Rapid expansion and extinction of antibiotic resistance mutations during treatment of acute bacterial respiratory infections.

Acute bacterial infections are often treated empirically, with the choice of antibiotic therapy updated during treatment. The effects of such rapid antibiotic switching on the evolution of antibiotic resistance in individual patients are poorly understood. Here we find that low-frequency antibiotic resistance mutations emerge, contract, and even go to extinction within days of changes in therapy. We analyzed Pseudomonas aeruginosa populations in sputum samples collected serially from 7 mechanically ventilated patients at the onset of respiratory infection. Combining short- and long-read sequencing and resistance phenotyping of 420 isolates revealed that while new infections are near-clonal, reflecting a recent colonization bottleneck, resistance mutations could emerge at low frequencies within days of therapy. We then measured the in vivo frequencies of select resistance mutations in intact sputum samples with resistance-targeted deep amplicon sequencing (RETRA-Seq), which revealed that rare resistance mutations not detected by clinically used culture-based methods can increase by nearly 40-fold over 5-12 days in response to antibiotic changes. Conversely, mutations conferring resistance to antibiotics not administered diminish and even go to extinction. Our results underscore how therapy choice shapes the dynamics of low-frequency resistance mutations at short time scales, and the findings provide a possibility for driving resistance mutations to extinction during early stages of infection by designing patient-specific antibiotic cycling strategies informed by deep genomic surveillance.

Evolution towards Virulence in a Burkholderia Two-Component System.

Bacteria in the Burkholderia cepacia complex (BCC) are significant pathogens for people with cystic fibrosis (CF) and are often extensively antibiotic resistant. Here, we assess the impacts of clinically observed mutations in fixL, which encodes the sensor histidine kinase FixL. FixL along with FixJ compose a two-component system that regulates multiple phenotypes. Mutations in fixL across two species, B. dolosa and B. multivorans, have shown evidence of positive selection during chronic lung infection in CF. Herein, we find that BCC carrying the conserved, ancestral fixL sequence have lower survival in macrophages and in murine pneumonia models than mutants carrying evolved fixL sequences associated with clinical decline in CF patients. In vitro phosphotransfer experiments found that one evolved FixL protein, W439S, has a reduced ability to autophosphorylate and phosphorylate FixJ, while LacZ reporter experiments demonstrate that B. dolosa carrying evolved fixL alleles has reduced fix pathway activity. Interestingly, B. dolosa carrying evolved fixL alleles was less fit in a soil assay than those strains carrying the ancestral allele, demonstrating that increased survival of these variants in macrophages and the murine lung comes at a potential expense in their environmental reservoir. Thus, modulation of the two-component system encoded by fixLJ by point mutations is one mechanism that allows BCC to adapt to the host infection environment. IMPORTANCE Infections caused by members of the Burkholderia cepacia complex (BCC) are a serious concern for patients with cystic fibrosis (CF) as these bacteria are often resistant to many antibiotics. During long-term infection of CF patients with BCC, mutations in genes encoding the FixLJ system often become prevalent, suggesting that these changes may benefit the bacteria during infection. The system encoded by fixLJ is involved in sensing oxygen and regulating many genes in response and is required for full virulence of the bacteria in a murine pneumonia model. Evolved fixL mutations seen later in infection improve bacterial persistence within macrophages and enhance infection within mice. However, these adaptations are short sighted because they reduce bacterial fitness within their natural habitat, soil.

Native American depopulation, reforestation, and fire regimes in the Southwest United States, 1492-1900 CE.

Native American populations declined between 1492 and 1900 CE, instigated by the European colonization of the Americas. However, the magnitude, tempo, and ecological effects of this depopulation remain the source of enduring debates. Recently, scholars have linked indigenous demographic decline, Neotropical reforestation, and shifting fire regimes to global changes in climate, atmosphere, and the Early Anthropocene hypothesis. In light of these studies, we assess these processes in conifer-dominated forests of the Southwest United States. We compare light detection and ranging data, archaeology, dendrochronology, and historical records from the Jemez Province of New Mexico to quantify population losses, establish dates of depopulation events, and determine the extent and timing of forest regrowth and fire regimes between 1492 and 1900. We present a new formula for the estimation of Pueblo population based on architectural remains and apply this formula to 18 archaeological sites in the Jemez Province. A dendrochronological study of remnant wood establishes dates of terminal occupation at these sites. By combining our results with historical records, we report a model of pre- and post-Columbian population dynamics in the Jemez Province. Our results indicate that the indigenous population of the Jemez Province declined by 87% following European colonization but that this reduction occurred nearly a century after initial contact. Depopulation also triggered an increase in the frequency of extensive surface fires between 1640 and 1900. Ultimately, this study illustrates the quality of integrated archaeological and paleoecological data needed to assess the links between Native American population decline and ecological change after European contact.