A swapped genetic code prevents viral infections and gene transfer.

Engineering the genetic code of an organism has been proposed to provide a firewall from natural ecosystems by preventing viral infections and gene transfer1-6. However, numerous viruses and mobile genetic elements encode parts of the translational apparatus7-9, potentially rendering a genetic-code-based firewall ineffective. Here we show that such mobile transfer RNAs (tRNAs) enable gene transfer and allow viral replication in Escherichia coli despite the genome-wide removal of 3 of the 64 codons and the previously essential cognate tRNA and release factor genes. We then establish a genetic firewall by discovering viral tRNAs that provide exceptionally efficient codon reassignment allowing us to develop cells bearing an amino acid-swapped genetic code that reassigns two of the six serine codons to leucine during translation. This amino acid-swapped genetic code renders cells resistant to viral infections by mistranslating viral proteomes and prevents the escape of synthetic genetic information by engineered reliance on serine codons to produce leucine-requiring proteins. As these cells may have a selective advantage over wild organisms due to virus resistance, we also repurpose a third codon to biocontain this virus-resistant host through dependence on an amino acid not found in nature10. Our results may provide the basis for a general strategy to make any organism safely resistant to all natural viruses and prevent genetic information flow into and out of genetically modified organisms.

Genomic Analyses of Longitudinal Mycobacterium abscessus Isolates in a Multicenter Cohort Reveal Parallel Signatures of In-Host Adaptation.

BACKGROUND: Nontuberculous mycobacteria (NTM) are ubiquitous in the environment and an increasingly frequent cause of opportunistic infections. Mycobacterium abscessus complex (MABC) is one of the major NTM lung pathogens that disproportionately colonize and infect the lungs of individuals with cystic fibrosis (CF). MABC infection can persist for years, and antimicrobial treatment is frequently ineffective. METHODS: We sequenced the genomes of 175 isolates longitudinally collected from 30 patients with MABC lung infection. We contextualized our cohort amidst the broader MABC phylogeny and investigated genes undergoing parallel adaptation across patients. Finally, we tested the phenotypic consequences of parallel mutations by conducting antimicrobial resistance and mercury-resistance assays. RESULTS: We identified highly related isolate pairs across hospital centers with low likelihood of transmission. We further annotated nonrandom parallel mutations in 22 genes and demonstrated altered macrolide susceptibility co-occurring with a nonsynonymous whiB1 mutation. Finally, we highlighted a 23-kb mercury-resistance plasmid whose loss during chronic infection conferred phenotypic susceptibility to organic and nonorganic mercury compounds. CONCLUSIONS: We characterized parallel genomic processes through which MABC is adapting to promote survival within the host. The within-lineage polymorphisms we observed have phenotypic effects, potentially benefiting fitness in the host at the putative detriment of environmental survival.

Necrotizing Enterocolitis and the Microbiome: Current Status and Future Directions.

Decades of research have failed to define the pathophysiology of necrotizing enterocolitis (NEC), a devastating pediatric gastrointestinal disorder of preterm infants. However, evidence suggests that host-microbiota interactions, in which microbial dysbiosis is followed by loss of barrier integrity, inflammation, and necrosis, are central to NEC development. Thus, greater knowledge of the preterm infant microbiome could accelerate attempts to diagnose, treat, and prevent NEC. In this article, we summarize clinical characteristics of and risk factors for NEC, the structure of the pre-event NEC microbiome, how this community interfaces with host immunology, and microbiome-based approaches that might prevent or lessen the severity of NEC in this very vulnerable population.

Acoustic tracking of fin whales: Habitat use and movement patterns within a Canadian Pacific fjord system.

Fin whale 20 Hz calls were detected, localized, and tracked using a 10 km aperture network of three acoustic receivers deployed for 11 months in a Pacific Canadian fjord system. The area has been historically important for fin whales and is located along a route that tankers will begin using in 2024. A total of 6712 calls were localized, and trajectories were fitted for 55 acoustic tracks. Fin whale tracks occurred throughout the monitoring site. Call activity peaked in September and was low during winter months. Swimming characteristics varied significantly between day- and nighttime: at night, whales swam faster (7.1 vs 4.0 km/h median, +75.2%), which resulted in longer (+34.7%), less predictable (-70.6%) tracks as compared to daylight hours. Call frequencies varied between 16 and 32 Hz. Beside stereotypical song frequencies, fin whales also used irregular frequency components, which contributed the majority of calls in the summer but did not occur in the winter. The results suggest that the area is primarily used as a summer feeding ground, where fin whales follow a diel behavioral cycle. The observed activity patterns will aid in the assessment of strike risk and harassment mitigation and provide a baseline to document behavioral change.

