Draft genome of Kazachstania heterogenica var. weizmannii, a commensal fungus of the mouse digestive tract

TL;DR

This paper presents the genome of a commensal fungus found in mice, which is part of a fungal complex typically associated with disease.

Contribution

The discovery and genome sequencing of a commensal strain of Kazachstania heterogenica from mice gut.

Findings

A commensal strain of Kazachstania heterogenica, named var. weizmannii, was isolated from the mouse gut.

The genome of K. heterogenica var. weizmannii was sequenced, providing insights into its commensal nature.

Abstract

Kazachstania heterogenica is a member of the K. telluris complex, where all members to date are reported to be pathogenic fungi. We have isolated a strain, K. heterogenica var. weizmannii, from the gut of mice that seems to be a commensal strain and sequenced its genome.

Tables

TABLE 1. Sequencing statistics

Illumina read pairs	5820035
Illumina bases	1,737,163,443 bp
Illumina coverage	121×
Nanopore Raw Read N50	5,450 bp
Nanopore trimmed reads	400,977
Nanopore trimmed bases	951,032,193 bp
Nanopore coverage	66×

Full text

ANNOUNCEMENT

Kazachstania heterogenica is a fungus in the Kazachstania telluris species complex of the family Saccharomycetaceae. Other complex members (K. telluris, K. bovina, K. slooffiae, and K. pintolopesii) were isolated from various hosts in the context of pathogenicity (1). We isolated a putative commensal fungus from mouse feces while studying the effects of Candida albicans infection on immunocompromised mice. Our isolate has not shown signs of pathogenicity (2).

K. heterogenica var. weizmannii was cultured from the feces of mice housed in an SPF facility (2) and grown on solid YPD media at 37°C. DNA was isolated from fungal pellets using a DNeasy Blood & Tissue Kit (Qiagen, cat. 69504). Sequencing and hybrid (Nanopore and Illumina) assembly were performed by SeqCenter (Pittsburgh, PA), as follows: Illumina libraries were prepared using an Illumina DNA Prep kit and IDT 10 bp UDI indices, and sequenced on an Illumina NextSeq 2000 producing 2 × 151 bp reads. For all programs mentioned, default parameters were used except where otherwise noted. The data were demultiplexed and adapters were removed using bcl2fastq (v2.20.0.445) (3). Samples were prepared for sequencing using Oxford Nanopore’s “Genomic DNA by Ligation” kit (SQK-LSK109) with no fragmentation or size selection. The samples were run on Nanopore R9 flow cells (R9.4.1) on a MinION. Basecalling was performed with Guppy (version 4.2.2), in high-accuracy mode (Default parameters + effbaf8). Quality control and adapter trimming were performed with Porechop (version 0.2.2_seqan2.1.1) (4). Statistics are shown in Table 1.

Long read assembly with ONT reads was performed with flye (5) (version 2.8). The long read assembly was polished with Pilon (1.23) for one round of base correction (6). Final assembly statistics were recorded with QUAST (5.0.2) (7). The final assembly of 33 contigs has a length of 14.37 Mb, 30.39% GC, and an N50 of 1.03 Mb.

Annotation was performed with the Yeast Gene Annotation Pipeline (YGAP) (8) with the Post-WGD settings, and Companion (9) for Fungi with the reference organism Candida glabrata CBS138. The output of both programs was compared with CD-HIT (version 4.8.1) (10) with c = 1 to reduce redundancy, and the remaining genes were combined using YGAP as the base annotation using in-house scripts. The assembly has 5,217 protein-coding genes, 223 tRNAs, and 7 putative long non-coding RNA genes. The genome has 86.9% complete BUSCOs (11) (v 5.4.6; analysis was performed on the proteins using the Galaxy website usegalaxy.org (12).

We compared our sequence to the K. heterogenica type strain NRRL-Y27499, as follows: the Illumina reads of the type strain (NCBI accession: SRR25935784) were mapped to the K. heterogenica var. weizmannii genome using BWA-mem (0.7.13) (13), and variant calling was performed with GATK (4.4.0.0) (14), SAMtools (1.9) (15), and Picard (2.22.8) (16). 96.34% of the reads mapped to the genome of our isolate. There were 18,779 SNPs in the entire genome, for a 0.13% difference. In addition, there were 7,506 indels, though most were short and in repetitive DNA elements. Taken together with virtually no differences in the rDNA locus (only one difference in 18S and two differences in ITS2 out of the entire locus), we believe that our isolate is Kazachstania heterogenica and should be known as the weizmannii strain.