, Marquès L (2015) The antifungal plant defensin AhPDF1.1b is a beneficial factor involved in adaptive response to zinc overload when it is expressed in yeast cells. Microbiol Open 4(3):409–422. https://doi.org/10.1002/mbo3.248
Moosavi A, Ghazvini RD, Ahmadikia K, Hashemi SJ, Geramishoar M, Mohebali M, Yekaninejad MS, Bakhshi H, Khodabakhsh M (2019) The frequency of fungi isolated from the skin and hair of asymptomatic cats in rural area of Meshkin-shahr-Iran. J Mycol Med 29(1):14–18. https://doi.org/10.1016/j.mycmed.2019.01.004
Mukherjee PK, Chandra J, Retuerto M, Sikaroodi M, Brown RE, Jurevic R, Salata RA, Lederman MM, Gillevet PM, Ghannoum MA (2014) Oral mycobiome analysis of HIV-infected patients: identification of pichia as an antagonist of opportunistic fungi. PLoS Pathog. https://doi.org/10.1371/journal.ppat.1003996
Oberacker P, Stepper P, Bond DM, Höhn S, Focken J, Meyer V, Schelle L, Sugrue VJ, Jeunen GJ, Moser T, Hore SR, von Meyenn F, Hipp K, Hore TA, Jurkowski TP (2019) Bio-on-magnetic-beads (BOMB): open platform for high-throughput nucleic acid extraction and manipulation. PLoS Biol 17(1):1–16. https://doi.org/10.1371/journal.pbio.3000107
Reagan KL, Dear JD, Kass PH, Sykes JE (2019) Risk factors for Candida urinary tract infections in dogs and cats. J Vet Intern Med 33(2):648–653. https://doi.org/10.1111/jvim.15444
Richard ML, Sokol H (2019) The gut mycobiota: insights into analysis, environmental interactions and role in gastrointestinal diseases. Nat Rev Gastroenterol Hepatol 16(6):331–345. https://doi.org/10.1038/s41575-019-0121-2
Rissi DR, Kirby KD, Sanchez S (2016) Systemic Trichosporon loubieri infection in a cat. J Vet Diagn Invest 28(3):350–353. https://doi.org/10.1177/1040638716640313
Rodríguez-Pérez H, Ciuffreda L, Flores C (2020) NanoCLUST: a species-level analysis of 16S rRNA nanopore sequencing data. Bioinformatics 37(11):1600–1601. https://doi.org/10.1093/bioinformatics/btaa900
Sharman MJ, Stayt J, McGill SE, Mansfield CS (2010) Clinical resolution of a nasal granuloma caused by Trichosporon loubieri. J Feline Med Surg 12(4):345–350. https://doi.org/10.1016/j.jfms.2009.11.004
Suchodolski JS (2011) Intestinal microbiota of dogs and cats: a bigger world than we thought. Vet Clin N Am Small Anim Pract 41(2):261–272. https://doi.org/10.1016/j.cvsm.2010.12.006
Tay DD, Siew SW, Razali MN, Ahmad HF (2021) Metadata analysis for gut microbiota between indoor and street cats of Malaysia. Curr Sci Technol 1(1):56–65. https://doi.org/10.15282/cst.v1i1.6443
Tedersoo L, Anslan S, Bahram M, Põlme S, Riit T, Liiv I, Kõljalg U, Kisand V, Nilsson RH, Hildebrand F, Bork P, Abarenkov K (2015) Shotgun metagenomes and multiple primer pair-barcode combinations of amplicons reveal biases in metabarcoding analyses of fungi. MycoKeys 10(May):1–43. https://doi.org/10.3897/mycokeys.10.4852
Tong Y, Tang J (2017) Candida albicans infection and intestinal immunity. Microbiol Res 198:27–35. https://doi.org/10.1016/j.micres.2017.02.002
Vaser R, Sović I, Nagarajan N, Šikić M (2017) Fast and accurate de novo genome assembly from long uncorrected reads. Genome Res 27(5):737–746. https://doi.org/10.1101/gr.214270.116
Vernocchi P, Chierico FD, Putignani L (2020) Gut microbiota metabolism and interaction with food components. Int J Mol Sci 21(10):1–18. https://doi.org/10.3390/ijms21103688
