Biology subjects

Fuller, D. T.

Publications and source records attributed to Fuller, D. T..

3 recordsLinked to original sources

bioRxiv2024

Dirichlet distribution parameter estimation with application in microbiome analyses

Microbiomes are of vital importance for understanding human and environmental health. However, quantifying microbial composition remains challenging and relies on statistical modeling of either the raw taxonomic counts or the relative abundances. Relative abundance measures are commonly preferred over the absolute counts to analyze and interpret microbiome (as the sampling fraction are unknown in sequence data) but currently there is no ideal distribution for carrying out this modeling . In this work, the Dirichlet distribution is proposed to model the relative abundances of taxa directly without the use of any further transformation. In a comprehensive simulation study, we compared biases and standard errors of two Methods of Moments Estimators (MMEs) and Maximum Likelihood Estimator (MLE) of the Dirichlet distribution. Comparison of each estimator is done over three different cases of differing sample size and dimension: (i) small dimension and small sample size; (ii) small dimension and large sample size; (iii) large dimension with both large and small sample size. We demonstrate the Dirichlet modeling methodology with four real world microbiome datasets and show how the results of the Dirichlet model differ from those obtained by a commonly used method, namely Bayesian Dirichlet-Multinomial estimation (BDME). We find that the results of parameter estimation can be dependent upon the sequencing depth and sequencing technique used to produce a given microbiome dataset. However, for all datasets, the Dirichlet MLE (DMLE) results are comparable to the BDME results while requiring less computational time in each case.

bioinformatics↗

bioRxiv2023

Application of nanopore sequencing for accurate identification of bacterial colonies

Culture based detection remains to be one of the most reliable and acceptable techniques to detect extremely low quantity pathogens present in a sample. The process typically involves inoculating the sample on an agar plate to allow growth of the microorganisms to form colonies, followed by the identification of the individual colonies, commonly by DNA sequencing of a PCR-amplified targeted gene. Sanger method is often the default choice of sequencing as it offers affordable and accurate results for a single species. However, the technique could pose limitations in certain situations such as identification of multi-species microbial colonies. In this work, we compared the performance of Sanger sequencing with MinION nanopore sequencing in identifying bacterial colonies derived from bioaerosol samples. We conducted Sanger and nanopore sequencing of full-length 16S rRNA genes from seven bacterial colonies derived from bioaerosol samples and compared the outcome by alignment against NCBI 16S reference database. We found that for five out of seven colonies both techniques indicated the presence of the same bacterial genus. For one of the remaining colonies, a noisy Sanger electropherogram failed to generate a meaningful sequence, but nanopore sequencing identified it to be a mix of two bacterial genera Alkalihalobacillus and Kocuria. For the other remaining colony, the Sanger sequencing suggested Micrococcus with a clean electropherogram, however, the nanopore sequencing suggested the presence of an additional genus Paraburkholderia. Further corroborating these findings with mock multispecies colonies from pure bacterial DNA samples, we confirm that nanopore sequencing is comparable to the Sanger method in identifying colonies with single bacterial species but is the superior method in classifying individual bacterial components with their relative abundances in multispecies colonies. Our results suggest that nanopore sequencing could be advantageous over Sanger sequencing for colony identification in culture-based analysis of environmental samples such as bioaerosol where direct inoculation of the sample to culture plate might lead to formation of multispecies colonies.

microbiology↗

bioRxiv2020

Digital spatial profiling of coronary plaques from persons living with HIV reveals high levels of STING and CD163 in macrophage enriched regions

BackgroundChronic innate and adaptive immune activation may contribute to high prevalence of cardiovascular disease in persons living with HIV (PLWH). MethodsWe assessed coronary plaques from deceased PLWH (n=6) and HIV-negative (n=6) persons matched by age and gender. Formalin-fixed, paraffin-embedded 5m thick sections were processed using Movat, hematoxylin and eosin, immunohistochemical and immunofluorescence stains. Immune cell populations were measured using surface antibodies, and immune-related protein expression from macrophage rich, T-cell rich and perivascular adipose tissue regions using GeoMx(R) digital spatial profiling. ResultsCoronary plaques from PLWH and HIV-negative persons had similar plaque area and percent stenosis. Percent CD163+ cells as measured by immunohistochemical staining was significantly higher in PLWH, median 0.29% (IQR 0.11-0.90) vs. 0.01% (IQR 0.0013-0.11) in HIV-negative plaque, p = 0.02 (Figure 1A). Other surface markers of innate cells (CD68 +, p=0.18), adaptive immune cells (CD3+, p=0.39; CD4+, p=0.09; CD8+, p=0.18) and immune trafficking markers (CX3CR1+, p=0.09) within the coronary plaque trended higher in HIV-positive plaques but did not reach statistical significance. GeoMx(R) digital spatial profiling showed higher differential protein expression of CD163 (scavenger receptor for hemoglobin-haptoglobin complex), stimulator of interferon gamma (STING, a cytosolic DNA sensor), CD25 and granzyme-B in the HIV-positive compared to HIV-negative, p<0.05(Figure 1B). O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/221325v1_fig1.gif" ALT="Figure 1"> View larger version (51K): org.highwire.dtl.DTLVardef@1a4757forg.highwire.dtl.DTLVardef@1e2236org.highwire.dtl.DTLVardef@582f4borg.highwire.dtl.DTLVardef@188f376_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 1.C_FLOATNO Characterization of coronary plaques. Representative coronary plaques from three HIV-positive and three HIV-negative individuals were stained with H&E and Movat stain (A). Plaque area (m2) and % plaque stenosis were measured and calculated in 6 individuals per group (B). Statistical analysis, Mann-Whitney; ns not significant C_FIG ConclusionsIncreased inflammation within the coronary plaques of PLWH is characterized by more innate and adaptive immune cells. Higher STING expression in PLWH suggests that immune response to viral antigens within the plaque might be a driver above other stimulants. STING inhibitors are available and could be investigated as a future therapeutic target in PWH if these results are replicated with a larger number of plaques. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=146 SRC="FIGDIR/small/221325v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@27fa00org.highwire.dtl.DTLVardef@8e4a65org.highwire.dtl.DTLVardef@1b65320org.highwire.dtl.DTLVardef@10b7c9a_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIImmunohistochemical and fluorescent stains combined with GeoMx(R) digital spatial profiling allowed for deep characterization of immune cells within intact coronary plaques and perivascular adipose tissue C_LIO_LICoronary plaques from HIV-positive persons had higher proportion of CD163+ immune cells compared to HIV-negative persons C_LIO_LIDifferential protein expression of immune-rich regions of interest within intact 5m sections of coronary plaques revealed higher levels of stimulator of interferon gamma (STING) in HIV-positive persons C_LI

immunology↗