Stable isotope probing with 18O-water to investigate microbial growth and death in environmental samples.

Growth and mortality of microorganisms have been characterized through DNA stable isotope probing (SIP) with 18O-water in soils from a range of ecosystems. Conventional SIP has been improved by sequencing a marker gene in all fractions retrieved from an ultracentrifuge tube to produce taxon density curves, which allow estimating the atom percent isotope composition of each microbial taxon's genome. Very recent advances in SIP with 18O-water include expansion of the technique to aquatic samples, investigations of microbial turnover in soil, and the first use of 18O-water in RNA-SIP studies.

Impact of saline irrigation and topical corticosteroids on the postsurgical sinonasal microbiota.

BACKGROUND: Topical treatments with nasal saline irrigation, topical steroid sprays, or corticosteroid rinses can improve sinonasal symptoms in chronic rhinosinusitis (CRS). However, the impact of these therapies on commensals (Corynebacterium) and on biofilm pathogens associated with CRS (Staphylococcus aureus and Pseudomonas) is not well characterized. METHODS: Paired nasal and sinus swabs were collected endoscopically from 28 controls and 14 CRS patients with nasal polyposis (CRSwNP) who had not received systemic antibiotics or corticosteroids in the previous 8 weeks. Total DNA from swab eluents were extracted and analyzed by 16S rRNA gene-based pyrosequencing. A total of 359,077 reads were obtained and classified taxonomically. The association of use of topical therapies with sinonasal microbiota composition was assessed by factor/vector-fitting. The proportional abundances of sinonasal bacteria between topical therapy users and nonusers were further compared by 2-tailed Kolmogorov-Smirnov test among controls and among CRSwNP participants. RESULTS: Nasal saline irrigation, with or without added budesonide, was not associated with significantly distinct sinonasal microbiota composition or significantly decreased Pseudomonas or S. aureus abundances among either controls or CRSwNP participants. Corynebacterium was slightly lower in controls that reported using saline irrigation than those who did not. No significant association was found between nasal saline irrigation and the proportional abundances of Pseudomonas, S. aureus, and Corynebacterium in CRSwNP participants. However, male CRSwNP patients were noted to have significantly higher Corynebacterium proportional abundances than their female counterparts. The use of topical steroid sprays was associated with a distinct microbiota in control subjects, characterized by higher proportional abundances of Dolosigranulum and Simonsiella and a lower proportional abundance of Campylobacter. CONCLUSION: Nasal saline irrigation is not associated with a distinct alteration in the proportional abundance of commensal bacteria or biofilm-forming pathogens in CRSwNP patients. However, use of topical intranasal corticosteroid sprays in control subjects is associated with a distinct sinonasal microbiota.

Rapid differentiation between livestock-associated and livestock-independent Staphylococcus aureus CC398 clades.

Staphylococcus aureus clonal complex 398 (CC398) isolates cluster into two distinct phylogenetic clades based on single-nucleotide polymorphisms (SNPs) revealing a basal human clade and a more derived livestock clade. The scn and tet(M) genes are strongly associated with the human and the livestock clade, respectively, due to loss and acquisition of mobile genetic elements. We present canonical single-nucleotide polymorphism (canSNP) assays that differentiate the two major host-associated S. aureus CC398 clades and a duplex PCR assay for detection of scn and tet(M). The canSNP assays correctly placed 88 S. aureus CC398 isolates from a reference collection into the human and livestock clades and the duplex PCR assay correctly identified scn and tet(M). The assays were successfully applied to a geographically diverse collection of 272 human S. aureus CC398 isolates. The simple assays described here generate signals comparable to a whole-genome phylogeny for major clade assignment and are easily integrated into S. aureus CC398 surveillance programs and epidemiological studies.

Medical therapy reduces microbiota diversity and evenness in surgically recalcitrant chronic rhinosinusitis.

BACKGROUND: Chronic rhinosinusitis (CRS) is a highly prevalent and heterogeneous condition frequently treated with antibiotics and corticosteroid therapy. However, the effect of medical therapy on sinus microbiota remains unknown. METHODS: We enrolled CRS patients (n = 6) with patent maxillary antrostomies and active mucosal inflammation, who had not received antibiotics or corticosteroids in the previous 8 weeks. A pretreatment and posttreatment maxillary sinus swab was collected, from which DNA was extracted, pyrosequenced, and analyzed using a naïve Bayesian classifier and ecological analyses. RESULTS: Four patients showed significant improvement in endoscopic appearance. The shifts in microbiota in response to therapy were highly individualized. There was no single common microbiota profile among patients with similar clinical outcomes, but overall there was significant decrease in microbiota diversity (t(5) = 2.05, p = 0.10) and evenness (t(5) = 2.28, p = 0.07) after treatment. CONCLUSION: Our findings strongly correlate with earlier studies that examined the impact of antibiotics on human microbiota. We observed that posttreatment, patients frequently became colonized by taxa that are less susceptible to the prescribed antibiotics. Our findings highlight the challenge in seeking generalizable diagnostic and therapeutic options in CRS, particularly regarding microbiological response and outcomes.