Search

Background Sequencing of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viral genome from patient samples is an important epidemiological tool for monitoring and responding to the pandemic, including the emergence of new mutations in specific communities. Methods SARS-CoV-2 genomic sequences were generated from positive samples collected, along with epidemiological metadata, at a walk-up, rapid testing site in the Mission District of San Francisco, California during 22 November to 1 December, 2020, and 10–29 January 2021. Secondary household attack rates and mean sample viral load were estimated and compared across observed variants. Results A total of 12 124 tests were performed yielding 1099 positives. From these, 928 high-quality genomes were generated. Certain viral lineages bearing spike mutations, defined in part by L452R, S13I, and W152C, comprised 54.4% of the total sequences from January, compared to 15.7% in November. Household contacts exposed to the "California" or "West Coast" variants (B.1.427 and B.1.429) were at higher risk of infection compared to household contacts exposed to lineages lacking these variants (0.36 vs 0.29, risk ratio [RR] = 1.28; 95% confidence interval [CI]: 1.00–1.64). The reproductive number was estimated to be modestly higher than other lineages spreading in California during the second half of 2020. Viral loads were similar among persons infected with West Coast versus non-West Coast strains, as was the proportion of individuals with symptoms (60.9% vs 64.3%). Conclusions The increase in prevalence, relative household attack rates, and reproductive number are consistent with a modest transmissibility increase of the West Coast variants. Summary : We observed a growing prevalence and modestly elevated attack rate for "West Coast" severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants in a community testing setting in San Francisco during January 2021, suggesting its modestly higher transmissibility. [ABSTRACT FROM AUTHOR]

Preparation of high-quality sequencing libraries is a costly and time-consuming component of metagenomic next generation sequencing (mNGS). While the overall cost of sequencing has dropped significantly over recent years, the reagents needed to prepare sequencing samples are likely to become the dominant expense in the process. Furthermore, libraries prepared by hand are subject to human variability and needless waste due to limitations of manual pipetting volumes. Reduction of reaction volumes, combined with sub-microliter automated dispensing of reagents without consumable pipette tips, has the potential to provide significant advantages. Here, we describe the integration of several instruments, including the Labcyte Echo 525 acoustic liquid handler and the iSeq and NovaSeq Illumina sequencing platforms, to miniaturize and automate mNGS library preparation, significantly reducing the cost and the time required to prepare samples. Through the use of External RNA Controls Consortium (ERCC) spike-in RNAs, we demonstrated the fidelity of the miniaturized preparation to be equivalent to full volume reactions. Furthermore, detection of viral and microbial species from cell culture and patient samples was also maintained in the miniaturized libraries. For 384-well mNGS library preparations, we achieved cost savings of over 80% in materials and reagents alone, and reduced preparation time by 90% compared to manual approaches, without compromising quality or representation within the library. [ABSTRACT FROM AUTHOR]

Background: Ocular infections remain a major cause of blindness and morbidity worldwide. While prognosis is dependent on the timing and accuracy of diagnosis, the etiology remains elusive in ~50% of presumed infectious uveitis cases. The objective of this study is to determine if unbiased metagenomic deep sequencing (MDS) can accurately detect pathogens in intraocular fluid samples of patients with uveitis. Methods: This is a proof-of-concept study, in which intraocular fluid samples were obtained from five subjects with known diagnoses, and one subject with bilateral chronic uveitis without a known etiology. Samples were subjected to MDS, and results were compared with those from conventional diagnostic tests. Pathogens were identified using a rapid computational pipeline to analyze the non-host sequences obtained from MDS. Results: Unbiased MDS of intraocular fluid produced results concordant with known diagnoses in subjects with (n = 4) and without (n = 1) uveitis. Samples positive for Cryptococcus neoformans, Toxoplasma gondii, and herpes simplex virus 1 as tested by a Clinical Laboratory Improvement Amendments-certified laboratory were correctly identified with MDS. Rubella virus was identified in one case of chronic bilateral idiopathic uveitis. The subject's strain was most closely related to a German rubella virus strain isolated in 1992, one year before he developed a fever and rash while living in Germany. The pattern and the number of viral identified mutations present in the patient's strain were consistent with long-term viral replication in the eye. Conclusions: MDS can identify fungi, parasites, and DNA and RNA viruses in minute volumes of intraocular fluid samples. The identification of chronic intraocular rubella virus infection highlights the eye's role as a long-term pathogen reservoir, which has implications for virus eradication and emerging global epidemics. [ABSTRACT FROM AUTHOR]