Detection and classification of narrow-band high frequency echolocation clicks from drifting recorders.

In the California Current off the United States West Coast, there are three offshore cetacean species that produce narrow-band high frequency (NBHF) echolocation pulses: Dall's porpoise (Phocoenoides dalli) and two species of Kogia. NBHF pulses exist in a highly specialized acoustic niche thought to be outside the hearing range of killer whales and other potential mammal-eating odontocetes. Very little is known about the dwarf and pygmy sperm whales (K. sima and K. breviceps), including their NBHF pulse characteristics. This paper presents a multivariate clustering method using data from unmanned drifting acoustic recorders and visually verified porpoise recordings to discriminate between probable porpoise and Kogia clicks. Using density clustering, this study finds three distinct clusters whose geographic distributions are consistent with the known habitat range for Kogia and Dall's porpoise. A Random Forest classification model correctly assigned 97% of the clicks to their cluster. Visually verified Dall's porpoise clicks from towed hydrophones were strongly associated with one of the clusters, while a second cluster tended to be outside the geographic range of Dall's porpoise and unlike the Dall's porpoise cluster. These clicks, presumed to be made by Kogia, exhibited greater spectral variance than previous Kogia echolocation studies. It is possible that the structure of Kogia NBHF pulses may not be as stereotypical as previously described.

Automated localization of whales in coastal fjords.

Localization and tracking of vocalizing marine mammals are powerful tools for understanding and mitigating the impacts of anthropogenic stressors such as vessel noise on habitat use of cetaceans. A large-aperture hydrophone network has been installed in the Kitimat Fjord System, an ecologically, culturally, and economically valued marine environment in northern British Columbia, Canada. This network consists of four synchronized bottom-mounted hydrophones that permanently record and radio-transmit data to a land-based laboratory. An automated system has been developed which includes routines to localize transient bio-acoustic signals from three or more streaming hydrophones in near real-time. These routines comprise the correlation of hydrophone signals, the construction of a time lag model, and signal localization and error estimation from a spatial likelihood surface. The localization method was tested experimentally and subsequently applied to vocalizations from humpback whales, fin whales, and killer whales. Refractive and reflective sound propagation effects in the confined fjords are assessed using ray tracing propagation models. Automated localization results are compared to ground-truth data and shown to provide good accuracy.

A century of phage research: bacteriophages and the shaping of modern biology.

2015 marks the centennial of the discovery of bacteriophages, viruses that infect bacteria. Phages have been central to some of biology's most meaningful advances over the past hundred years (shown here); they greatly influence the workings of the biosphere, and are poised to play expanded roles in biomedicine, biotechnology, and ecology.

Collecting baleen whale blow samples by drone: A minimally intrusive tool for conservation genetics.

In coastal British Columbia, Canada, marine megafauna such as humpback whales (Megaptera novaeangliae) and fin whales (Balaenoptera physalus velifera) have been subject to a history of exploitation and near extirpation. While their populations have been in recovery, significant threats are posed to these vulnerable species by proposed natural resource ventures in this region, in addition to the compounding effects of anthropogenic climate change. Genetic tools play a vital role in informing conservation efforts, but the associated collection of tissue biopsy samples can be challenging for the investigators and disruptive to the ongoing behaviour of the targeted whales. Here, we evaluate a minimally intrusive approach based on collecting exhaled breath condensate, or respiratory 'blow' samples, from baleen whales using an unoccupied aerial system (UAS), within Gitga'at First Nation territory for conservation genetics. Minimal behavioural responses to the sampling technique were observed, with no response detected 87% of the time (of 112 UAS deployments). DNA from whale blow (n = 88 samples) was extracted, and DNA profiles consisting of 10 nuclear microsatellite loci, sex identification and mitochondrial (mt) DNA haplotypes were constructed. An average of 7.5 microsatellite loci per individual were successfully genotyped. The success rates for mtDNA and sex assignment were 80% and 89% respectively. Thus, this minimally intrusive sampling method can be used to describe genetic diversity and generate genetic profiles for individual identification. The results of this research demonstrate the potential of UAS-collected whale blow for conservation genetics from a remote location.

