Biology subjects

Callaghan, J. D.

Publications and source records attributed to Callaghan, J. D..

4 recordsLinked to original sources

bioRxiv2024

Spatio-temporal dynamics of cyanobacterium Dolichospermum lemmermannii populations in a bloom-prone region of Lake Superior

Cyanobacterial Harmful Algal Blooms (cHABs) are increasingly common in marine and freshwater environments, including the Laurentian Great Lakes (LGL). Lake Superior has seen two large-scale cyanobacterial blooms (2012 and 2018) along the Wisconsin shoreline west of the Apostle Islands, caused by the cyanobacterium Dolichospermum lemmermannii. The drivers of bloom formation in Lake Superior are not yet certain, with many factors known to trigger blooms elsewhere in the LGL being absent in Lake Superior. Furthermore, little is known about D. lemmermanniis spatial structure or phenology. Thus, we sought to track the seasonal population dynamics of D. lemmermannii to shed light on its growth, physiology, and abundance. In 2021, we used 16S rRNA amplicon and shotgun metagenomic sequencing to characterize spatiotemporal patterns of D. lemmermannii abundance and diversity along the bloom-prone Wisconsin shore of Lake Superior. In 2022, we performed net tows and direct colony counts in another localized area. No large-scale bloom event was observed during either year, though several smaller blooms were observed. D. lemmermannii abundances were low at nearly all sites and sampling times. Spikes in abundance occurred in July and September, particularly near Siskiwit Bay, a hotspot of bloom formation. We also observed a seasonal shift in heterocyte and akinete abundance indicative of late-season nutrient limitation. Most striking was the seasonal turnover of D. lemmermannii strains, suggesting strain adaptation to specific environmental conditions. These results offer valuable and actionable insights for managers and provide a foundation for additional work to clarify drivers of bloom formation in Lake Superior.

microbiology↗

bioRxiv2023

Rise in frequency of lasR mutant Pseudomonas aeruginosa among keratitis isolates between 1993 and 2021

Pseudomonas aeruginosa causes severe vision threatening keratitis. LasR is a transcription factor that regulates virulence associated genes in response to the quorum sensing molecule N-3-oxo-dodecanoyl-L-homoserine lactone. P. aeruginosa isolates with lasR mutations are characterized by an iridescent high sheen phenotype caused by a build-up of 2-heptyl-4-quinolone. A previous study indicated a high proportion (22 out of 101) of P. aeruginosa keratitis isolates from India between 2010 and 2016 were sheen positive and had mutations in the lasR gene, and the sheen phenotype correlated with worse clinical outcomes for patients. In this study, a longitudinal collection of P. aeruginosa keratitis isolates from Eastern North America were screened for lasR mutations by the sheen phenotype and sequencing of the lasR gene. A significant increase in the frequency of isolates with the sheen positive phenotype was observed in isolates between 1993 and 2021. Extracellular protease activity was lower among the sheen positive isolates and a defined lasR mutant. Cloned lasR alleles from the sheen positive isolates were loss of function or dominant negative and differed in sequence from previously reported ocular lasR mutant alleles. Insertion elements were present in a subset of independent isolates and may represent an endemic source from some of the isolates. Retrospective analysis of patient information suggested significantly better visual outcomes for patients with infected by sheen positive isolates. Together, these results indicate an increasing trend towards lasR mutations among keratitis isolates at a tertiary eye care hospital in the United States.

microbiology↗

bioRxiv2023

Nitrogen dynamics and fixation control cyanobacterial abundance, diversity, and toxicity in Lake of the Woods (USA, Canada)

