Social interactions among animals mediate essential behaviours, including mating, nurturing, and defence 1,2 . The gut microbiota contribute to social activity in mice 3,4 , but the gut-brain connections that regulate this complex behaviour and its underlying neural basis are unclear 5,6 . Here we show that the microbiome modulates neuronal activity in specific brain regions of male mice to regulate canonical stress responses and social behaviours. Social deviation in germ-free and antibiotic-treated mice is associated with elevated levels of the stress hormone corticosterone, The Author(s), under exclusive licence to Springer Nature Limited 2021

A fundamental goal in microbiome studies is determining which microbes affect host physiology. Standard methods for determining changes in microbial taxa measure relative, rather than absolute abundances. Moreover, studies often analyze only stool, despite microbial diversity differing substantially among gastrointestinal (GI) locations. Here, we develop a quantitative framework to measure absolute abundances of individual bacterial taxa by combining the precision of digital PCR with the high-throughput nature of 16S rRNA gene amplicon sequencing. In a murine ketogenic-diet study, we compare microbial loads in lumenal and mucosal samples along the GI tract. Quantitative measurements of absolute (but not relative) abundances reveal decreases in total microbial loads on the ketogenic diet and enable us to determine the differential effects of diet on each taxon in stool and smallintestine mucosa samples. This rigorous quantitative microbial analysis framework, appropriate for diverse GI locations enables mapping microbial biogeography of the mammalian GI tract and more accurate analyses of changes in microbial taxa in microbiome studies.

Isothermal amplification assays, such as loopmediated isothermal amplification (LAMP), show great utility for the development of rapid diagnostics for infectious diseases because they have high sensitivity, pathogen-specificity and potential for implementation at the point of care. However, elimination of non-specific amplification remains a key challenge for the optimization of LAMP assays. Here, using chlamydia DNA as a clinically relevant target and high-throughput sequencing as an analytical tool, we investigate a potential mechanism of non-specific amplification. We then develop a real-time digital LAMP (dLAMP) with high-resolution melting temperature (HRM) analysis and use this single-molecule approach to analyze approximately 1.2 million amplification events. We show that single-molecule HRM provides insight into specific and non-specific amplification in LAMP that are difficult to deduce from bulk measurements. We use real-time dLAMP with HRM to evaluate differences between polymerase enzymes, the impact of assay parameters (e.g. time, rate or florescence intensity), and the effect background human DNA. By differentiating true and false positives, HRM enables determination of the optimal assay and analysis parameters that leads to the lowest limit of detection (LOD) in a digital isothermal amplification assay.

Early detection of SARS-CoV-2 infection is critical to reduce asymptomatic and pre-symptomatic transmission, curb the spread of variants, and maximize treatment efficacy. Low-analytical-sensitivity nasal-swab testing is commonly used for surveillance and symptomatic testing, but the ability of these tests to detect the earliest stages of infection has not been established. In this study, conducted between September 2020 and June 2021 in the greater Los Angeles County, California area, initially-SARS-CoV-2-negative household contacts of individuals diagnosed with COVID-19 prospectively self-collected paired anterior-nares nasal-swab and saliva samples twice daily for viral-load quantification by high-sensitivity RT-qPCR and digital-RT-PCR assays. We captured viral-load profiles from the incidence of infection for seven individuals and compared diagnostic sensitivities between respiratory sites. Among unvaccinated persons, testing saliva with a high-analytical-sensitivity assay detected infection up to 4.5 days before viral loads in nasal swabs reached concentrations detectable by low-analytical-sensitivity nasal-swab tests. For most participants, nasal swabs reached higher peak viral loads than saliva, but were undetectable or at lower loads during the first few days of infection. High-analytical-sensitivity saliva testing was most reliable for earliest detection. Our study illustrates the value of acquiring early (within hours after a negative high-sensitivity test) viral-load profiles to guide the appropriate analytical sensitivity and respiratory site for detecting earliest infections. Such data are challenging to acquire but critical to design optimal testing strategies with emerging variants in the current pandemic and to respond to future viral pandemics.

Antimicrobial-resistant Neisseria gonorrhoeae is an urgent public-health threat, with continued worldwide incidents of infection and rising resistance to antimicrobials. Traditional culture-based methods for antibiotic susceptibility testing are unacceptably slow (1–2 days), resulting in the use of broad-spectrum antibiotics and the further development and spread of resistance. Critically needed is a rapid antibiotic susceptibility test (AST) that can guide treatment at the point-of-care. Rapid phenotypic approaches using quantification of DNA have been demonstrated for fast-growing organisms (e.g. E. coli) but are challenging for slower-growing pathogens such as N. gonorrhoeae. Here, we investigate the potential of RNA signatures to provide phenotypic responses to antibiotics in N. gonorrhoeae that are faster and greater in magnitude compared with DNA. Using RNA sequencing, we identified antibiotic-responsive transcripts. Significant shifts (>4-fold change) in transcript levels occurred within 5 min of antibiotic exposure. We designed assays for responsive transcripts with the highest abundances and fold changes, and validated gene expression using digital PCR. Using the top two markers (porB and rpmB) we correctly determined the antibiotic susceptibility and resistance of 49 clinical isolates after 10 min exposure to ciprofloxacin. RNA signatures are therefore promising as an approach on which to build rapid AST devices for N. gonorrhoeae at the point-of-care, which is critical for disease management, surveillance, and antibiotic stewardship efforts.