Correction: Fin whales of the Great Bear Rainforest: Balaenoptera physalus velifera in a Canadian Pacific fjord system.

[This corrects the article DOI: 10.1371/journal.pone.0256815.].

Clinical sequelae of gut microbiome development and disruption in hospitalized preterm infants.

Aberrant preterm infant gut microbiota assembly predisposes to early-life disorders and persistent health problems. Here, we characterize gut microbiome dynamics over the first 3 months of life in 236 preterm infants hospitalized in three neonatal intensive care units using shotgun metagenomics of 2,512 stools and metatranscriptomics of 1,381 stools. Strain tracking, taxonomic and functional profiling, and comprehensive clinical metadata identify Enterobacteriaceae, enterococci, and staphylococci as primarily exploiting available niches to populate the gut microbiome. Clostridioides difficile lineages persist between individuals in single centers, and Staphylococcus epidermidis lineages persist within and, unexpectedly, between centers. Collectively, antibiotic and non-antibiotic medications influence gut microbiome composition to greater extents than maternal or baseline variables. Finally, we identify a persistent low-diversity gut microbiome in neonates who develop necrotizing enterocolitis after day of life 40. Overall, we comprehensively describe gut microbiome dynamics in response to medical interventions in preterm, hospitalized neonates.

High-throughput digital cough recording on a university campus: A SARS-CoV-2-negative curated open database and operational template for acoustic screening of respiratory diseases.

Objective: Respiratory illnesses have information-rich acoustic biomarkers, such as cough, that can potentially play an important role in screening populations for disease risk. To realize that potential, datasets of paired acoustic-clinical samples are needed for the development and validation of acoustic screening models, and protocols for collecting acoustic samples must be efficient and safe. We collected cough acoustic signatures at a high-throughput SARS-CoV-2 testing site on a college campus. Here, we share logistical details and the dataset of acoustic cough signatures paired with the gold standard in SARS-CoV-2 testing of SARS-CoV-2 genomic sequences using qRT-PCR. Methods: Cough recordings were collected in winter-spring 2021 at a rural residential college (Sewanee, TN, USA), where approximately 2000 students were tested for SARS-CoV-2 on a weekly basis. Cough collection was managed by student volunteers using custom software. Results: 4302 coughs were recorded from 960 participants over 11 weeks. All coughs were COVID-19 negative. Approximately 30 s were required to check-in a participant and collect their cough. Conclusion: The value of acoustic screening tools depends upon our ability to develop and implement them reliably and quickly. For that to happen, high-quality datasets and logistical insights must be collected and shared on an ongoing basis.

Longitudinal passive cough monitoring and its implications for detecting changes in clinical status.

Research question: What is the impact of the duration of cough monitoring on its accuracy in detecting changes in the cough frequency? Materials and methods: This is a statistical analysis of a prospective cohort study. Participants were recruited in the city of Pamplona (Northern Spain), and their cough frequency was passively monitored using smartphone-based acoustic artificial intelligence software. Differences in cough frequency were compared using a one-tailed Mann-Whitney U test and a randomisation routine to simulate 24-h monitoring. Results: 616 participants were monitored for an aggregated duration of over 9 person-years and registered 62 325 coughs. This empiric analysis found that an individual's cough patterns are stochastic, following a binomial distribution. When compared to continuous monitoring, limiting observation to 24 h can lead to inaccurate estimates of change in cough frequency, particularly in persons with low or small changes in rate. Interpretation: Detecting changes in an individual's rate of coughing is complicated by significant stochastic variability within and between days. Assessing change based solely on intermittent sampling, including 24-h, can be misleading. This is particularly problematic in detecting small changes in individuals who have a low rate and/or high variance in cough pattern.

Acoustic surveillance of cough for detecting respiratory disease using artificial intelligence.

