Draft genome sequences for 6 isolates of endospore forming class Bacilli species isolated from soil from a suburban, wooded, developed space

Introduction

The publication by [Anna L. McLoon, Thomas T. Awad, Molly F. Bogardus, Meredith G. Buono, Kaitlyn A. Devine, Rebecca M. Draper, Brianna Femenella, Hannah M. Gallagher, Laura A. Morelock, Mishal Razi, Jacqueline R. Rennick, Abigail K. Sheridan, Righlee J. Thibault, Katie L. Touchette, and Grace E. Zuchowski] can be found here: [URL]

Table of Contents

1. Background and Experimental Methods
2. Import and annotation
3. QC, Assembly, and Annotation
4. Taxonomic Classification
5. References

Access to Narrative workflows and sequence data

Background and Experimental Methods

Sample Collection

We collected a soil sample on January 5, 2022 from a heavily impacted wooded area between a parking lot and a marsh overgrown with phragmites reeds on the Siena College campus (42°43'11.0"N 73°44'59.6"W 42.719711, -73.749876) near a willow (Salix sp.) tree. The soil in the area is Stafford loamy fine sand, and was frozen at the time of collection.

Isolation

Subsamples of soil were suspended in water and boiled for 10 minutes to isolate endospores, then cultured on 5% sheep blood TSA plates overnight at 37ºC. 6 isolates were recultured and assessed via Gram staining and other tests to ensure purity. All grow well on TSA at temperatures between 22 and 37 ºC. All 6 isolates are in the Bacilli class (3), several in newly reclassified genera (4, 5).

Genome Sequencing

For genomic DNA isolation, cells were grown in tryptic soy broth at 37 ºC for 3-4 hours, until the culture was visibly dense. Cells were resuspended in 50 mM EDTA, and peptidoglycan was digested for 1 hour at 37 ºC with the addition of 30 µl of 20 mg/ml lysozyme, then genomic DNA was purified using a Promega Wizard Genomic DNA purification kit. Genomic DNA was sequenced as 151bp paired end reads using an Illumina NextSeq2000 by the Microbial Genome Sequencing Center, Pittsburgh, PA.

QC, Assembly, and Annotation

Adapter sequences were removed by the sequencing facility. All subsequent analysis steps were carried out within the KBase environment (6, 7). Analyses were carried out in parallel for each isolate in its own Narrative. Sequence quality was validated with FastQC v0.11.9 (8), then reads were trimmed with Trimmomatic v0.36 (9). Trimmed reads were assembled into genome assemblies using SPAdes v3.15.3, annotated using RAST v1.073 and/or Prokka v.1.14.5, the species identity was assigned using GTDB-Tk v1.7.0 and TYGS, and metabolic predictions were made using DRAM v0.1.0 (10–21). Default parameters were used. Genome assemblies range in size from 4.38 to 6.07 Mb bp and are AT rich (Table 1).

Taxonomic Identification

To assign each isolate to the appropriate genus and, where possible, species, we used the GTDB-Tk classification program v1.7.0 using the draft genome assemblies. Each identification was independently achieved using the TYGS type strain server (Meier-Kolthoff et al., 2022). See the individual narratives for full taxonomies, with genus and species summarized here:

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