Wanasinghe DN, Jones EBG, Camporesi E, Dissanayake AJ, Kamolhan S, Mortimer PE, Xu J, Abd-Elsalam KA, Hyde KD (2016) Taxonomy and phylogeny of Laburnicola gen. nov. and Paramassariosphaeria gen. nov. (Didymosphaeriaceae, Massarineae, Pleosporales). Fungal Biol 120(11):1354–1373. https://doi.org/10.1016/j.funbio.2016.06.006
Wu X, Xia Y, He F, Zhu C, Ren W (2021) Intestinal mycobiota in health and diseases: from a disrupted equilibrium to clinical opportunities. Microbiome 9(1):1–18. https://doi.org/10.1186/s40168-021-01024-x
Yeoman CJ, White BA (2014) Gastrointestinal tract microbiota and probiotics in production animals. Annu Rev Anim Biosci 2:469–486. https://doi.org/10.1146/annurev-animal-022513-114149
Zhang N, Cai G, Price DC, Crouch JA, Gladieux P, Hillman B, Khang CH, Lebrun MH, Lee YH, Luo J, Qiu H, Veltri D, Wisecaver JH, Zhu J, Bhattacharya D (2018) Genome wide analysis of the transition to pathogenic lifestyles in Magnaporthales fungi. Sci Rep 8(1):1–13. https://doi.org/10.1038/s41598-018-24301-6
Zhang M, Wang L, Wu G, Wang X, Lv H, Chen J, Liu Y, Pang H, Tan Z (2021) Effects of Lactobacillus plantarum on the fermentation profile and microbiological composition of wheat fermented silage under the freezing and thawing low temperatures. Front Microbiol 12(June):1–14. https://doi.org/10.3389/fmicb.2021.671287
Zhao J, Mou Y, Shan T, Li Y, Zhou L, Wang M, Wang J (2010) Antimicrobial metabolites from the endophytic fungus pichia guilliermondii Isolated from Paris polyphylla var. yunnanensis. Molecules 15(11):7961–7970. https://doi.org/10.3390/molecules15117961
Zheng D, Liwinski T, Elinav E (2020) Interaction between microbiota and immunity in health and disease. Cell Res 30(6):492–506. https://doi.org/10.1038/s41422-020-0332-7
Acknowledgements
The authors would like to thank the cat owners who provided cat stool samples and data for this research.
Funding
This work was financially supported the Ministry of Higher Education Malaysia, Universiti Malaysia Pahang and Supercat International Sdn. Bhd. for supporting this study through research funding. HFA was awarded for FRGS/1/2019/WAB13/UMP/03/1 and MNR was granted with UMP-MTUN Industry Matching Grant RDU192802/UIC190812, respectively.
Author information
Authors and Affiliations
Contributions
HFA, MNR, and SSK conceived the study and participated in its design. DDT and SSW conducted the experiment and analyzed the data. DDT and SSW drafted the manuscript. All authors read, revised, and approved the manuscript.
Corresponding author
Ethics declarations
Conflict of interest
The authors declare that they have no conflict of interest in the publication.
Ethical approval
This is an observational study. The Institutional Animal Care and Use Committee (IACUC) of Universiti Malaysia Pahang confirmed that no ethical approval is required because only freely passed stool samples were collected from healthy cats, no specific ethical approval was required. All samples used in this analysis were collected with verbal owner consent, who were aware that these samples were taken for research purpose only.
Additional information
Communicated by Erko Stackebrandt.
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Information
Below is the link to the electronic supplementary material.