Research question: Can smartphones be used to detect individual and population-level changes in cough frequency that correlate with the incidence of coronavirus disease 2019 (COVID-19) and other respiratory infections? Methods: This was a prospective cohort study carried out in Pamplona (Spain) between 2020 and 2021 using artificial intelligence cough detection software. Changes in cough frequency around the time of medical consultation were evaluated using a randomisation routine; significance was tested by comparing the distribution of cough frequencies to that obtained from a model of no difference. The correlation between changes of cough frequency and COVID-19 incidence was studied using an autoregressive moving average analysis, and its strength determined by calculating its autocorrelation function (ACF). Predictors for the regular use of the system were studied using a linear regression. Overall user experience was evaluated using a satisfaction questionnaire and through focused group discussions. Results: We followed-up 616 participants and collected >62 000 coughs. Coughs per hour surged around the time cohort subjects sought medical care (difference +0.77 coughs·h-1; p=0.00001). There was a weak temporal correlation between aggregated coughs and the incidence of COVID-19 in the local population (ACF 0.43). Technical issues affected uptake and regular use of the system. Interpretation: Artificial intelligence systems can detect changes in cough frequency that temporarily correlate with the onset of clinical disease at the individual level. A clearer correlation with population-level COVID-19 incidence, or other respiratory conditions, could be achieved with better penetration and compliance with cough monitoring.

Clinical performance of scientifically designed, hot isostatic-pressed (HIP'd) zirconia cores in a bilayered all-ceramic system.

PURPOSE: The success of bilayered all-ceramic restorations is dependent upon the combination and contributions of the three principal components of these restorations: core material, core design, and core-veneer interface. The purpose of this paper is to describe the fabrication and clinical survival of optimized ceramic restorations having an explicit, scientifically designed core, machined from HIP'd isotropic zirconia and veneered using a specific protocol with thermally compatible porcelain. MATERIALS AND METHODS: Using a consistent clinical and laboratory protocol in a multicenter setting, 3,192 bilayered single and 797 bilayered splinted units were fabricated and placed on teeth and implant abutments in 1,007 patients. Approximately 61.7% (n = 2,462) were posterior restorations and 38.3% (n = 1,527) were anterior. Of the total, approximately 5.7% (n = 227) were placed on implant abutments. Survival of the restorations was determined with the Kaplan-Meier (KM) method by tooth number. RESULTS: For the 3,989 units placed, 9 failures were recorded. The KM survival of most zirconia restorations, when segregated by tooth number, was 100%. Exceptions were the 9 failed units, with a KM survival between 88% and 99% for those restorations. Six restorations failed within the first year of service, including three failed cores. Examination of those restorations revealed failure was related to initial design, quality assessment, or fabrication inconsistencies. CONCLUSION: The incorporation of a reinforcing ring beam onto an anatomically shaped core made from end-state HIP'd zirconia, in partnership with a thermally compatible veneering porcelain and a specific application protocol, resulted in extremely high survival rates for both anterior and posterior all-ceramic restorations after medium-term clinical use. These results equal or surpass the equivalent-term success rates of porcelain-fused-to-metal restorations.

Social survival: Humpback whales (Megaptera novaeangliae) use social structure to partition ecological niches within proposed critical habitat.

Animal culture and social bonds are relevant to wildlife conservation because they influence patterns of geography, behavior, and strategies of survival. Numerous examples of socially-driven habitat partitioning and ecological-niche specialization can be found among vertebrates, including toothed whales. But such social-ecological dynamics, described here as 'social niche partitioning', are not known among baleen whales, whose societies-particularly on foraging grounds-are largely perceived as unstructured and incidental to matters of habitat use and conservation. However, through 16 years of behavioral observations and photo-identifications of humpback whales (Megaptera novaeangliae) feeding within a fjord system in the Canadian Pacific (primarily within Gitga'at First Nation waters), we have documented long-term pair bonds (up to 12 years) as well as a complex societal structure, which corresponds closely to persistent patterns in feeding strategy, long-term site fidelity (extended occupancy and annual rate of return up to 75%), specific geographic preferences within the fjord system, and other forms of habitat use. Randomization tests of network congruency and clustering algorithms were used to test for overlap in patterns of social structure and habitat use, which confirmed the occurrence of social niche partitioning on the feeding grounds of this baleen whale species. In addition, we document the extensive practice of group bubble net feeding in Pacific Canada. This coordinated feeding behavior was found to strongly mediate the social structure and habitat use within this humpback whale society. Additionally, during our 2004-2019 study, we observed a shift in social network structure in 2010-2012, which corresponded with environmental and demographic shifts including a sudden decline in the population's calving rate. Our findings indicate that the social lives of humpback whales, and perhaps baleen whales generally, are more complex than previously supposed and should be a primary consideration in the assessment of potential impacts to important habitat.

Comparative Genomics of Mycobacterium avium Complex Reveals Signatures of Environment-Specific Adaptation and Community Acquisition.