Background Mass distributions of oral azithromycin have long been used to eliminate trachoma, and they are now being proposed to reduce childhood mortality. The observed benefit appears to be augmented with each additional treatment, suggesting a possible community-level effect. Here, we assess whether 2 biannual mass treatments of preschool children affect the community's gut microbiome at 6 months after the last distribution. Methods In this cluster-randomized controlled trial, children aged 1–60 months in the Dossa region of Niger were randomized at the village level to receive a single dose of azithromycin or placebo every 6 months. Fecal samples were collected 6 months after the second treatment for metagenomic deep sequencing. The prespecified primary outcome was the Euclidean PERMANOVA of the gut microbiome, or effectively the distance between the genus-level centroid at the community level, with the secondary outcome being the Simpson's α diversity. Results In the azithromycin arm, the gut microbial structures were significantly different than in the placebo arm (Euclidean PERMANOVA, P <.001). Further, the diversity of the gut microbiome in the azithromycin arm was significantly lower than in the placebo arm (inverse Simpson's index, P =.005). Conclusions Two mass azithromycin administrations, 6 months apart, in preschool children led to long-term alterations of the gut microbiome structure and community diversity. Here, long-term microbial alterations in the community did not imply disease but were associated with an improvement in childhood mortality. Clinical Trials Registration NCT02048007. [ABSTRACT FROM AUTHOR]

BACKGROUND: Mass distribution of azithromycin to preschool children twice yearly for 2 years has been shown to reduce childhood mortality in sub-Saharan Africa but at the cost of amplifying macrolide resistance. The effects on the gut resistome, a reservoir of antimicrobial resistance genes in the body, of twice-yearly administration of azithromycin for a longer period are unclear. METHODS: We investigated the gut resistome of children after they received twice-yearly distributions of azithromycin for 4 years. In the Niger site of the MORDOR trial, we enrolled 30 villages in a concurrent trial in which they were randomly assigned to receive mass distribution of either azithromycin or placebo, offered to all children 1 to 59 months of age every 6 months for 4 years. Rectal swabs were collected at baseline, 36 months, and 48 months for analysis of the participants' gut resistome. The primary outcome was the ratio of macrolide-resistance determinants in the azithromycin group to those in the placebo group at 48 months. RESULTS: Over the entire 48-month period, the mean (±SD) coverage was 86.6±12% in the villages that received placebo and 83.2±16.4% in the villages that received azithromycin. A total of 3232 samples were collected during the entire trial period; of the samples obtained at the 48-month monitoring visit, 546 samples from 15 villages that received placebo and 504 from 14 villages that received azithromycin were analyzed. Determinants of macrolide resistance were higher in the azithromycin group than in the placebo group: 7.4 times as high (95% confidence interval [CI], 4.0 to 16.7) at 36 months and 7.5 times as high (95% CI, 3.8 to 23.1) at 48 months. Continued mass azithromycin distributions also selected for determinants of nonmacrolide resistance, including resistance to beta-lactam antibiotics, an antibiotic class prescribed frequently in this region of Africa. CONCLUSIONS: Among villages assigned to receive mass distributions of azithromycin or placebo twice yearly for 4 years, antibiotic resistance was more common in the villages that received azithromycin than in those that received placebo. This trial showed that mass azithromycin distributions may propagate antibiotic resistance. [ABSTRACT FROM AUTHOR]