Biology subjects

Beall, C. J.

Publications and source records attributed to Beall, C. J..

3 recordsLinked to original sources

bioRxiv2023

Fungi and Bacteria Occupy Distinct Spatial Niches within Carious Dentin

The role of bacteria in the etiology of dental caries is long established, while the role of fungi has only recently gained more attention. The microbial invasion of dentin in advanced caries especially merits additional research. We evaluated the fungal and bacterial community composition and spatial distribution within carious dentin. Amplicon 16S rRNA gene sequencing together with quantitative PCR was used to profile bacterial and fungal species in caries-free children (n=43) and 4 stages of caries progression from children with severe early childhood caries (n=32). Additionally, healthy (n=10) and carious (n=10) primary teeth were decalcified, sectioned, and stained with Grocotts methenamine silver, periodic acid Schiff (PAS) and calcofluor white (CW) for fungi. Immunolocalization was also performed using antibodies against fungal {beta}-D-glucan, gram-positive bacterial lipoteichoic acid, Streptococcus mutans, and Candida albicans. We also performed field emission scanning electron microscopy (FESEM) to visualize fungi and bacteria within carious dentinal tubules. Bacterial communities observed included a high abundance of S. mutans and the Veillonella parvula group, as expected. There was a higher ratio of fungi to bacteria in dentin-involved lesions compared to less severe lesions with frequent preponderance of C. albicans, C. dubliniensis, and in one case C. tropicalis. Grocotts silver, PAS, CW and immunohistochemistry (IHC) demonstrated the presence of fungi within carious dentinal tubules. Multiplex IHC revealed that fungi and gram-positive bacteria primarily occupied separate dentinal tubules, with rare instances of colocalization. Similar findings were observed with multiplex immunofluorescence using anti-S. mutans and anti-C. albicans antibodies. Electron microscopy showed monomorphic bacterial and fungal biofilms within distinct dentin tubules. We demonstrate a previously unrecognized phenomenon in which fungi and bacteria occupy distinct spatial niches within carious dentin and seldom co-colonize. The potential significance of this phenomenon in caries progression warrants further exploration.

microbiology↗

bioRxiv2019

Oral bacteriophages are maintained at high levels for months in individuals but infrequently transmitted between mothers and infants

In this work, we exploit recent advances in metagenomic assembly and bacteriophage identification to describe the phage content of saliva from 5 mother-baby pairs sampled twice 7 - 11 months apart during the first year of the babies lives. We identify 25 phage genomes that are comprised of one to 71 contigs, with 16 having a single contig. At the detectable level, phage were sparsely distributed with the most common one being present in 4 of the 20 samples, derived from two mothers and one baby. However, if they were present in the early time point sample from an individual, they were also present in the later sample from the same person more frequently than expected by chance. The nucleotide diversity ({pi}) in phage from the same sample or the same person was much lower than between different individuals, indicating dominance of one strain in each person. This was different from bacterial genomes, which had higher diversity indicating the presence of multiple strains within an individual. We identify likely bacterial hosts for 16 of the 25 phage, including an apparent inovirus that is capable of integrating in the dif site of Haemophilus species. It appears that phage in the oral cavity are sparsely distributed, but can be maintained for months once acquired.

microbiology↗

bioRxiv2018

High-resolution ISR amplicon sequencing reveals personalized oral microbiome

Background: Sequencing of the 16S rRNA gene has been the standard for studying the composition of microbial communities. While it allows identification of bacteria at the level of species, it does not usually provide sufficient information to resolve at the sub-species level. Species-level resolution is not adequate for studies of transmission or stability, or for exploring subspecies variation in disease association. Current approaches using whole metagenome shotgun sequencing require very high coverage that can be cost-prohibitive and computationally challenging for diverse communities. Thus there is a need for high-resolution, yet cost-effective, high-throughput methods for characterizing microbial communities.\n\nResults: Significant improvement in resolution for amplicon-based bacterial community analysis was achieved by combining amplicon sequencing of a high-diversity marker gene, the ribosomal operon ISR, with a probabilistic error modeling algorithm, DADA2. The resolving power of this new approach was compared to that of both standard and high-resolution 16S-based approaches using a set of longitudinal subgingival plaque samples. The ISR strategy achieved a 5.2-fold increase in community richness compared to reference-based 16S rRNA gene analysis, and showed 100% accuracy in predicting the correct source of a clinical sample. Individuals microbial communities were highly personalized, and although they exhibited some drift in membership and levels over time, that difference was always smaller than the differences between any two subjects, even after one year. The construction of an ISR database from publicly available genomic sequences allowed us to explore genomic variation within species, resulting in the identification of multiple variants of the ISR for most species.\n\nConclusions: The ISR approach resulted in significantly improved resolution of communities, and revealed a highly personalized, stable human oral microbiota. Multiple ISR types were observed for all species examined, demonstrating a high level of subspecies variation in the oral microbiota. The approach is high-throughput, high-resolution yet cost-effective, allowing subspecies-level community fingerprinting at a cost comparable to that of 16S rRNA gene amplicon sequencing. It will be useful for a range of applications that require high-resolution identification of organisms, including microbial tracking, community fingerprinting, and potentially for identification of virulence-associated strains.

microbiology↗