Biology subjects

Spormann, A. M.

Publications and source records attributed to Spormann, A. M..

4 recordsLinked to original sources

bioRxiv2022

Double emulsions as a high-throughput enrichment and isolation platform for slower-growing microbes

Our understanding of in situ microbial physiology is primarily based on physiological characterization of fast-growing and readily-isolatable microbes. Microbial enrichments to obtain novel isolates with slower growth rates or physiologies adapted to low nutrient environments are plagued by intrinsic biases for fastest-growing species when using standard laboratory isolation protocols. New cultivation tools to minimize these biases and enrich for less well-studied taxa are needed. In this study, we developed a high-throughput bacterial enrichment platform based on single cell encapsulation and growth within double emulsions (GrowMiDE). We showed that GrowMiDE can cultivate many different microorganisms and enrich for novel taxa that are never observed in traditional batch enrichments. For example, preventing dominance of the enrichment by fast-growing microbes due to nutrient privatization within the double emulsion droplets allowed cultivation of novel Negativicutes and Methanobacteria from stool samples in rich media enrichment cultures. In competition experiments between growth rate and growth yield specialist strains, GrowMiDE enrichments prevented competition for shared nutrient pools and enriched for slower-growing but more efficient strains. Finally, we demonstrated the compatibility of GrowMiDE with commercial fluorescence-activated cell sorting (FACS) to obtain isolates from GrowMiDE enrichments. Together, GrowMiDE + DE-FACS is a promising new high-throughput enrichment platform that can be easily applied to diverse microbial enrichments or screens.

microbiology↗

bioRxiv2022

Metabolic diversity in commensal protists regulates intestinal immunity and trans-kingdom competition

The microbiota influences intestinal health and physiology, yet the contributions of commensal protists to the gut environment have been largely overlooked. Here, we identified several new rodent- and human-associated parabasalid protists. Genomic and metabolomic analyses of murine parabasalids from the genus Tritrichomonas revealed species-level differences in the excretion of the metabolite succinate. This metabolic dissimilarity results in distinct small intestinal immune responses during protist colonization. Metabolic differences between Tritrichomonas species also determine their ecological niche within the microbiota. By manipulating dietary fibers and developing in vitro protist culture, we show that different parabasalid species preferentially rely on dietary polysaccharides or mucus glycans. These polysaccharide preferences create trans-kingdom competition with specific commensal bacteria, which affects intestinal immunity in a diet-dependent manner. Our findings reveal unappreciated diversity in commensal parabasalids, elucidate differences in commensal protist metabolism, and suggest how dietary interventions could regulate their impact on gut health.

microbiology↗

bioRxiv2020

Gold-standard diagnostics are tarnished by lytic bacteriophage

BackgroundA fundamental clinical and scientific concern is how lytic bacteriophage, as well as antibiotics, impact diagnostic positivity. MethodsCholera was chosen as a model disease to investigate this important question. Patients with diarrheal disease were enrolled at two remote hospitals in Bangladesh. Diagnostic performance was assessed as a function of lytic bacteriophage detection, as well as exposure to the first-line antibiotic azithromycin detected by mass spectrometry. ResultsAmong diarrheal samples positive by nanoliter quantitative PCR for Vibrio cholerae (n=78/849), the odds that a rapid diagnostic test (RDT) or qPCR was positive was reduced by 89% (OR 0.108; 95%CI 0.002-0.872) and 87% (OR 0.130; 95%CI 0.022-0.649) when lytic bacteriophage were detected, respectively. The odds that a rapid diagnostic test (RDT) or qPCR was positive was reduced by more than 99% (OR 0.00; 95% CI: 0.00-0.28) and 89% (OR 0.11; 95% CI: 0.03-0.44) when azithromycin was detected, respectively. ConclusionsEstimations of cholera burden may improve by accommodating for the negative effect of antimicrobial exposure on diagnostic positivity. Furthermore, the findings herein challenge our current approach to interpreting and developing bacterial diagnostics given variable rates of lytic bacteriophage and antibiotic exposure.

microbiology↗

bioRxiv2019

High-throughput multi-parallel enteropathogen quantification via nano-liter qPCR

Quantitative molecular diagnostic methods, such as qPCR, can effectively detect pathogen-specific nucleic acid sequences. However, costs associated with multi-pathogen quantitative molecular diagnostics hinder their widespread use. Nano-liter qPCR (nL-qPCR) is a miniaturized tool for quantification of multiple targets in large numbers of samples based on assay parallelization on a single chip, with potentially significant cost-savings due to rapid throughput and reduced reagent volumes. We evaluated a suite of novel and published assays to detect 17 enteric pathogens using a commercially available nL-qPCR technology. Assay efficiencies ranged from 88-98% (mean 91%) and were reproducible across four operators at two separate facilities. When applied to complex fecal material, assays were sensitive and selective (99.8% of DNA amplified were genes from the target organism). Detection limits were 1-2 orders of magnitude higher for nL-qPCR than an existing enteric TaqMan Array Card (TAC), due to nanofluidic volumes. Compared to the TAC, nL-qPCR displayed 97% (95% CI 0.96, 0.98) negative percent agreement and 63% (95% CI 0.60, 0.66) overall positive percent agreement. Positive percent agreement was 90% for target concentrations above the nL-qPCR detection limits. nL-qPCR assays showed an underestimation bias of 0.34 log10 copies/gram of stool [IQR -0.41, -0.28] compared with the enteric TAC. Higher detection limits, inherent to nL-qPCR, do not hinder detection of clinically relevant pathogen concentrations. With 12 times higher throughput for a sixth of the per-sample cost of the enteric TAC, the nL-qPCR chip described here is a viable alternative for enteropathogen quantification for studies where other technologies are cost-prohibitive.

microbiology↗