Characterisation of the koala (Phascolarctos cinereus) pouch microbiota in a captive population reveals a dysbiotic compositional profile associated with neonatal mortality

Study information

Reviewed Marked as Reviewed by Folakunmi on 2024-4-3

study design

laboratory experiment

Citation

PMID PubMed identifier for scientific articles.

37060097

DOI Digital object identifier for electronic documents.

https://doi.org/10.1186/s40168-023-01527-9

URI

Authors

Maidment TI, Bryan ER, Pyne M, Barnes M, Eccleston S, Cunningham S, Whitlock E, Redman K, Nicolson V, Beagley KW, Pelzer E

Journal

Microbiome

Year

2023

Keywords:

Dysbiosis, Endangered species, Enterobacteriaceae, Klebsiella pneumoniae, Koala, Marsupial, Muribaculaceae, Phascolarctos cinereus, Pluralibacter gergoviae, Pouch, Reproduction

BACKGROUND: Captive koala breeding programmes are essential for long-term species management. However, breeding efficacy is frequently impacted by high neonatal mortality rates in otherwise healthy females. Loss of pouch young typically occurs during early lactation without prior complications during parturition and is often attributed to bacterial infection. While these infections are thought to originate from the maternal pouch, little is known about the microbial composition of koala pouches. As such, we characterised the koala pouch microbiome across the reproductive cycle and identified bacteria associated with mortality in a cohort of 39 captive animals housed at two facilities. RESULTS: Using 16S rRNA gene amplicon sequencing, we observed significant changes in pouch bacterial composition and diversity between reproductive time points, with the lowest diversity observed following parturition (Shannon entropy - 2.46). Of the 39 koalas initially sampled, 17 were successfully bred, after which seven animals lost pouch young (overall mortality rate - 41.18%). Compared to successful breeder pouches, which were largely dominated by Muribaculaceae (phylum - Bacteroidetes), unsuccessful breeder pouches exhibited persistent Enterobacteriaceae (phylum - Proteobacteria) dominance from early lactation until mortality occurred. We identified two species, Pluralibacter gergoviae and Klebsiella pneumoniae, which were associated with poor reproductive outcomes. In vitro antibiotic susceptibility testing identified resistance in both isolates to several antibiotics commonly used in koalas, with the former being multidrug resistant. CONCLUSIONS: This study represents the first cultivation-independent characterisation of the koala pouch microbiota, and the first such investigation in marsupials associated with reproductive outcomes. Overall, our findings provide evidence that overgrowth of pathogenic organisms in the pouch during early development is associated with neonatal mortality in captive koalas. Our identification of previously unreported, multidrug resistant P. gergoviae strains linked to mortality also underscores the need for improved screening and monitoring procedures aimed at minimising neonatal mortality in future. Video Abstract.

Experiment 1

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Reviewed Marked as Reviewed by Folakunmi on 2024-4-3

Curated date: 2024/03/06

Curator: Ehi

Revision editor(s): Ehi, Folakunmi

Subjects

Location of subjects: Australia

Host species Species from which microbiome was sampled. Contact us to have more species added.: Phascolarctos cinereus

Body site Anatomical site where microbial samples were extracted from according to the Uber Anatomy Ontology: Marsupium Marsupial pouch,Pouch,Marsupium,marsupium

Condition The experimental condition / phenotype studied according to the Experimental Factor Ontology: Neonatal death Neonatal lethal,Neonatal death,neonatal death

Group 0 name Corresponds to the control (unexposed) group for case-control studies: Successful breeders

Group 1 name Corresponds to the case (exposed) group for case-control studies: Unsuccessful breeders

Group 1 definition Diagnostic criteria applied to define the specific condition / phenotype represented in the case (exposed) group: Captive Koalas who lost their pouch young during lactation.

Group 0 sample size Number of subjects in the control (unexposed) group: 10

Group 1 sample size Number of subjects in the case (exposed) group: 7

Lab analysis

Sequencing type: 16S

16S variable region One or more hypervariable region(s) of the bacterial 16S gene: V3-V4

Sequencing platform Manufacturer and experimental platform used for quantifying microbial abundance: Illumina

Statistical Analysis

Data transformation Data transformation applied to microbial abundance measurements prior to differential abundance testing (if any).: raw counts

Statistical test: DESeq2

Significance threshold p-value or FDR threshold used for differential abundance testing (if any): 0.05

MHT correction Have statistical tests be corrected for multiple hypothesis testing (MHT)?: Yes

Alpha Diversity

Shannon Estimator of species richness and species evenness: more weight on species richness: unchanged

Richness Number of species: unchanged

Signature 1

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Reviewed Marked as Reviewed by Folakunmi on 2024-4-3

Curated date: 2024/03/07

Curator: Ehi

Revision editor(s): Ehi, Folakunmi

Source: supplementary Table 11

Description: Differentially abundant genera in the pouch microbiota of successful vs. unsuccessful breeding koalas during early lactation.

Abundance in Group 1: increased abundance in Unsuccessful breeders

NCBI	Quality Control	Links
Phascolarctobacterium faecium
Pseudomonas
Escherichia/Shigella sp.
Klebsiella
Pluralibacter
Lonepinella

Revision editor(s): Ehi, Folakunmi

Signature 2

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Your review change was submitted

Reviewed Marked as Reviewed by Folakunmi on 2024-4-3

Curated date: 2024/03/07

Curator: Ehi

Revision editor(s): Ehi, Folakunmi

Source: supplementary Table 11

Description: Differentially abundant genera in the pouch microbiota of successful vs. unsuccessful breeding koalas during early lactation.

Abundance in Group 1: decreased abundance in Unsuccessful breeders

NCBI	Quality Control	Links
Muribaculum
Phascolarctobacterium faecium

Revision editor(s): Ehi, Folakunmi