Our understanding of drivers of cyanobacterial harmful algal blooms (cHABs) is evolving, but it is apparent that not all lakes are created equal. Nitrogen (N) is an important component of all cHABs and is crucial for cyanotoxin production. It is generally assumed that external nitrogen inputs are the primary N source for cHABs. However, in northern lakes, nitrogen inputs are typically low, and suggests that internal nitrogen cycling, through heterotrophic organic matter decomposition or nitrogen fixation, may play a significant role in cHAB development and sustainment. Using Lake of the Woods as a testbed, we quantified nutrients, cyanotoxins, nitrogen fixation, and the microbial community in the southern extent of the lake. During our temporal study, inorganic nitrogen species (NO3-+NO2- and NH4+) were either at very low concentrations or below detection, while phosphorus was in excess. These conditions resulted in nitrogen-deficient growth and thereby favored nitrogen fixing cyanobacterial species. In response, nitrogen fixation rates increased exponentially throughout the summer and coincided with the Aphanizomenon sp. bloom. Despite nitrogen limitation, microcystin, anatoxin, saxitoxin, and cylindrospermopsin were all detected, with microcystin being the most abundant cyanotoxin detected. Microcystin concentrations were highest when free nitrogen was available and coincided with an increase in Microcystis. Together, our work suggests that internal nitrogen dynamics are responsible for the dominance of nitrogen fixing cyanobacteria and that additions of nitrogen may increase the likelihood of other cyanobacterial species, currently at low abundance, to increase growth and cyanotoxin production. Statement of SignificanceThis study is the first assessment of nitrogen fixation rates and water column 16S rRNA gene amplicon sequencing in Lake of the Woods during a harmful algal bloom season. The aim of this study is to better understand nitrogen dynamics and the microbial ecology of cyanobacterial harmful algal blooms on Lake of the Woods. Result from this study reveal that internal nitrogen cycling via nitrogen fixation may alleviate nitrogen deficiencies, and structure and control the cyanobacterial community and cyanotoxin production. Molecular analysis reveals that cyanotoxins in Lake of the Woods are produced by less abundant cyanobacteria that are limited by nitrogen. This study has significant management implication as agencies continue to mitigate toxic blooms on Lake of the Woods, the largest shoreline lake in the United States. Our work is an important initial assessment and jumping off point for further research on Lake of the Woods when assessing how nitrogen plays a role in bloom formation and toxicity. Submitting to L&O, we believe would allow for the greatest outreach and access to an audience that will continue to build upon our findings. Additionally, submitting with L&O our work will reach beyond the scientific audience, but also reach other parties participating in the mitigation of harmful algal blooms.

microbiology↗

bioRxiv2020

Generation of Xylose-inducible promoter tools for Pseudomonas species and their use in implicating a role for the Type II secretion system protein XcpQ in inhibition of corneal epithelial wound closure

Tunable control of gene expression is an invaluable tool for biological experiments. In this study, we describe a new xylose-inducible promoter system and evaluate it in both Pseudomonas aeruginosa and P. fluorescens. The Pxut promoter derived from the P. flurorescens xut operon was incorporated into a broad host-range pBBR1-based plasmid and compared to the Escherichia coli-derived PBAD promoter using gfp as a reporter. GFP-fluorescence from the Pxut promoter was inducible in both Pseudomonas species, but not in E. coli, which may facilitate cloning of toxic genes using E. coli to generate plasmids. The Pxut promoter was expressed at a lower inducer concentration than PBAD in P. fluorescens and higher gfp levels were achieved using Pxut. Flow cytometry analysis indicated that Pxut was more leaky than PBAD in the tested Pseudomonas species, but was expressed in a higher proportion of cells when induced. D-xylose did not support growth of P. aeruginosa or P. fluorescens as a sole carbon source and is less expensive than many other commonly used inducers which could facilitate large scale applications. The efficacy of this system aided in demonstrating a role for the P. aeruginosa type II secretion system gene from xcpQ in bacterial inhibition of corneal epithelial cell wound closure. This study introduces a new inducible promoter system for gene expression for use in Pseudomonas species. ImportancePseudomonas species are enormously important in human infections, biotechnology, and as a model system for interrogating basic science questions. In this study we have developed a xylose-inducible promoter system and evaluated it in P. aeruginosa and P. fluorescens and found it to be suitable for the strong induction of gene expression. Furthermore, we have demonstrated its efficacy in controlled gene expression to show that a type 2 secretion system protein from P. aeruginosa, XcpQ, is important for host-pathogen interactions in a corneal wound closure model.

microbiology↗