Nontuberculous mycobacteria, including those in the Mycobacterium avium complex (MAC), constitute an increasingly urgent threat to global public health. Ubiquitous in soil and water worldwide, MAC members cause a diverse array of infections in humans and animals that are often multidrug resistant, intractable, and deadly. MAC lung disease is of particular concern and is now more prevalent than tuberculosis in many countries, including the United States. Although the clinical importance of these microorganisms continues to expand, our understanding of their genomic diversity is limited, hampering basic and translational studies alike. Here, we leveraged a unique collection of genomes to characterize MAC population structure, gene content, and within-host strain dynamics in unprecedented detail. We found that different MAC species encode distinct suites of biomedically relevant genes, including antibiotic resistance genes and virulence factors, which may influence their distinct clinical manifestations. We observed that M. avium isolates from different sources-human pulmonary infections, human disseminated infections, animals, and natural environments-are readily distinguished by their core and accessory genomes, by their patterns of horizontal gene transfer, and by numerous specific genes, including virulence factors. We identified highly similar MAC strains from distinct patients within and across two geographically distinct clinical cohorts, providing important insights into the reservoirs which seed community acquisition. We also discovered a novel MAC genomospecies in one of these cohorts. Collectively, our results provide key genomic context for these emerging pathogens and will facilitate future exploration of MAC ecology, evolution, and pathogenesis. IMPORTANCE Members of the Mycobacterium avium complex (MAC), a group of mycobacteria encompassing M. avium and its closest relatives, are omnipresent in natural environments and emerging pathogens of humans and animals. MAC infections are difficult to treat, sometimes fatal, and increasingly common. Here, we used comparative genomics to illuminate key aspects of MAC biology. We found that different MAC species and M. avium isolates from different sources encode distinct suites of clinically relevant genes, including those for virulence and antibiotic resistance. We identified highly similar MAC strains in patients from different states and decades, suggesting community acquisition from dispersed and stable reservoirs, and we discovered a novel MAC species. Our work provides valuable insight into the genomic features underlying these versatile pathogens.

Fin whales of the Great Bear Rainforest: Balaenoptera physalus velifera in a Canadian Pacific fjord system.

Fin whales (Balaenoptera physalus) are widely considered an offshore and oceanic species, but certain populations also use coastal areas and semi-enclosed seas. Based upon fifteen years of study, we report that Canadian Pacific fin whales (B. p. velifera) have returned to the Kitimat Fjord System (KFS) in the Great Bear Rainforest, and have established a seasonally resident population in its intracoastal waters. This is the only fjord system along this coast or elsewhere in which fin whales are known to occur regularly with strong site fidelity. The KFS was also the only Canadian Pacific fjord system in which fin whales were commonly found and killed during commercial whaling, pointing to its long-term importance. Traditional knowledge, whaling records, and citizen science databases suggest that fin whales were extirpated from this area prior to their return in 2005-2006. Visual surveys and mark-recapture analysis documented their repopulation of the area, with 100-120 whales using the fjord system in recent years, as well as the establishment of a seasonally resident population with annual return rates higher than 70%. Line transect surveys identified the central and outer channels of the KFS as the primary fin whale habitat, with the greatest densities occurring in Squally Channel and Caamaño Sound. Fin whales were observed in the KFS in most months of the year. Vessel- and shore-based surveys (27,311 km and 6,572 hours of effort, respectively) indicated regular fin whale presence (2,542 detections), including mother-calf pairs, from June to October and peak abundance in late August-early September. Seasonal patterns were variable year-to-year, and several lines of evidence indicated that fin whales arrived and departed from the KFS repeatedly throughout the summer and fall. Additionally, we report on the population's social network and morphometrics. These findings offer insights into the dynamics of population recovery in an area where several marine shipping projects are proposed. The fin whales of the Great Bear Rainforest represent a rare exception to general patterns in this species' natural history, and we highlight the importance of their conservation.

Microbiome Restoration by RBX2660 Does Not Preclude Recurrence of Multidrug-Resistant Urinary Tract Infection Following Subsequent Antibiotic Exposure: A Case Report.

A 62-year-old woman received RBX2660, an investigational microbiome restoration therapeutic, for recurrent multidrug-resistant (MDR) urinary tract infection (UTI). RBX2660 increased gut microbiome diversity but did not eliminate uropathogen carriage, and MDR UTI recurred after subsequent antibiotic exposure. Thus, restoration of microbiome diversity does not preclude disease recurrence by residual MDR pathogens.