Biology subjects

Andrews, J. R.

Publications and source records attributed to Andrews, J. R..

4 recordsLinked to original sources

bioRxiv2025

A colorimetric method for detecting virulent bacteriophage to Vibrio cholerae in fecal and environmental samples

BackgroundV. cholerae are often infected with vibriophages and these phages may be found associated with V. cholerae bacteria or may be detected independently from the bacteria, especially in environmental water samples. When detected, vibriophages serve as a surrogate for detection of V. cholerae and they also have an important role in the ecology of V. cholerae. Vibriophages can be detected using plaque assays or PCR, but these methods are time-consuming and require specialized laboratory resources. MethodologyTo address limitations of other methods, we developed a simple, rapid, and inexpensive colorimetric assay to detect vibriophage that can be scaled up to quickly and easily to screen a large number of samples. The assay uses resazurin and a bacterium, AC6169 that is susceptible to vibriophages ICP1, 2, and 3. Resazurin is a dye that turns color from blue to pink when added to a culture broth with growing bacteria. When a bacteria-free test sample, such as a Millipore filtered wastewater sample containing vibriophages, is added to a culture broth with AC6169, the bacteria will be lysed by the phages and will not grow, and the color of the broth will remain blue. However, if there are no phages in the sample, the bacteria will grow rapidly, and the culture broth will turn pink. We developed the assay using ICP1 spiked samples of environmental water, stool and frozen bile peptone and found it to be sensitive with a limit of detection of 4 to 40 plaque forming units/ml. ConclusionThis colorimetric assay promises to provide a convenient method to detect vibriophages on a larger scale than was possible earlier to better understand their role as a surrogate for detecting V. cholerae and to better understand their role in the pathogenesis, ecology, and epidemiology of cholera. Author SummaryWe developed a simple and inexpensive colorimetric assay to detect virulent vibriophage in samples of wastewater and stool. The assay uses resazurin, a blue dye and a unique bacterium, AC6169, that is known to be susceptible to vibriophages. Bacteria-free samples (e.g. Millipore filtrates) suspected of having vibriophage are incubated for 2 or 3 hours in a broth with AC6169, following which resazurin is added. After a further 30-minute incubation, the color of the broth is observed. If the sample contains vibriophage, the broth will remain blue, but if there are no vibriophage, the broth will turn pink. The change in color from blue to pink is due to the metabolic reaction from the growing bacteria. When vibriophage are present, they lyse the bacterium AC6169, thus preventing the color change, but if vibriophage are not present, the bacteria multiply rapidly, and the metabolic reaction causes the broth to change color to pink. Because the assay procedure is inexpensive and simple, laboratories in cholera endemic areas should find it convenient for their epidemiologic and clinical surveillance activities.

microbiology↗

bioRxiv2021

SARS-CoV-2 infects human adipose tissue and elicits an inflammatory response consistent with severe COVID-19

The COVID-19 pandemic, caused by the viral pathogen SARS-CoV-2, has taken the lives of millions of individuals around the world. Obesity is associated with adverse COVID-19 outcomes, but the underlying mechanism is unknown. In this report, we demonstrate that human adipose tissue from multiple depots is permissive to SARS-CoV-2 infection and that infection elicits an inflammatory response, including the secretion of known inflammatory mediators of severe COVID-19. We identify two cellular targets of SARS-CoV-2 infection in adipose tissue: mature adipocytes and adipose tissue macrophages. Adipose tissue macrophage infection is largely restricted to a highly inflammatory subpopulation of macrophages, present at baseline, that is further activated in response to SARS-CoV-2 infection. Preadipocytes, while not infected, adopt a proinflammatory phenotype. We further demonstrate that SARS-CoV-2 RNA is detectable in adipocytes in COVID-19 autopsy cases and is associated with an inflammatory infiltrate. Collectively, our findings indicate that adipose tissue supports SARS-CoV-2 infection and pathogenic inflammation and may explain the link between obesity and severe COVID-19. One sentence summaryOur work provides the first in vivo evidence of SARS-CoV-2 infection in human adipose tissue and describes the associated inflammation.

immunology↗

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

Genomic variant identification methods alter Mycobacterium tuberculosis transmission inference

Pathogen genomic data are increasingly used to characterize global and local transmission patterns of important human pathogens and to inform public health interventions. Yet there is no current consensus on how to measure genomic variation. We investigated the effects of variant identification approaches on transmission inferences for M. tuberculosis by comparing variants identified by five different groups in the same sequence data from a clonal outbreak. We then measured the performance of commonly used variant calling approaches in recovering variation in a simulated tuberculosis outbreak and tested the effect of applying increasingly stringent filters on transmission inferences and phylogenies. We found that variant calling approaches used by different groups do not recover consistent sets of variants, often leading to conflicting transmission inferences. Further, performance in recovering true outbreak variation varied widely across approaches. Finally, stringent filters rapidly eroded the accuracy of transmission inferences and quality of phylogenies reconstructed from outbreak variation. We conclude that measurements of genetic distance and phylogenetic structure are dependent on variant calling approach. Variant calling algorithms trained upon true sequence data outperform other approaches and enable inclusion of repetitive regions typically excluded from genomic epidemiology studies, maximizing the information gleaned from outbreak genomes.

genomics↗