203_2022_2929_MOESM2_ESM.tif
Supplementary file2 Fig. S1. Gut mycobiome composition for cats were conducted with Indoor Cats (IC) samples (n=5) and Outdoor Cats (OC) samples (n=5) through ITS1 gene amplicon sequencing. a). Taxa summary - Bar plot illustrating the relative abundant fungal features present within the two groups which combines all appropriate samples at phylum level where Ascomycota (abundance=0.6551) is more prevalent in Indoor Cats (IC) and Basidiomycota (abundance=0.7400) in Outdoor Cats (OC); b). Taxa summary - Bar plot illustrating the top 10 relative abundant fungal features present within the two groups which combines all appropriate samples at genus level where Peniophorella (abundance=0.3274) is most abundant in Indoor Cats (IC) samples while Trichosporon (abundance= 0.3484) and Pichia (abundance= 0.3641) are most abundant in Outdoor Cats (OC) samples (TIF 159 KB)
203_2022_2929_MOESM3_ESM.tif
Supplementary file3 Fig. S2. Gut mycobiome composition for cats were conducted with Indoor Cats (IC) samples (n=5) and Outdoor Cats (OC) samples (n=5) through ITS1 gene amplicon sequencing. Results were extracted from MicrobiomeAnalyst after post-processing in QIIME2: a). Taxa summary - Bar plot illustrating the relative abundant fungal features present within the two groups for each sample at class level; b). Taxa summary - Bar plot illustrating the relative abundant fungal features present within the two groups which combines all appropriate samples at class level where Agaricomycetes (abundance=0.7352) is more prevalent in Indoor Cats (IC) and Saccharomycetes (abundance=0.5733) in Outdoor Cats (OC); c). Taxa summary - Bar plot illustrating the top relative abundant fungal features present within the two groups for each sample at order level; d). Taxa summary - Bar plot illustrating the top relative abundant fungal features present within the two groups which combines all appropriate samples at order level where Agaricales (abundance=0.4077) and Corticiales (abundance=0.3274) are most abundant in Indoor Cats (IC) samples while Saccharomycetales (abundance= 0.5733) and Trichosporonales (abundance=0.3331) are most abundant in Outdoor Cats (OC) samples (TIF 535 KB)
203_2022_2929_MOESM4_ESM.tif
Supplementary file4 Fig. S3. Gut mycobiome composition for cats were conducted with Indoor Cats (IC) samples (n=5) and Outdoor Cats (OC) samples (n=5) through ITS1 gene amplicon sequencing. Results were extracted from MicrobiomeAnalyst after post-processing in QIIME2: a). Taxa summary - Bar plot illustrating the relative abundant fungal features present within the two groups for each sample at family level; b). Taxa summary - Bar plot illustrating the relative abundant fungal features present within the two groups which combines all appropriate samples at family level where Corticiaceae (abundance=0.3274) is more prevalent in Indoor Cats (IC) while Trichosporonaceae (abundance=0.3331) and Pichiaceae (abundance=0.3481) in Outdoor Cats (OC); c). Taxa summary - Bar plot illustrating the top 10 relative abundant fungal features present within the two groups for each sample at species level; d). Taxa summary - Bar plot illustrating the top 10 relative abundant fungal features present within the two groups which combines all appropriate samples at species level where Peniophorella odontiiformis (abundance=0.3274) is the most abundant in Indoor Cats (IC) samples while unclassified Trichosporon (abundance=0.3484) and Pichia kudriavzevii (abundance=0.3641) are most abundant in Outdoor Cats (OC) samples (TIF 436 KB)
203_2022_2929_MOESM5_ESM.tif
Supplementary file5 Fig. S4. Analysis was performed on both Indoor Cats (IC) (n=5) and Outdoor Cats (OC) (n=5) sample data obtained from ITS1 gene amplicon sequencing. Results were extracted from MicrobiomeAnalyst after post-processing in QIIME2: a). Plot of alpha diversity results using index observed feature at sample level which shows higher diversity in Outdoor Cats (OC) samples compared to Indoor Cats (IC). b). Alpha rarefaction plot showcasing that full diversity was observed within all samples (TIF 120 KB)
Rights and permissions
About this article
Cite this article
Tay, D.D., Siew, S.W., Shamzir Kamal, S. et al. ITS1 amplicon sequencing of feline gut mycobiome of Malaysian local breeds using Nanopore Flongle. Arch Microbiol 204, 314 (2022). https://doi.org/10.1007/s00203-022-02929-3
Received:
Accepted:
Published:
DOI: https://doi.org/10.1007/s00203-022-02929-3